I"!- m LIBRAA Y I 3 1293 00659 1774 Michigan State University This is to certify that the thesis entitled Control of Hydrogen Sulfide Odors Fran Anaerobic lagoons by Purple Sulfur Bacteria presented by Theodorus Johannes Maria van Iotringen has been accepted towards fulfillment of the requirements for H1. D. degree in Agricultural Engineering Major pr fessor DateM 0-7 639 @1151 U 5 :993 ‘5- CONTROL OF HYDROGEN SULFIDB ODORS FROM ANAEROBIC LAGOONS BY PURPLE SULFUR BACTERIA BY Theodorus Johannes Maria van Lotringen A DISSERTATION Submitted to Michigan State University in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY Department of Agricultural Engineering 1978 PLEASE NOTE: Some pages contain colored illustrations which will not reproduce well. Filmed as received. UNIVERSITY MICROFILMS INTERNATIONAL ABSTRACT CONTROL OF HYDROGEN SULFIDE ODORS FROM ANAEROBIC LAGOONS BY PURPLE SULFUR BACTERIA BY Theodorus Johannes Maria van Lotringen Purple sulfur bacteria have been found to reduce the odor levels of anaerobic lagoons. A serious problem is getting a culture of purple sulfur bacteria to survive a cold winter season. This thesis puts forward a quantitative analysis of the purple sulfur bacterial processes in a lagoon environment. A model based largely on literature expresses these processes in mathematical equations. Also presented are experiments to support the model where critical values were unavailable. The processes considered in the model are the major components of the sulfur cycle, since hydrogen sulfide is believed to be an important contributor to lagoon odor. The physical environment (with special emphasis on heat balance) is modeled in order to obtain better insight into its influence on odor production. In support of the heat balance model a temperature history was recorded during the spring warm-up of an Theodorus Johannes Maria van Lotringen anaerobic swine waste lagoon. Evidence suggests that a froth-type scum cover increases the heat loss of such a lagoon system, tending to prolong the warm-up period. In support of the model of the sulfur cycle, the rate at which hydrogen sulfide is oxidized and the rate at which sulfate is formed were measured under various light- and temperature conditions. At low light intensities the rate of oxidation of hydrogen sulfide seems to be independent of the temperature. The work with the anaerobic swine waste lagoons at MSU has led to new insights in design and management of such lagoons. A system involving two lagoons has been shown effective in reducing the time during which odors can be pro- duced; the basic features of the system include a strategy to preserve a population of purple sulfur bacteria during the cold Michigan winter. ACKNOWLEDGMENTS I am grateful to Dr. J. B. Gerrish, whose advice and guidance profoundly affected my development as a graduate student. I thank Drs. D. E. Linvill, T. L. London, E. R. Miller, R. L. Uffen and M. Yokoyama for their advice and suggestions. I thank V. E. Shull for his assistance with the electron microprobe, P. K. Ku for his assistance with the iron analysis, and P. Colwell for the spectral analysis of the light source. I thank J. G. Funkhouser for the use of the double beam spectroPhotometer. Special thanks is given to Jane Repko for her assistance in the work, for the many helpful suggestions and for her encouragement. I am grateful to Fumiko Mikami for her assistance in the preparation of the thesis and her support for all the work. I am also grateful for financial assistance from the MSU Agricultural Experiment Station and from the National Pork Producers Council. ii LIST OF TABLES LIST OF FIGURES LIST OF SYMBOLS INTRODUCTION . TABLE OF CONTENTS PART I. REVIEW OF LITERATURE CHAPTER CHAPTER CHAPTER CHAPTER 1. O O ubUONI-J 3 3. 3. 3 4 LAGOONS . . . . . . . . . . . . Introduction . . . . . . . . . Aerobic lagoons . . . . . . . Facultative lagoons . . . . . Anaerobic lagoons . . . . . . ANAEROBIC TREATMENT . . . . . . Introduction . . . . . . . . . Anaerobic digestion processes The influence of the physical environment . . . . . . . The influence of the chemical environment . . . . . . . Models and simulation of the anaerobic treatment . . . . PURPLE SULFUR BACTERIA . . . . Introduction . . . . . . . . . Classification . . . . . . . . Metabolism . . . . . . . Interactions with the environment SWINE WASTE MANAGEMENT AT MSU . iii Page vi viii xi ONU'lbbJ u (D 14 17 22 26 26 27 32 36 41 Page PART II. THEORY AND MODEL DEVELOPMENT CHAPTER 5. HEAT BALANCE . . . . . . . . . . . . . . 53 1 Introduction . . . . . . . . . . . 53 2 Transfer of heat by radiation . . . . . 55 3 Transfer of heat by evaporation or condensation . . . . . . . . . . . . 60 4 Transfer of sensible heat . . . . . . . 67 5 The distribution of heat in the lagoon . . . . . . . . . . . . . . . 68 5.6 Scum formation . . . . . . . . . . . . 69 CHAPTER 6. THE SULFUR CYCLE AND KINETICS . . . . . 74 6.1 Introduction . . . . . . . . . . . . 74 6.2 Description and mathematical formulation of the lagoon . . . . . . 76 6.3 Description and mathematical formula- tion of gas bubbles rising in the lagoon liquid . . . . . . . . . . . . 79 6.4 Transfer of hydrogen sulfide . . . . . 84 6.5 Microbial conversion processes . . . . 87 6.5.1 Sulfide balance . . . . . . . . 87 6.5.2 Sulfate balance . . . . . . . 94 6.5.3 Sulfur balance . . . . . . . . . 95 6.5.4 Balance of purple sulfur bacteria . . . . . . . . . . . 95 6.5.5 Balance of desulfurizing bacteria . . . . . . . . . . . 96 6.6 The influence of pH . . . . . . . . . . 96 6.7 The influence of temperature . . . . . 97 6.8 Changes in the lagoon volume . . . . . 98 6.9 The total model . . . . . . . . . . . . 99 PART III. EXPERIMENTS CHAPTER . HEAT BALANCE . . . . . . . . . . . . . . 102 7 7.1 Introduction . . . . . . . . . . . 102 7.2 The shape of the lagoons . . . . . . . 103 7.3 Temperature distribution . . . . . . . 108 8 CHAPTER 0 KINETICS O O O O O O O O O O O O O O O O 118 Introduction . . . . . . . . . . . . . 118 8.1 8.2 Experimental procedures . . . . . . . . 113 8.3 Results and discussion . . . . . . . . 123 iv PART IV. CHAPTER 9. CONCLUSIONS APPENDICES Appendix A. Constants and Parameter Values B. Analytical Methods C. Environmental Data BIBLIOGRAPHY MANAGEMENT IMPLICATIONS DISCUSSION Page 148 153 155 162 166 169 Table 3.1 5.1 5.2 LIST OF TABLES CHAPTER 3 Carotenoid Composition (as g of total caro- tenoids) of Three Purple Sulfur Bacteria, the Absorption Maxima of These Carotenoids and the Extinction Coefficients of the Middle Main Maxima . . . . . . . . . . . . . . . . CHAPTER 4 Swine Feed Mix . . . . . . . . . . . . . . . . Nutrient Composition of Dried Swine Feces (DSF) o o o o o o o o o o o o o o o o o o 0 Amino Acid Composition of Dried Swine Feces (DSF) o o o o o o o o o o o o o o o o o o o Swine Input-output Summary . . . . . . . . . Pollutional Characteristics of Swine Waste . . Mineral Analysis--(Feed, Feces, Urine) SWine O O O O O O O O O O O O O O O O O O O O Lagoon Analysis in 1974 . . . . . . . . . . . . CHAPTER 5 Parameter Values for the Equation Based on Vapor Pressure Deficit and Wind Velocity . . Volatile Fatty Acids in the Lagoons on April 30, 1977 . . . . . . . . . . . . . . . vi Page 31 42 43 43 45 46 47 51 62 73 Table A.1 A.2 A.3 A.4 A.5 CHAPTER 7 Depth Profile of the West Lagoon CHAPTER 8 Oxidation of Hydrogen Sulfide at Light Intensities . . . . . . Oxidation of Hydrogen Sulfide at Light Intensities . . . . . . Oxidation of Hydrogen Sulfide at Light Intensities . . . . . . Oxidation of Hydrogen Sulfide at 540 1x . . . . . . . . . . . . Oxidation of Hydrogen Sulfide at 10°C and Four 20°C and Four 25°C and Four 0 O I O O 25°C and 25°C and 540 1x .143 with Increased Starting Concentrations . . . Oxidation of Hydrogen Sulfide at Four Temperatures . . . . . . APPENDIX A Conversion of Units (Weast, 1976) Physical Constants (Weast, 1976) Waste Characteristics . . . . . Lagoon Characteristics . . . . . Microbial Characteristics . . . APPENDIX C Solar Radiation Data . . . . . . Local Climatological Data . . . vii 540 1x and Page .106 .125 .126 .127 137 . 144 155 156 158 159 160 166 167 LIST OF FIGURES Figure Page CHAPTER 4 4.1 MSU Swine Farm Lagoons . . . . . . . . . . . . . 48 4.2 The Color of the West Lagoon in the Summer of 1977 . . . . . . . . . . . . . . . . . . . . . 52 CHAPTER 5 5.1 Direct Solar Radiation as a Function of the Time of the Year at 45°N Latitude . . . . . . 57 5.2 Scum Formation in Spring . . . . . . . . . . . . 71 CHAPTER 6 6.1 The Trapezoidal Shaped Lagoon . . . . . . . . . 77 6.2 A Spherical Cap . . . . . . . . . . . . . . . . 80 6.3 Lineweaver-Burk Plot for Light-limited Growth ... . . . . . . . . . . . . . . . . . . 93 6.4 Model of the Sulfur Cycle . . . . . . . . . . . 100 6.5 Flowchart of the Main Loop . . . . . . . . . . . 101 CHAPTER 7 7.1 Water Depth of the West Lagoon . . . . . . . . . 104 7.2 Sediment Depth of the West Lagoon . . . . . . . 107 7.3 Installation of Temperature Measurement system 0 O O O O O O O O O O O O O O O O O O O 109 7.4 Temperature Measurement System in Operation . . 109 viii Figure 7.5 7.10 8.1 8.2 8.6 8.7 8.8 Page Temperatures in the East Lagoon from May 1, 9 P.M. to May 4, 8 A.M. . . . . . . . . . . . 110 Temperatures in the East Lagoon from May 18, 9 A.M. to May 20, 11 P.M., with a Three Hour Interruption at the Thirtieth Hour . . . . . lll Temperatures in the West Lagoon from May 1, 9 P.M. till May 4, 8 A.M. . . . . . . . . . . 112 Temperatures in the West Lagoon from May 18, 9 A.M. till May 20, ll P.M. with a Three Hour Interruption at the Thirtieth Hour . . . 113 Heat Content of the Two Lagoons from May 1, 9 P.M. to May 4, 8 A.M. . . . . . . . . . . . 116 Heat Content of the Two Lagoons from May 18, 9 A.M. to May 20, 11 P.M. with an Interruption at the Thirtieth Hour . . . . . . . . . . . . 117 CHAPTER 8 Spectral Distribution of the Incandescent Light Source . . . . . . . . . . . . . . . . 120 Oxidation of Hydrogen Sulfide at 10°C and Four Light Intensities . . . . . . . . . . . 128 Formation of Sulfate at 10°C and Four Light IntenSitieS O I O O 0 O O O O I O I O O O O O 129 Oxidation of Hydrogen Sulfide at 20°C and Four Light Intensities . . . . . . . . . . . . . . 130 Formation of Sulfate at 20°C and Four Light IntenSitieS O O O O O O O O O O O O O O O O O 1'31 Oxidation of Sulfide at 25°C and Four Light Intensities . . . . . . . . . . . . . . . . . 132 Formation of Sulfate at 25°C and Four Light IntenSities I O O O O I O O O O O O O O O O O 133 Concentration of Hydrogen Sulfide in the West Lagoon During the Fall of 1977 (measured at 5-6 P.M.) o o o o o o o o o o o o o o 9, o o o 134 ix 9.1 Page Oxidation of Hydrogen Sulfide at 25°C and 540 1x . . . . . . . . . . . . . . . . . . . 138 Oxidation of Hydrogen Sulfide at 25°C and 540 1x (plotted by computer subroutine) . . 139 Computer Simulation of the Oxidation of Hydro- gen Sulfide at 25°C and 540 lx . . . . . . . 141 Oxidation of Hydrogen Sulfide at 25°C and 540 1x with Increased Starting Concentrations. . 145 Formation of Sulfate at 25°C and 540 1x with Increased Starting Concentrations of Hydrogen Sulfide . . . . . . . . . . . . . . 146 Oxidation of Hydroqen Sulfide at 540 1x and Four Temperatures . . . . . . . . . . . . . 147 CHAPTER 9 Pumping the East Lagoon in the Fall 1977 . . . 151 Symbol U LIST OF SYMBOLS Description Surface area Surface area of a bubble Cross-sectional area of the lagoon at height h above the bottom Atomic weight of sulfur Constant Fraction of the sky covered by clouds Constant Drag coefficient Constant Velocity of light in vacuo Diffusivity Diffusivity of hydroqen sulfide in water Constant Equivalent diameter of a bubble The actual diameter of a bubble Radiant energy at a wavelength Efitvos number Rate of feces production per unit weight of animal Froude number Gravitational acceleration xi m/s m /s m /s W/m2 m m3/kg 5 m/s Symbol K. K" Description The total water depth in a lagoon Variable height measured from the lagoon bottom Planck's constant Height of compensation point Rise of a bubble Incident light intensity Compensation point for purple sulfur bacteria Light intensity at height h above the bottom of a lagoon with total height H Saturation constant for the reduction of sulfate Saturation constant for the oxidation of hydrogen sulfide Saturation constant for the oxidation of sulfur Saturation constant for the reduction of sulfur Saturation constant of light Henry's constant for carbon dioxide First dissociation constant for H28 Henry's constant for hydrogen sulfide The inhibition constant for hydrogen sulfide First dissociation constant for H28 Second dissociation constant for H28 Boltzmann constant Mass transfer coefficient for H28 at the bubble surface xii Units 1x 1x 1x 3 mole/m Pa mole/m3 mole/m3 Pa mole/m3 mole/m3 mole/m3 J/K m/s Symbol Description Factor depending on the cloud type Mass transfer coefficient for carbon dioxide Mass transfer coefficient for water Mass transfer coefficient for hydrogen sulfide Latent heat of evaporation at Tw Length of the lagoon at the bottom Molecular weight of air Molecular weight of water Constant The number of animals whose manure is transported to the lagoon Number of bubbles in the lagoon Mass transfer rate of carbon dioxide Mass transfer rate of water Mass transfer rate of hydrogen sulfide Average number of sulfur globules in a cell Nusselt number Constant Refractive index of an emitter Atmospheric pressure at the altitude of the surface above sea level Partial pressure of carbon dioxide in the gas phase Partial pressure of hydrogen sulfide in the gas phase xiii Units m/s m/s m/s j/kg kg/mole kg/mole mole/s mole/s mole/s Pa Pa Pa Symbol Pr Re Description Prandtl number Instantaneous deviation of the water vapor pressure from the time-averaged water vapor pressure Vapor pressure of water Saturated water vapor pressure at Tw Water vapor pressure at height 21 Water vapor pressure at height 22 Long wave radiant energy from the atmosphere Gas production rate Indirect solar radiation Reflected solar radiation Direct solar radiation Long wave radiant energy from the water Net input of heat by evaporation and condensation Net total input of heat Net input of heat by radiation Net input of heat by radiation during the night Net input of heat by rainfall Net input of heat by direct heat transfer Gas constant Reflectivity Bowen ratio The radius of the bubble surface Reynolds number xiv Units Pa Pa Pa Pa Pa 2 S 21 23 S S S W W W m3Pa/mole K Symbol Description Surface area of the sulfur globules per unit volume lagoon water Schmidt number Sherwood number Specific surface of the bubbles Absolute temperature Absolute temperature of the air Reference temperature Absolute temperature of the water time Rate of urine production per unit weight of animals Rising velocity of a bubble Wind velocity in main air stream Wind velocity at the height 21 Wind velocity at the height 22 Wind velocity at a height 2 The total volume of a lagoon The volume of a bubble Volume of the lagoon below height h Molecular gas volume Width of the lagoon at the bottom Average weight of the animals Instantaneous deviation of the vertical velocity from the time-averaged vertical velocity Concentration of desulfurizing bacteria XV Units 79 '23 91 9Q m3/mole m kg/animal m/s cells/m3 Symbol Description Concentration of purple sulfur bacteria Size parameter of the lagoon surface Yield coefficient for the reduction of sulfate Yield coefficient for the oxidation of hydrogen sulfide Yield coefficient for the oxidation of sulfur Yield coefficient for the reduction of sulfur Group, defined in equation (6.22) Group, defined in equation (6.28) Two different heights above the water surface Heat transfer coefficient The angle of the side slopes of a lagoon Extinction coefficient temperature Wavelength measured in vacuo Heat conductivity of air Heat conductivity of water .growth rate Specific growth rate of the desulfurizing bacteria Specific growth rate of the purple sulfur bacteria oxidizing hydrogen sulfide Specific growth rate of the purple sulfur bacteria, reducing sulfur Specific growth rate of the purple sulfur bacteria for light-limited growth xvi Units 3 cells/m m cells/mole cells/mole cells/mole cells/mole W/mZK deg l/m °C W/m K W/m K 1/3 1/3 l/s 1/s 1/s 1! Description Dynamic viscosity of air The maximum growth rate of the desul- furizing bacteria The theoretical maximum growth rate for the oxidation of hydrogen sulfide The maximum growth rate for the oxidation of sulfur The maximum growth rate for the reduction of sulfur The theoretical maximum growth rate for light limited growth Reference dynamic viscosity Dynamic viscosity of water Density of air at Ta Density of sulfur in globules Density of water The density difference between a bubble and water Surface tension of water Stefan-Boltzmann constant Contact time of a water particle with a bubble The time a bubble is in the water Angle of incidence Angle of refraction Solar angle Concentration Equilibrium concentration xvii Units kg/m 5 l/s l/s l/s l/s l/s kg/m 3 kg/m 5 kg/m3 kg/m3 kg/m3 kg/m3 kg/s2 W/mzK4 rad rad rad mole/m3 mole/m3 Subscripts: a b air bubble feces gas height sulfur urine water xviii INTRODUCTION Since the introduction of confined housing for ani- mal production, manure management has become a problem: the manure was not defecated directly on pasture but in a building. Consequently, manure had to be transported. It is possible to design different combinations of storage and transport techniques, which vary from labor-intense systems with low capital investment to systems having almost no labor but requiring a high capital investment. The system under consideration in this thesis treats the manure as a liquid. Swine manure is transported out of a slatted—floor growing-finishing building several times each day by flushing with water. The manure thus diluted is carried to an anaerobic lagoon. In order to have adequate storage capacity in a small volume lagoon, water is pumped back to the building and used for the flushing operation. .An anaerobic lagoon is currently considered to be the most economical and convenient system for storage and stabiliza- tion of manure. A major disadvantage of the anaerobic lagoon is the potential for odor production. The simple storage of animal manure in an anaerobic lagoon is an example of the complexity of nature. Not only do the processes of anaerobic digestion take place, but also photosynthesis by bacteria or algae. The Open structure makes it also susceptible to variations induced by meteoro- logical and climatic conditions. Consequently the processes and their rates are functions of time and place. In this thesis I will evaluate a number of variables, which play a role in the behavior of an anaerobic lagoon. In Part I the basic characteristics are described. In Part II these variables are integrated into a model, designed to improve the understanding of the complex inter- actions. I will attempt to make the model credible by com- paring it with experimental results. Finally I will use this model to propose management practices which will result in a better lagoon performance. Particular attention is paid to lagoon operation in the North Central States, where anaerobic lagoons work in less than favorable climatic conditions. PART I REVIEW OF LITERATURE CHAPTER 1 LAGOONS 1.1 Introduction A lagoon is an open structure in the soil for the storage and/or treatment of wastewater. There is no clear distinction between different types of lagoon, but rather a continuum from one extreme to the other. Generally lagoons are subdivided into aerobic, facultative and anaerobic lagoons. This subdivision is based on the availability and presence of oxygen. Other subdivisions, which can be made, are according to the type of waste (i.e., agricultural, industrial or municipal) according to the type of discharge (i.e., no dis- charge, irrigation or discharge to surface waters) according to the type of climate (i.e., cold, warm or tropical, wet or dry climates) or according to the depth (i.e., shallow or deep lagoons). All these subdivisions are arbitrary and examples of all of these can be found. The lagoon type of interest in this thesis could be described as an anaerobic cool-and-wet climate lagoon for swine waste with land application as discharge. Major sources of information about lagoons are conference papers, edited by Gloyna gt a1. (1976) and the annual literature reviews of the Water Pollution Control Federation (Burkhead and O'Brien, 1974; and O'Brien, 1975, 1976 and 1977). The following general discussion will be based on the subdivision into aerobic, facultative and anaerobic lagoons. 1.2 Aerobic lagoons Aerobic lagoons are characterized by the presence of oxygen throughout the body of water. They are used for the storage of irrigation and drinking water and as a polishing step for wastewater treatment. Aerobic lagoons are only used for wastewater treatment in cases where stringent odor or effluent requirements exist. Design of these lagoons is based on the use of the receiving water. Therefore criteria are used such as algal growth potential, median toxicity limits for fish, viral plaque forming units and selective disinfection kinetics of algal-bacterial effluents. Conse- quently the treatment is directed toward reduction of sus- pended solids, such as algae, reduction of enteric bacteria and reduction of nutrients such as nitrogen and phosphorous. If the effluent is used for irrigation, additional criteria such as the sodium:potassium ratio and dissolved salt limit should be considered (Gloyna, 1976). 0 Generally aerobic lagoons are shallow to permit natural aeration, but sometimes they are built with a con- siderable depth and forced-aeration is applied at a low rate. Aerobic lagoons are not suitable for the treatment of wastes from livestock production units. These wastes are too concentrated. 1.3 Facultative lagoons Facultative lagoons are lagoons in which an anaerobic and an aerobic environment coexist. Sometimes a lagoon is facultative the whole year around, but quite often an aerobic lagoon turns anaerobic near the bottom during adverse condi- tions or an anaerobic lagoon acquires an aerobic layer at the surface during periods of lower loading rates or other action which reduces the biochemical oxygen demand (BOD). Facultative lagoons are also called waste stabiliza- tion ponds, since their main application is in the reduction of the BOD of wastewater. Compared with activated sludge processes, in which forced aeration is applied, a faculta- tive lagoon requires more land and has a higher content of biological solids, such as algae. There are however, many advantages, such as a higher BOD removal, less energy and chemical consumption and much lower manpower requirements. In addition, lagoons are less sensitive to influent char- acteristics than are activated sludge treatment plants. The anaerobic environment at the bottom of a facul- tative lagoon serves to keep nutrients available for the algae. Gloyna (1971) describes an empirical relationship for the design of facultative lagoons, which includes a correction factor for the temperature and for sulfide toxicity. A sulfide concentration of 7 ppm is toxic to algae. A facultative lagoon will function at temperatures above 5°C. The most important design parameter is, however, the surface area. Loading rates are therefore expressed per unit area of lagoon surface. A larger area will result in a larger input of oxygen from the air and will permit the penetration of more light into the water. Shallow lagoons are very susceptible to mixing by the wind resulting in the suspension of solids and reduced light penetration. Photosynthetic bacteria can occur in facultative lagoons at the upper boundary of the anaerobic zone. Their coloring of the lagoon water is, however, masked by the large number of algae, which predominate in these lagoons. 1.4 Anaerobic lagoons Characteristic of anaerobic lagoons is the complete absence of oxygen. Oxygen which enters via the surface is very rapidly consumed in the uppermost film of lagoon water. Although all lagoons, which lack oxygen are called anaerobic, there is still quite a variation possible depending on how low the oxidation-reduction ("redox") potential is. With decreasing redox potential, the following processes take place: denitrification, acid fermentation, sulfate reduction and finally, methane fermentation. In the next chapter these processes will be discussed in greater detail. In an anaerobic lagoon the several processes will occur simulta- neously with process rates being affected by depth. Anaerobic lagoons can be built much deeper than aerobic or facultative lagoons. A minimum depth is required, however, to maintain anaerobic conditions. Since anaerobic bacteria have low growth rates, an anaerobic lagoon should not be pumped out completely because it is necessary to retain a minimum bacterial culture. This lowest allowable level is called the design volume. Consequently the design of anaerobic lagoons is not based on the load per unit sur- face as for facultative lagoons, but on the load per unit volume. If conditions are suitable, photosynthetic bacteria can establish themselves in large numbers in anaerobic lagoons. Their characteristics and their occurrence in lagoons are described in Chapter 3. Most types of algae cannot survive in anaerobic lagoons because of their oxygen requirement. There are however, some types which can perform fermentative reactions (Wiedeman and Bold, 1965). Often, however, the sulfide concentration is above the toxic level of 7 ppm. Photosyn- thetic sulfur bacteria can stimulate the occurrence of these algae. CHAPTER 2 ANAEROBIC TREATMENT 2.1 Introduction In the presence of high concentrations of organic material, anaerobic processes will occur as long as the supply of oxygen is insufficient. Even in ancient times, it was known that in swamps a combustible gas is formed. This is mentioned, for example, in the Historia Naturalis of Pliny (Haberly, 1957). For centuries this gas has been used for cooking and heating. It was Volta (1776) who dis- covered that the formation of this gas was related to the decomposition of organic material. For about a century, use has been made of the anaerobic processes in the treatment of organic waste. This has developed in a wide variety of constructions in which the environment is controlled. Starting in 1860, the first anaerobic digesters were developed especially for the treat- ment of sewage sludge (Cameron and Travis in England, Mouras in France and Imhoff in Germany). In 1906 the Imhoff-Tank or Emscherbrunnen was used for the first time (Imhoff, 1916L In the Imhoff-tank, the digestion area is built directly under the sedimentation basin of a sewage treatment plant. In 1914 Imhoff improved the distribution of the sludge supply and in 1916 he introduced the principle of mixing the incoming sludge with the content of the digester. Since that time, study of many environmental factors has resulted in the design of improved systems. These developments were generally accompanied by an increase in complexity of operation, by an increase in cost and therefore by an increase in the economical size of operation. In comparison, farms are small scale systems. Farming is already such a diverse enterprise that one must avoid any unnecessary complication in equipment and manage- ment. A gas collection system was shown to contribute more costs than benefits (Nordstedt, 1976). The principal biological processes for anaerobic lagoons and anaerobic digestion and the governing natural laws are the same. The anaerobic lagoon, however, avoids the complexity introduced in the environment controlled digestion process. Some important sources of information about the anaerobic treatment processes include the biblio- graphy of Shadduck and Moore (1975) about the digestion of livestock wastes, the review of Regan (1975) and the annual literature review of the Water Pollution Control Federation (Ghosh and Conrad, 1974 and 1975; Ghosh et 21., 1976 and 1977). 10 2.2 Anaerobic digestion processes The basic feature of these processes is the degra- dation of volatile organic compounds through the action of microorganisms. These compounds are hydrolyzed with the aid of extracellular enzymes to a form which can be taken up by the microorganisms. The organisms which produce these extracellular enzymes are named acid-forming microorganisms or "Acid-formers," because they convert these hydrolyzed organic compounds mainly into lower molecular weight fatty acids. These acids can be used directly or indirectly by methane forming organisms, which convert them into methane and carbon dioxide. As a result of the methane digestion, organic compounds are converted almost completely into gases. The principle organic compounds in sludge and manure are carbohydrates, proteins and fats. Each of these will be described shortly in the following discussion. The most important carbohydrates are starch, cellu- lose and glucose. Starch and glucose are decomposed along well known pathways. Their decomposition is a relatively fast process. The breakdown of cellulose, however, requires specific environmental conditions (Bryant, 1973 and Leather- wood, 1973). The predominant cellulose-hydrolyzing bacteria use ammonia as the only source of nitrogen and are unable to use amino acids. These bacteria also need sodium, B—vitamins and branched volatile fatty acids for the synthesis of amino acids. The most important form of the enzyme cellulase is 11 supposed to be built up of two groups: a hydrolyzing group and a group which attaches itself to the cellulose (Smith, 1973). The intensity of attachment to cellulose is strongly reduced by the presence of lignin and silicon, and by the crystallinity of the cellulose. If the attachment were not reduced by lignin and the other factors, hydrolysis would proceed much faster. The enzyme cellulase can be excreted by the following organisms (Leatherwood, 1973 and Dehority, 1973): Ruminococcus albus, Butyrivibrio fibri- solvens and Bacteroides ruminicola. The cellulose is con- verted by cellulase to the disaccharide cellobiose, which is further converted intracellularly to glucose. The decomposition of cellulose is a slow process. Proteins are broken down outside the cell by pro- teolytic enzymes. A wide variety of organisms can do this. The resulting amino acids are used within cells for biosyn- thesis or are further broken down with the formation of CO H H S, NH and HCN (Chanin, 1961). 2 3 Fats are hydrolyzed by the enzyme lipase to gly- 2' 2' cerol and fatty acids. The glycerol is converted to pyru— vic acid. The fatty acids are decomposed by B-oxidation (Jeris and McCarty, 1965). The decomposition of fats results, therefore, mainly in production of acetic acid. According to Jeris and McCarty (1965) formic acid is con- verted into methane by reduction and acetic acid is con- verted into carbon dioxide and methane by transmethylation. 12 Acetic and formic acid are intermediate products formed during the degradation of all longer-chain fatty acids. Formic acid is converted so fast that it is seldom detected. Methane forming organisms can utilize only a limited range of compounds. The most important substrates are hydrogen (with carbon dioxide) and formic acid. Andrews (1965), Cookson and Burbank (1965) and Laskin and Lechevalier (1973) list a number of methane forming orga- nisms with their substrates. Some of the organisms listed are now considered to be symbiotic associations between two organisms. The best known example for this is Methano- bacterium omelianskii. Each organism performs a part of the ethanol decomposition (Bryant gt 31., 1967 and Bryant, 1967). 2 CH3 CH2 OH + 2 H20 + 2 CH3 COOH + 4 H2 C02 + 4 H2 + CH4 + 2 H20 The first organism is inhibited by hydrogen which makes isolation difficult. Winfrey and Zeikus (1977) and Winfrey 23 31. (1977) describe how methanogenesis is inhibited by small quantities of sulfate. The results of measurements in Lake Mendota sediments are explained very well by the pure culture studies of Bryant et_§l, (1977). Bryant 22 31. found, that desulfurizing bacteria can have a similar symbiotic association with H -utilizing methanogenic bacteria as 2 13 described for Methanobacterium omelianskii, provided sulfate is absent. Not only will this association metabolize ethanol as described above, but the desulfurizing bacteria produce acetate, hydrogen and carbon dioxide from lactate. Small concentrations of sulfate, however, change the meta- bolism of the desulfurizing bacteria. Instead of producing hydrogen they will use it seemingly at a much faster rate than the methanogenic bacteria, resulting in suppression of methanogenesis. D'Allessandro gt gt. (1974) describes how the desul- furizing bacteria use the hydrogen from lactate metabolism for the reduction of sulfate: 3 8042- + 4H+ + NH4+ + 7 CH3 CHOHCOO- + 3H8 + 6 CO2 + C5 H7 O2 N + 5 CH3 COO + 9 H20 In the anaerobic environment, denitrification also takes place. Nitrates and nitrites are reduced to the gaseous products nitrous oxide and molecular nitrogen. This loss of nitrogen out of the environment is considered as a favorable aspect in wastewater treatment. For agricultural purposes, however, this means a loss of fertilizer value. Since nitrates and nitrites are unlikely to form in anaerobic lagoons, this loss is probably negligible in comparison with the loss of nitrogen by the volatilization of ammonia. 14 2.3 The influence of thegphysical environment The most important physical factors in the anaerobic treatment are the intensity of mixing, the temperature and the availability of light. As mentioned above, Imhoff (1916) found that mixing can considerably improve the rate of digestion. Some twenty years ago, Myers (1961) and Schreiber (1962) worked on the effects of mixing on the anaerobic digestion process. Their recommendations are incorporated by Bargman (1966) in a Manual of Practice for sludge digestion. Mixing has the following positive effects. a. Newly introduced sludge comes rapidly to the same temperature as the bulk of the digestion tank. b. The mixing of new sludge with large quantities of digesting and buffered sludge causes a fast contact between substrate and microorganisms resulting in a rapid start of the decomposition process. o. By maintaining a uniform mixture of all compounds the negative effect of high local concentrations of decomposition products and toxic substances is diminished. d. By mixing a digester, the whole volume is used. If "dead" corners are avoided in this way the size of the digester can be reduced. e. Mixing results in a better separation of the produced gas and therefore reduces the accumulation 15 of scum. Scum is a layer of solids and gas at the surface. The negative effects of mixing are: a. It requires energy. b. Some gas is lost. Because of the negative effects, mixing is generally limited to short periods before and after the introduction of new sludge, which takes place once or twice daily. In order to obtain the beneficial effects of mixing, the visco- sity has to be kept low. The viscosity rises dramatically with an increase in total solids. A practical upper limit for the total solids is therefore about twelve percent. The temperature has a strong influence on the digestion process. Most attention has been given to meso- philic digestion (ls-35°C) because this temperature range is fairly close to environmental conditions. In a few instances, thermophilic digestion (about 60°C) is applied. Psychrophilic organisms can occur during the winter in unheated digesters and lagoons. Digestion is then very slow and incomplete. Fair and Moore (1937), Burd (1968) and Maly and Fadrus (1971) present graphically the influence of tempera- ture on the time after which the digestion can be con- sidered as completed. These times apply to digesters in which the environmental conditions are kept fairly constant. At higher temperatures the decomposition is more complete, 16 resulting in an increased gas production. Thermophilic digestion, however, requires a lot of energy and results in a bad-smelling effluent. According to Garber (cited in Burd, 1968) thermophilic-digested sludge has better de- watering characteristics because of the more complete degradation of proteins and the formation of larger particles. Speece and Kem (1970) studied the influence of short-time temperature variations as they occur during the introduction of new sludge. Even if the temperature drops a few degrees the gas production stOps. After a time which depends on the time and intensity of the temperature dis- turbance, gas production will resume. Organisms require about two hours to adapt after a step decrease in tempera- ture of 10°C. Up to 45°C an increase in temperature will result in an increased rate of decomposition. Above this temperature, however, a rapid decrease occurs. In lagoons the temperature will never become too high. Light plays no role in digesters, because of the closed structure. Besides the influence on photosynthetic organisms in lagoons (to be discussed in the next chapter), light can also influence the performance of some methano- genic bacteria. Pantskhava (1973a and b) studied the con- version of methanol to formaldehyde, carbon dioxide, pyruvic acid and vitamin 812 by cell-free extracts of Methanobac- ;§§rium kuzneceovii in the light. The influence of light is 17 stronger at higher temperatures. Light has also a stronger effect when the reaction occurs in a hydrogen atmosphere rather than a nitrogen atmosphere. Light is thus reversing the degradation processes which take place in the dark. Light seems to inhibit methane formation in at least some methanogenic organisms. 2.4 The influence of the chemical environment The most important chemical factors in the anaerobic environment are the pH, the concentration of volatile or- ganic acids and the concentration of cations. In some cases compounds foreign to the system have a detrimental effect on the decomposition process. After the start-up of a digestion process the pH will go down, because the acid forming bacteria grow faster than the methane forming bacteria. When the number of methanogenic bacteria increases, the pH will be restored to near neutral conditions. At the same time the alkalinity builds up, protecting the system against fast pH changes. If, however, the loading rate is increased too fast or if the loading is irregular the methane formers cannot keep pace with the acid formers and the alkalinity drops. Once the alkalinity becomes too low the pH starts falling. As this happens methane formation is the first process to be :hahibited. Since the acid-forming bacteria have a lower Ifli optimum (Albertson, 1961) the pH will initially fall 18 faster and faster until a pH of about 6 is reached. If no action is taken the pH can go down as low as a value of 3. The above sequence of events can be initiated by an increased loading, a drop in temperature or introduction of inhibitors of methanogenesis (e.g., oxygen). In animal waste lagoons the alkalinity is generally quite high. A rapid change in pH is therefore unlikely. If, however, a farmer discharges the contents of a manure storage pit into a lagoon, the tolerable loading rate can be far exceeded. Andrews and Graef (1971) among others describe the action of the bicarbonate buffer. Carbon dioxide occurs in several different forms: [C02] gas, [C02] dissolved, [HZCO3], [H CO3-] and [C032-]. The equi- librium between gaseous CO and dissolved CO is described 2 2 by Henry's Law: I = * [C02w Kco2 Pco2 (2'1) in which: [C02‘; = the saturated concentration of dissolved carbon dioxide gas in equilibrium with the carbon dioxide in the gas phase (mole/m3) KCO2 = Henry's constant for carbon dioxide (mole/m3. Pa) PCO2 = partial pressure of carbon dioxide in the gas phase (Pa). 19 In reality, equilibrium is never attained because there is a resistance to mass transfer at the gas-liquid interface. The mass transport rate becomes: NCO2 = kCO2 * A * (lcozlw - [COzl'wl (2.2) in which: NCO = mass transfer rate of carbon dioxide 2 (mole/s) kCO = mass transfer coefficient for carbon 2 dioxide (m/s) A = transfer surface area (mZ/m3) [C02]w = actual concentration of carbon dioxide in water (mole/m3) At equilibrium the concentration of [C02]w far exceeds the concentration of H2C03. The concentration of C032- is negligible as a pH of about 7. un< €02 = 10.25). There remains the equilibrium between carbonic acid and bicar- bonate: 1 = 4-3 * 10-7 (2.3) + K — [H30 ] [HCO3 CO2 [H2 CO3] According to Andrews, neglecting the resistance to mass transfer at the interface causes an error of about 10%. Wood (1962) demonstrates the strong influence of pH on the reaction mechanism with an experiment in which Escherichia coli was grown on glucose at a pH of 6.2 and 20 7.8. At the higher pH for example, the enzyme is inhibited which dissociates formic acid into hydrogen and carbon dioxide. Albertson (1961) among others gives a pH optimum for the anaerobic digestion between 6.8 and 7.2. At a higher pH (pH > 8) free ammonia can become toxic. According to Andrews and Graef (1971) the inhibition of methanoqenic bacteria at a lower pH is caused by the concentration of undissociated volatile fatty acids. In the next section I will come back to this point. McCarty and McKinney (1961b) on the other hand state, that more often digester indigestion occurs because of excessive concentrations of cations rather than anions. Kugelman and McCarty (1965) introduce a 50% inhibition index, i.e., the concentration of a cation (in eq/l) at which the reaction rate is decreased to 50% of the rate of a control unit. They get the following results: cation: Na+ NH4+ K+ Ca2+ Mg2+ 50% inhibition index: .32 .25 .15 .23 .16 Low concentrations of cations other than the one applied reduce the effect. The inhibition by Na+, K+ and NH4+ can be eliminated almost completely by the antagonistic action of Mg2+ and Ca2+. Optimum concentrations for all cations appear to be around .01 eq/l. The wastewater from industry often contains chemical compounds which are normally not found in the natural (anaerobic environment. The suitability of a lagoon or 21 digester treatment depends, then, on the tolerance of the system towards these compounds. In the most favorable case, the system is not affected. Waste from animal production units can also be toxic, e.g., because of antibiotics or because of copper additives to the feed. McDermott (1963a and b, 1965) finds that nickel concentrations below 40 ppm have almost no effect. Copper on the other hand inhibits the organisms above 10 ppm. Zinc has an effect intermediate to those of copper and nickel. Pallasch and Triebel (1969) state that concentra- tions of 1 wt% copper, chromium and nickel based on dry solids will totally inhibit the fermentation and concen- trations of respectively .5, .5 and .3 wt% will reduce the gas-production to 50%. Hydrogen sulfide gives a precipitate with heavy metals. Lawrence and McCarty (1965), Masselli (1967) and Goebgen and Brockman (1969) suggest, therefore, the use of H28 to counteract heavy metal toxicity. Tenney and Budzin (1972) state that fluoride is toxic at a concentration of about 1000 ppm. Such a concen- tration of fluoride normally does not occur. Much is written about the inhibition by chlorinated .hydrocarbons. Prins (1972) states that chlorinated analogs of methane inhibit in the micromolar range. According to Bauchop (1967) the methane forming bacteria are quite Specifically inhibited by chloroform. Sykes and Kirsch (1972) give a concentration of 16 ppm above which carbon 22 tetrachloride becomes toxic. Carbon tetrachloride has also a direct influence on the acid production: hydrogen accumu- lates, production the production of According inhibits the rate (1972) and Jensen 50% of the DDT is Hernandez of acetate and propionate is reduced and valerate and caproate increases. to McBride and Wolfe (1971) 1 uM DDT of methane formation by 75%. Albone (1972) report however, that after 7 hours converted into analogs. and Bloodgood (1971) studied the influence of linear alkyl benzene sulfonates (ABS). Concentrations above 1 wt % on a dry matter basis cause inhibition. The ABS can, however, be degraded by the process. Nitrilotriacetate (NTA), used as a substitute for phosphates in detergents, is degraded by anaerobic fermen- tation and can serve even as the sole source of carbon (Enfors and Molin, 1973a and b). Bishop (1972) studied the conversion of mercury to the very toxic methylmercury. Mercuric salts, however, in the presence of H28 result in the very insoluble HgS. 2.5 Models and simulation of the anaerobic treatment Many authors have composed models of the digestion process in order to obtain a better understanding of the .process dynamics and of the microorganism-substrate inter- lactions under different environmental conditions. In 1967 Lawrence and McCarty (1969) tested the Dhanod model for application to steady state anaerobic 23 digestion and in 1969 they added to this the possibility of varying the solids retention time to account for the re- cycling of solids. They found that the Monod model gave a good description of the system. An increase in the solids retention time permits higher loading rates. Concurrently, Pfeffer (1968) modeled the influence of the recycling of solids. Pfeffer's model was later extended by Fan (1973) to a two stage digestion with mixed cultures. In the meanwhile Andrews and Graef (1971) developed their model based on earlier experimental results (Andrews, 1965). The improved model included an inhibition function with undissociated acetic acid as both growth limiting sub- strate and inhibitor. The model also included the inter- actions between gas, liquid and biological phases. An impor- tant aspect is the above mentioned bicarbonate buffer (Andrews, 1971 and Graef and Andrews, 1973). Later Andrews and Graef added to their model the influence of base addi- tion, organism recycling and gas scrubbing. Base addition counteracts pH and alkalinity decreases and organism recycling conserves the slow growing biomass. Gas scrubbing removes ammonia, hydrogen sulfide and carbon dioxide. The basic features of their model are applied by Hill and Barth (1974) to explain the results of bench-scale lagoon models. By empirically adjusting certain values (1975) they could approximate the bench-scale lagoon loehavior at different temperatures; much deviation between 24 model and experimental results still remains, however. In 1977, Hill and Nordstedt added ammonia inhibition to this model in order to account for failure at a high pH. Ammonia inhibition is illustrated by Abeliovich and Azov (1976) who observed complete inhibition of algal growth at ammonia concentrations above 3 mole/m3. A more experimental approach was taken by Ghosh and Pohland (1974), who measured the performance of a two stage digester with recycling of solids. McCarty (1971) added to the modeling literature a completely different aspect by taking as basis for his cal- culations the energy transformation of the different meta- bolic processes. The major discrepancy in applying the results of process simulations to practical situation arises from the occurrence of process instabilities, caused by large vari- ations in influent flow rate and influent concentrations. This is obvious from the results of Hill and Barth, who loaded their units once a week. The transient behavior of biological processes was modeled by three investigators at the Rice University in Houston: Schaezler, McHarg and Busch (1971) present a model which takes into account such features as the lag phase, linear growth, and logarithmic growth. Their model uses three sets of relationships, one of which operates according to the growth phase of the organisms. The transition algorithm which selects the 25 operating relationships is missing from their paper, however. They managed to make their model describe the experimental results quite well. CHAPTER 3 PURPLE SULFUR BACTERIA 3.1 Introduction In several instances purple sulfur bacteria have been found in waste treatment lagoons (van Lotringen and Gerrish, 1977). Where found, they are always associated with reduced odor levels. George (1976) estimates that about 40% of all livestock waste lagoons in Missouri are purple as a result of these organisms. In order to be able to make use of these bacteria for odor control it is important to know which types of bacteria are active, what metabolic characteristics might possibly be important in odor control and which environ- mental factors influence their proliferation. It is clear that the major environmental factors are the type of waste and the lagoon management program. This will also set the lindtations on their use. As will be described more extensively in one of the following paragraphs, the most frequently identified purple sulfur bacteria in lagoons are Chromatium vinosum, Thiocapsa roseopersicina and Thiopedia rosea. Therefore I will 26 27 direct the following discussion mainly to the characteri- stics of these species. Comprehensive descriptions of photosynthetic bac- teria are given by van Niel (1931) and Kondrat'eva (1965). A more recent review is written by Pfennig (1967). 3.2 Classification The purple sulfur bacteria can be classified in several different ways as to motility, morphology, photo— synthetic pigments, formation of gas vacuoles, distribution of sulfur-globules, ability to form slime capsules, meta- bolism, formation of poly-B-hydroxy butyrate (PHB) granules, storage carbohydrates or phosphate deposits and to the redox potential at which they occur. The description of Pfennig in Bergey's Manual (1974) serves as a guide for the classification. The purple sulfur bacteria belong to the order Rhodospirillales and form the family of the Chromatiaceae. Members of this family are defined as "cells which are able to grow with sulfide and sulfur as the sole photosynthetic electron donors. In the pmesence of sulfide, globules of elemental sulfur are formed inside or outside the cells and further oxidized to sulfate." The first criterion for subdividing the Chroma- tiaceae is the site of storage of sulfur globules: I. Sulfur globules stored inside the cells. II. Sulfur globules stored outside the cells. 28 To the second group belongs only one genus of purple sulfur bacteria: Ectothiorhodospira. The first group is further subdivided into: A. Cells without gas vacuoles. B. Cells with gas vacuoles. Both of these are further subdivided as to motility. To the motile cells without gas vacuoles belong: a. Chromatium, cells ovoid to rod-shaped. b. Thiocystis, cells spherical, typically diplococcus- shaped before cell division. c. Thiosarcina, cells spherical to ovoid. Grouped as regular sarcina packets. d. Thiospirillum, cells clearly spiral-shaped. The non-motile cells without gas vacuoles are represented by the Thiocapsa. The motile cells with gas vacuoles are the Lampro- cystis. To the non-motile cells with gas vacuoles belong: a. Thiodictyon, cells rod shaped. b. Thiopedia, cells spherical to ovoid, characteri- stically arranged in regular platelets (flat sheets). c. Amoebobacter, cells spherical. Above are given the main characteristics of the 10 genera. Another important aspect is the composition of ‘bacteriochlorophyll (BChl) and carotenoids, which helps in recognizing the cells. Belonging to the suborder 29 Rhodospirillineae the purple sulfur bacteria contain BChl a or b. According to Meyer (1973) Thiocapsa pfennigii is the only species of purple sulfur bacteria and Rhodopseudomonas viridis (a purple non-sulfur bacterium) the only other known bacterial species having BChl b. The subdivision according to carotenoids places the phototrophic bacteria (including purple sulfur bacteria) into one of five groups: 1. The normal spirilloxanthin series with lycopene, rhodopin and spirilloxanthin as major components. 2. The alternative spirilloxanthin series and keto- carotenoids of the spheroidenone type. The major components are spheroidene, hydroxyspheroidene, spheroidenone, hydroxyspheroidenone and spirillo- xanthin. 3. Okenone series with okenone as major component. 4. Rhodopinal series with lyc0pena1, lycopenol, rhodo- pin, rhodopinal and rhodopinol as major components. 5. Chlorobactene series with Chlorobactene, hydro- xychlorobactene, B-isorenieratene and isorenieratene as major components. Thiocgpsa pfennigii is again an exception. It contains TetrahydrOSpirilloxanthin as major carotenoid. There are no purple sulfur bacteria known to belong to the groups 2 and 5. Much work on the identification and biosynthesis of the carotenoids of the purple sulfur bacteria has been 30 done by Jensen (1963), Schmidt gt gt. (1963), Schmidt (1963), Jensen and Schmidt (1963) and Schmidt gt gt. (1965). The results of these reports are summarized in an article by Schmidt gt_gl. (1965). (See also Pfennig in Bergey's Manual, 1974.) To group 1 belong Chromatium vinosum, g. gracile, Q. minutissimum and Thiospirillum jenense, Thiocapsa roseo- persicina, the Amoebobacter sp., Thiopedia sp. and Ecto- thiorhodospira sp. To group 3 belong Chromatium okenii, Q. weissei, 9. minus and Thiocystis gelatinosa. To group 4 belong Chromatium warmingii, g. buderi, g. violascens and Thiocystis violacea, the Lamprocystis sp. and Thiodictyon sp. In general, Schmidt gt gt. (1965) and Pfennig (writing in Bergey's Manual (1974) agree on the carotenoid classification of the purple sulfur bacteria. In contradiction to the above, Pfennig (1974) describes Thigpedia rosea as belonging to the okenone series, based on a completely different group of pigments. In Schmidt et a1. (1965), the carotenoid compositions were reported as listed in Table 3.1. Okenone doesn't even appear as a minor constituent of Thiopedia rosea. Although Chromatium vinosum appears to have a different carotenoid cxnnposition, the absorption maxima of the carotenoids are 'very close to each other. The use of spectra fOr HammumHo an» «0 :oHuHmom map mH H o 31 ooom on mmv owe imam.ommv . u . mcocmxo oovm smm mmv mos Hmmm.momv «H OOH . mm 00H . om cHsucmonHHqum oovm smm mmq mos Hmmm.mmmv m H I o N u o :HsucmonHHuHmm pmumahnuwsmpocoz oesm sHm Mme ems Hmsm.vmmv o H u o v n o cHnnH>oeonm OOAN on mmv vmv Hmsm.emmc m H u o m I o :HunH>oeonuuouesnca oosm sHm mma «ma Loam.ammc OH 0 o :Hdoeonuuouesnmo v.m ooom mom was mas Hmom.mvmc mm m u o a u o chococm oovm vom mew mew Hmom.mamo o H u o H n o mammoosq scam Hmnuo Edmaouumm mmmmmmw. mmmmm. mcwoflmummoomou «H cH memes .th ssHumsouso mmmmmmmmm. .mmmmmmmmm .Hmoma .rmm.mm ucflenomv msflxmfi GAME mapcfls may mo mucmwowmmmoo noduocwuxm on» can mpflocmuoumo mmmcu mo MEfime cOHuQHOmnm on» .mflumuomn usuasm magnum owns» mo Hmpwocmuoumo Hmuou no a may cofiuflmomfioo pwocmuoumoilfl.m magma 32 identification is further complicated by the variability in the quantitative composition of carotenoids in a cell, which not only changes the intensity of absorption at different wavelengths, but also causes a shift of the absorption peaks. This can be caused by a difference in substrate, as is clearly demonstrated by Clayton (1963). All forms of purple sulfur bacteria are able to develop as single cells. Under unfavorable conditions and in many natural environments the cells can occur in more or less regular aggregates surrounded by slime. Thiocapsa roseOpersicina is always surrounded by a slime capsule. In contrast to most other purple sulfur bacteria Thiopedia rosea is normally arranged in regular platelets. Only two species of purple sulfur bacteria, Thio- capsa rosegpersicina and Amoebobacter roseus are shown to be able to grow under aerobic conditions in the dark (Kondrat'eva gt_gt., 1975). Both Thiocaposa roseopersicina and Chromatium vinosum can store polysaccharides, poly-B-hydroxybutyrate (PHB) and polyphosphates. Thiopedia rosea can store PHB; other storage products have not been shown. 3.3 Metabolism The purple sulfur bacteria display a wide range of rmetabolic processes. As source of energy they can use light, for which they can synthesize the necessary pigments, but during the dark they can use storage products. 33 Thiocapsa roseopersicina has even been shown to be capable of chemolithoautotrophy and chemolithoheterotrophy (Kondrat' eva, gt gt., 1975, Krasil'nikov gt gt., 1975). As a source of carbon the purple sulfur bacteria can use carbon dioxide,a1cohols, carbohydrates and a variety of simple organic acids. As source of reducing power they can use a whole range of reduced sulfur compounds and many strains are able to use molecular hydrogen. As a source of nitrogen several species can fix molecular nitrogen. Moreover, they can use ammonium salts and urea and a few use glutamate and aspartate as nitrogen sources. Purple sulfur bacteria, like the other photosynthe- tic bacteria, don't evolve oxygen during their photosynthe- tic activities. This led to a new concept of photosynthesis (van Niel, 1931). The fixation of carbon dioxide was studied by Eymers (1938) and later by Fuller gt gt. (1961) who identified several enzymes of the Calvin cycle and the citric acid cycle in Chromatium extracts. Fuller found that the glyoxylate cycle is modified in that the malic dehydrogenase reaction is missing. Trfiper (1964) found 'that acetate is used preferentially to carbon dioxide. According to Gest (1972) the photosynthetic bacrteria produce adenosine triphosphate (ATP) from radiant energy via a cyclic electron flow. Reduced nicotinamide dinuzcleotides are formed via a reversed electron transport chai;n, using organic compounds as source of reducing power. 34 The first extensive study of the sulfur metabolism of the purple sulfur bacteria was performed and published by Trfiper and Schlegel (1964), Trfiper (1964), Trfiper and Pfennig (1966) and Thiele (1968 a and b). They found a maximum storage of sulfur of 30.5% and a minimum content of .7% of the dry weight. In agreement with their results, van Gemerden (1968a) reported that the oxidation of inter— nally stored sulfur occurs simultaneously with the oxida- tion of sulfide. As a result, 58% of the sulfide is oxi- dized to sulfate at the moment of sulfide depletion. Using- a maximum storage of elemental sulfur of 30% of the cells dry weight he was able to quantitatively associate this oxidation with the production of bacterial cell material, which appears to have the approximate molecular formula C5 H802N (van Gemerden, 1968b). With ammonia as the source of nitrogen, van Gemerden gives the following overall reactions: 10 C0 + 21 H S + 2 NH + 2 (C H O N) + 218 + 16 H O 2 2 3 5 8 2 2 (3.1) 30 C02 + 21 S + 6NH3 + 36 H20 + 6 (CSHBOZN) + 21 H2504 (3.2) 40 C0 + 21 H S + 8 NH + 20H SO 2 2 3 0 I 8(C5H OZN) + 21 H2 (3.3) 2 8 4 35 These reactions are valid, however, only if carbon dioxide is used as source of carbon. In the presence of acetate the consumption of reduced sulfur compounds will be: 5 C2H4O2 + H28 + 2NH3 + 2 (CSHBOZN) + S + 6 H20 (3.4) 15 C H O + S + 6 NH + 6 (C 2 4 2 3 2N) + H SO + 14 H O 2 4 2 (3.5) 5H80 20 C2H4O2 + H25 + 8NH3 + 8 (CSHBOZN) + H2804 + 20 H20 (3.6) The above reactions lead me to estimate that for the same increase in cellular material, the carbon dioxide reduction would use 21 times the amount of sulfide as is used for acetate incorporation. For the production of storage carbohydrates, the overall equations with the use of carbon dioxide are (repre- senting the carbohydrates by (C6H1005)n): 12n CO2 + 28 nHZS + 2 (C6H1005)n + 14n H2O + 28nS (3.7) 42 n CO2 + 28 nS + 63 nHZO + 7 (C6H1005)n + 28 n H2 SO4 (3.8) 54 n CO + 28 n H S + 49 n H O + 9 (CGH SO 2 2 2 05)n + 28 n H 10 2 4 (3.9) 36 and using acetate: 3 n C H O 242"“C 6H1005)n + n H20 (3.10) So no H28 is used in the formation of storage carbohydrates from acetate. According to Hendley (1955) and van Gemerden (1968c) the storage carbohydrates are converted into PHB in the dark. During this conversion, internally stored sulfur is converted to sulfide: (C6H 05)n + n H O + 3nS + (C H O ) + 2nC02 + 3n H S 2 4 6 2 n 2 (3.11) 10 Not only CO2 but also acetic acid is formed. In the light the reverse overall process will take place. Consequently a lagoon with purple sulfur bacteria will theoretically produce more hydrogen sulfide, and thus theoretically more odor, during the night. During the day, the hydrogen sul- fide, and thus odor, is diminished. If colloidal sulfur is added to a culture, it can also be used by the purple sulfur bacteria. The rate of sulfur metabolism is determined by the available sulfur surface (intra- and extracellular sulfur combined) (van Gemerden and Jannasch, 1971), rather than by the sulfur concentration. 3.4 Interactions with the environment Although none of the purple sulfur bacteria (except Thiocapsa roseopersicina and Amoebobacter roseus) 37 have been shown able to grow in the presence of oxygen, Hurlbert (1967) found, that oxygen is not toxic to Chromatium strain D. Oxygen completely prevented the syn- thesis of BChl, but the cells were still able to oxidize thiosulfate and internally stored sulfur. This sulfur could not be oxidized to sulfate. If acetate is present it is mainly converted into PHB. As follows from the reactions given above, this conversion would result in a net production of reducing power. Hurlbert observed also, that in the presence of oxygen cells are unable to divide. Whether the cells were actually using oxygen was not determined. Holm and Vennes (1970) report an Optimum concen- tration of sulfide of 1.4 to 1.9 mmol/l for the growth of Thiocapsa roseopersicina and Chromatium vinosum (at pH between 7.5 and 8.2). Van Gemerden (1974) found an optimum concentration (at pH = 7.0) of about 0.08 mmol/l for Chromatium weissei and 0.11 mmol/l for Chromatium vinosum with saturation constants of 0.010 and 0.007 and inhibition constants of 0.7 and 2.5 mmol/l, respectively. Since undissociated hydrogen sulfide is the substrate and inhi- lbiting agent (van Gemerden and Jannasch, 1971) the optimum concentration will be lower at a lower pH. Pfennig (1967) gives the following order for tolerance to high concentra— ticnas of sulfide: Thiocapsa > Amoebobacter > Thiodictyon > Thiopedia. The above sequence is in agreement with the 38 observations of McFarlane and Melcer (1977) for lagoon bacteria arranged in order of decreasing lagoon load (and presumably sulfide load): Chromatium + Thiocapsa + Thiopedia For the optimum pH Holm and Vennes report 7.5 to 8.2. Meredith and Pohland (1970) report an optimum pH of 7.5 for a Chromatium sp. from a lagoon. As suggested above, the optimum pH and the optimum sulfide concentration are not independent. Generally the values reported for the optimum pH are between 7 and 8.5 (Kondrat'eva, 1965). The purple sulfur bacteria are found in nature at temperatures ranging from 8 to 80°C. Most species, however, have an Optimum between 18 and 30°C. For a lagoon system this means that on a couple of warm and sunny days, when the surface temperature can get as high as 35°C, the activity of the purple sulfur bacteria will be reduced, while the activity of the sulfide producing bacteria remains very high. Consequently a lagoon might be expected to produce more Odor under these conditions. Obviously, the light intensity has a strong influ- ence on the purple sulfur bacteria. Light is often the growth limiting factor under natural conditions, because of .a requirement for anaerobic conditions (i.e., the surface water which would get the most light may become aerobic). Kondrat'eva (1965) reports a light saturation at intensities :front 3 to 40 klx. Trfiper (1964) gives a light saturation 39 of 2 klx for Chromatium okenii and g. vinosum and van Gemerden (1968a) l klx for a Chromatium. Takahashi gt gt. (1970, 1972) studied the influence of the light intensity on Chromatium strain D. They found an optimum intensity of 2 klx and a compensation point between 0.05 and 0.01 klx. At the lower light intensities the bacteria show an increase in BChl content and an increase in efficiency of this BChl. At 1 and 20 klx they found growth rates of 0.087 and 0.179 hr-l respectively. The significance of the above-mentioned light intensities is only relative, since no description is given of the quality of the light. As described by Kondrat'eva (1965) the bacteria are only active at specific ranges of the spectrum. From the metabolic features it follows that the purple sulfur bacteria interact with many other species of bacteria. From the work of van Gemerden (1968c) it is clear, that different types of purple sulfur bacteria can exist at the same time, competing for the same substrates. By closing the sulfur cycle, purple sulfur bacteria and desulfurizing bacteria will stimulate each other. This effect is studied by Matheron and Baulaigue (1976) who tested it for the green sulfur bacteria. Since the purple sulfur bacteria use acetate Preferentially to carbon dioxide they will enhance the removal of volatile fatty acids from the lagoon environment. 40 Though they don't consume oxygen for this process, the purple sulfur bacteria will actually lower the biochemical oxygen demand levels in the liquid phase, as was found by Holm and Vennes (1970) and McFarlane and Melcer (1977). In their observations on lagoons Holm and Vennes found that the viable counts of the purple sulfur bacteria was usually about a factor of ten lower than the total counts. The maximum number of purple sulfur bacteria was reached after the desulfurizing bacteria reached a maximum, but before the other heterotrophic and the methanogenic bacteria reach their respective maxima. McFarlane and Melcer (1977) list as suitable condi- tions for the bacteria to flourish: l. underloaded anaerobic lagoons 2. overloaded facultative lagoons 3. selective effluents The first indication of the presence of the purple sulfur bacteria was the change in pH. In the succession of dominant organisms they place the purple sulfur bacteria between the primary anaerobes and the algae. By lowering the sulfide concentration the purple sulfur bacteria make the growth of algae possible. This would explain the frequent observation that a lagoon will turn from purple to green during the summer. CHAPTER 4 SWINE WASTE MANAGEMENT AT MSU This chapter is a description of the swine waste management system on which most of the work in this thesis has been done. The system consists of a slatted floor growing-finishing building from which the waste is trans- ported by flushing to two lagoons of equal volume, further identified as the east lagoon and the west lagoon. The growing-finishing building houses an average of 250 hogs with an average weight of 60 kg. For the first 8 weeks these hogs receive the standard grower ration MSU 16 (see Table 4.1) as their weight increases from approximately 18 to 50 kg. For the next 8 weeks, the animals receive the standard finisher ration MSU 13 (see Table 4.1) as their ‘weight goes from approximately 50 to 90 kg (see Miller, 1975). Nutrient and amino acid composition Of the dried swine feces were reported by Orr (1971, 1973) (see Table '4-32 and 4.3). A mineral analysis Of the swine manure has been made by Ngoddy (1971) . His results are reproduced in the 41 Table 4.1--Swine Feed Mix. 42 Grower Finisher MSU-16 MSU-13 wt % wt % Ground shelled corn 78.25 85.00 Soy bean meal (49% protein) 18.00 11.50 Calcium carbonate .75 .75 Defluor phosphate 1.25 1.25 MSU—VTM premix .50 .50 Salt .50 .50 Sel. E. premix .50 .50 Aureomycin SP-250 .25 -- 100% 100% 43 Table 4.2--Nutrient Composition of Dried Swine Feces (DSF). DSFa Nutrient l 2 Protein-nitrogen, % 3.48 3.44 Non-protein-nitrogen, % -- —- Crude protein, % 21.8 21.5 Calcium, % 2.8 2.2 Phosphorus, % 1.8 1.5 Sulfur, % 1.0 1.1 Potassium, % 1.2 0.9 Sodium, % 0.3 0.2 Chlorine, % -- -- Magnesium, % 0.1 0.1 Manganese, ppm 213 141 Iron, ppm 513 397 Zinc, ppm 432 586 Copper, ppm 117 98 aAnalyses of dried swine feces by P. K. Ku. Table 4.3--Amino Acid Composition of Dried Swine Feces (DSF). DSFa Amino Acid % Lysine 1.11 Histidine 0.40 Arginine 0.67 Aspartic acid 1.37 Threonine 0.80 Serine 0.58 Glutamic acid 3.37 Proline 0.91 Glycine 1.51 Alanine 1.14 Cystine 0.12 ‘Valine 1.04 Methionine 0 . 5 8 Isoleucine 1.03 Leucine 1.57 Tyrosine 0.65 Phenylalanine 0.87 aAnalysis by W. G. Bergen. 44 Tables 4.4, 4.5, and 4.6. The ratio of urine to feces as is given by Ngoddy is quite high in comparison with the values of Pratt (1975). The swine manure in transported to the two lagoons twice daily (at about 8:00 A.M. and 4:00 P.M.) by a flushing system. For this flushing operation, use is made of three tipping tanks and a sudden release tank, each having a volume of about 0.7 m3. A Y-valve is used to distribute the incoming manure to the two lagoons. Inlet conduits are corrugated plastic tubing, 0.15 in. diameter. A sketch of the lagoons is presented in Figure 4.1. Lagoon liquid is recycled to the barn for the flushing operation. Twice a year in April and November the lagoons are pumped out with a tractor-driver centrifugal pump. The lagoon is pumped via an irrigation line to the nearby cornfields. The lagoon operation at the MSU Swine Research Cen- ter was started in 1972. At that time, the lagoons were designed using the figure of 0.062 m3/kg of animal weight and each lagoon was to receive wastes from 140 hogs with an average weight Of 57 kg. Added to this was a 200 m3 volume to allow for the treatment of effluent from the oxidation ditches of two nurseries, each containing an average of 120-14 kg hogs. In summary, the design of each lagoon con- sisted of the following components (T. L. Loudon, 1977» 45 'onunu; Joann -p333 yo :uaazad u can 0013mm? 3 3 ”3'3 33"! "’0 «once—no ashascacoooo usumtnuHHHAAHHHHAHHAHHHHHHH :qirnn 9571 '91 000T cp/sqr "3 '3 '3 '3 '9 nun u01nuwnj33$$$323333333333333 (auxin + 03303) ("Pl‘qt) ““0950 MN“ 0 OHDONH outxuounuxmng‘e....°f ..."§."? NNHIflU‘Q §fl€|fl00 “VDQO‘OO are! qt ‘0an ‘11) "2 a: banana: N n (uzaaoaa 1) none: p00,; 3 3 0’2" on N 33335 :9»; H '3 >* “W"“)"TTT€ dentin auuwué~nnq nééecn n o (hp/Nu) _; '3 m up! 30 a: g ("U .e-.-8 3322:: 3223:: m SWIOAOEOJOAVnOONQ—c HMHNF‘H o-flHflHo-‘H fi’ (pm)wpq~~ncno HNMQWO Hwnvno g o u g z 3.3: u 3.2. H.:3 P‘ 1.9 —c c: -.o .n :5 -.o .4 u do u a U Go E “ass: 32:33: sagas: E seq-nu tit-1.1.3“ u < a. me SH.» as oemm owes coo.nm ooo.om omeue>< Hm N.@ on omHm comm ooo.om ooo.on OH em m.oH mm oeue coo» ooo.ee ooo.Hm m mm m.m em oewn oooe ooo.m~ ooo.me a «m m.m om ommH come ooo.He ooo.e~ a % mm ooé on Sen 83 25.3 08.8 e m.mm om.m oe moon oAHm ooo.~e .ooo.om n mm e.m me . nHmN omen ooo.m~ ooo.wo e mm m.e Hm emmm coon ooo.- ooo.He m em 0.» an owmm aces ooo.om ooo.o~ N on m.@ on HHoe oemm ooo.~m ooo.om H m> «we 2 Has a mo Ameev Hwaev H\mao HH\wao mmuHmoasou N N z n :2 N z a :2 z Hoz b .udom ’ ’ >Hsn F >mz b .umz D damn. r a Dem . Dov . oom floom (am/M) UOIQPTPPH IPIOS 209110 58 The reflected solar radiation is directly propor- tional to the incident radiation: 0R = R' * OS (5.4) in which R' is the reflectivity for direct solar radiation. The reflectivity depends on the solar height (which depen- dence is expressed in Fresnel's law as a function of the angle of incidence, Ti) and the angle of refraction (Wr). For an undisturbed water surface: . 2 . 2 . R' = 15 {31112 (9’1 - Wr) + tan2 (‘1’1 - 9%)} (5.5) sin (Wi + Wr) tan (Vi + 9r) Anderson (1952) gives an empirical formula for clear days: R' = 1.18 Ws-O'77 (5.6) in which TS is the solar angle. This empirical formula accounts for reflection of sun and sky radiation together. The reflection of radiation from the sky is estimated to be 7% for the total reflectivity. Johnsson (cited in Hutchin- son, 1957) gives a value of 6% in summer and 10% in winter. The long wave radiation from the atmosphere will depend on the temperature, and the quantity and type of cloud cover. Bolz and Fritz (1950) give the following equation: 4 2.5 - QA 0 Ta (1 + kc B ) (.820 .250 10 .126 * PW) (5.7) 59 in which QA is in cal/cm2 min and Stefan-Boltzmann constant Q II T = air temperature B = fraction of the sky covered by clouds Pw = the vapor pressure of water kC = factor depending on cloud type: Cloud type kC cirrus .04 cirro-stratus .08 altocirrus .17 altostratus .20 cumulus .20 stratus .24 The long wave radiation emitted from the water surface has a spectrum somewhat different from black body radiation. Anderson gives an emissivity of 97%: QW= .97 0' * tw4 (5.8) in which tw is the water temperature. Johnsson assumes for both air and water blackbody radiation: 11 4 4 (t' - t ) (5.9) .. = v: ' QW QA 8.26 10 w a in which Q is in cal/cm2 min. For the usual environmental temperatures he simplifies this formula further to: _ _ 2 0W - QA — ll (tW ta) cal/cm .day (5.10) 60 Using this equation we obtain for the radiation surplus during the night: _ _ . 2 an - ll (ta tw) cal/cm .day (5.11) 5.3 Transfer of heat by evaporation or condensation Since the temperature of the air is most often higher than the dew point temperature, the heat lost by evaporation plays a much more important role than the heat gained by condensation. Consequently 05 is generally negative. The main driving force for evaporation is the vapor pressure difference between the atmosphere and the water surface. Several empirical equations have been pro- posed for the relation between evaporation rate and more easily measured parameters. On the basis of the type of equation there are three different groups: 1. Equations based on the vapor pressure deficit and the wind velocity. 2. Equations based on the vapor pressure gradient and the heat or momentum flux. 3. Equations based on the vapor pressure fluctuations according to the Eddy correlation method. The first type of equation has the following general form: m n .- = * * Q Le pw (a + b uz) x e (pw - p&) (5.12) 61 in which Le = latent heat of evaporation at tw (cal/9) tw = surface water temperature (K) p; = saturated vapor pressure at tw (m bar) u = wind velocity at a height 2 above the water (cm/s) x = size parameter of the surface (cm) 9 = density of water at tw (g/cm3) a, b, m, n = parameters, which vary within a small range, depending on the roughness of the surface, the wind speed, the stability of the atmosphere and (except for n) the size of the water surface. The oldest form of this equation is obtained by substituting m = 0 and n = 1. Brutsaert and Yu (1968) brought the result of ten articles together in an equation for which a = 0. Using the Bowen ratio Dingman st 31. (1968) arrived at an equation with m = 0 and n = l. Yen and Landvatter (1970) reported an equation with m = 0. The different parameters from these articles are represented in Table 5.1. Since most of these equations are derived from the researchers' specific equipment, they have a limited range of application. Obviously, the empirical equation (5.12) is dimen- sionally inhomogeneous.‘ In principle it is based on the general equation for mass transfer: H O H O N = k * A * (”1201a - [1420135 (5.13) 2 Table 5.l--Parameter Values for the Equation Based on Vapor Pressure Deficit and Wind Velocity. 3 Author m n a*10 b*10 Carpenter (1891) O l 2.17 .233 Rohwer (1931) 0 1 2.44 .148 Johnsson (1946) 0 0.8 0 2.40 Kohler (1952) 0 l 1.18 .132 Brutsaert (1968) 0 l 1.80 25.3 Brutsaert (1968) .124 .60 0 3.80 Dingman (1968) 0 1 1.08 25.3 Yen (1970) O 1.52 5.17 33.2 in which NH O = mass transfer rate of water (mole/s) 2 kH O = mass transfer coefficient for water (m/s) 2 A = surface area (m2) [H20]a = concentration of water vapor in the air (mole/m3) [H20]; = saturated water vapor concentration in the air at Tw (mole/m3) The mass transfer rate is related to the heat transfer rate via the latent heat of evaporation (Le); the concentration of water vapor is related to the vapor pressure via the ideal gas law. The kH O can be expressed as a function of 2 63 the wind velocity and a size parameter with the dimension- less Sherwood number (Sh): * kHZO X c d Sh = ——-————— = a + b * Re * Sc (5.14) D H20 DH 0 = diffusivity of water in air (mz/s) 2 x = size parameter of the surface (m) Re = Reynolds number (dimensionless) Sc = Schmidt number (dimensionless) a, b, c, d = dimensionless parameters The Reynolds and Schmidt numbers can be expressed as: pa * u * x u a and 0 SC = p *aD (5.16) a H O 2 in which pa = density of air at Ta (kg/m3) u = wind velocity of the main air stream (m/s) pa = viscosity of the air (kg/m 5) Combining the equations (5.14), (5.15) and (5.16) kH O can be expressed as: 2 c-d * c * c 1 * l-d DHZO Pa u x DH20 _ * * kH O a x + b c-d (5.17) 64 Comparing this with equation (5.12) shows, that the parameters m and n are not independent (m + n = l). The value for c is generally between .5 and .8 and the value for d is 1/3. a and b have to be determined experimentally. At very low wind velocities the second part of this equation becomes many times larger than the first part. As a result the first part can generally be neglected. Several equations are based on the vapor pressure gradient and the heat or momentum flux; two such equations are the relation of Thornthwaite and Holzman, as given by Rosenberg (1974): (P -P )(u -u) Q = Le * a * p * k2 * w2 wl 2 l e a 1n (2 /z )2 (5-18) 2 1 - 'IT-T Rb=Qo =61*105*1.'———a*p (5.19) Qe Pw - Pw in which: Pwl' sz = water vapor pressure at two different heights 21 and 22 (Pa) ul, u2 = wind velocities at the corresponding heights 21 and 22 (m/s) kH O = mass transfer coefficient (m/s) 2 Rb = Bowen ratio P = atmospheric pressure at altitude h of the surface above sea level. 65 Use of these equations requires a large number of variables to be measured and requires a correction factor for the stability of the atmosphere. In these equations it is assumed that the Reynolds analogy is valid. The Reynolds analogy describes the transfer of heat with equations simi- lar to the equations (5.13) and (5.17) and assumes that the transfer of mass and heat by conduction is negligible. In addition the coefficients b,c and d in equation (5.17) are assumed equal for heat and mass transfer. This means, that the heat transfer is described with: QO = a * A * (Tw - Ta) (5.20) in which a = heat transfer coefficient (W/mzk) *3 ll temperature of the water *3 II temperature of the air The heat transfer coefficient is related to the wind velo- city and the above mentioned size parameter via the Nusselt number (Nu): a * x d Nu = = b * ReC * Pr Xa (5.21) in which 1a = heat conductivity of the air (W/mK) Pr = Prandl number (dimensionless) 66 The Prandl number can be expressed as: Pr = “5.2 (5.22) in which cp is the heat capacity of air. With the equations (5.13) and (5.20) the Bowen ratio can be expressed as: Q a * A * (T - T ) * R * T Rb = 62 = W a a (5.23) * * * * v - 6 Le Mw k A (Pw PW) in which R is the gas constant (m3Pa/mole K). At an air temperature of 20°C and standard atmospheric pressure the Bowen ratio is: Rb = 5.34 * 10'7 * %— * w a o ‘T‘:“"* P (5.24) 2 Pw PW The values for R, P, Le and Mw are given in Appendix A.2. The value for a/kH O can be calculated from the equations 2 (5.21) and (5.14) by substituting d = 1/3: A d _ a Pr 1/3 2 H2O .A value for Pr can be calculated with the data in Appendix A.2, but can also be found directly in Perry (1973). Sub- stitution of the constants in equation (5.25) results in a value of a/kH o = 1.17 * 103. With equation (5.24) this 2 results in a constant in the equation for the Bowen ratio of 67 6.2 * 10-4 as is given in equation (5.19). The equations based on the vapor pressure fluctuations according to the Eddy correlation method describe the flow of water vapor in the vertical direction as follows: M/M Q€=Le*-—w;——§*Da*W (5.26) in which M = molecular weight w' = instantaneous deviation of the vertical velocity from the time-averaged vertical velocity. p' = instantaneous deviation of the water vapor pressure from the time-averaged water vapor pressure. In order to be able to measure instantaneous temperature, wind velocity and vapor pressure, the instrumentation has to be quite sensitive and responsive. The combination of anemometer, humidity sensor and thermometer give a large quantity of data which make an on-line computer arrangement almost a necessity. Golz §E_al. (1970) find with their instruments a good agreement with lysimeter measurements, although their system still had many problems. 5.4 Transfer of sensible heat Sensible heat is transported to and from the water surface by conductive and convective transport. As described above the contribution of conductive transport is already negligible at very low wind speeds. Johnsson (as 68 cited by Hutchinson (1957))gives the following relationship for the convective transport: 0.8 QO = 4.4 * (Tw - Ta) * u (5.27) in which QO is expressed in cal/cm2 day and u in m/s. Comparison of Johnsson's equation with the equa- tions (5.20) and (5.21) results in a value for b and c of 4.59 and 0.8 respectively. In most cases the convective heat transfer is smaller than the heat transfer by evapo- ration. If, however, the surface is covered by ice the Opposite can be true. The transfer of sensible heat can, of course, also be calculated with the Bowen ratio, making use of the Reynolds analogy. 5.5 The distribution of heat inithe lagoon The transfer of latent and sensible heat depends on the surface temperature and the surface roughness. The surface temperature is a function of the distribution of heat in the lagoon and is affected by mixing. The inten- sity of mixing depends on the wind velocity and the sta- bility of stratification. Mixing is also caused by density currents. Generally stratification in a shallow lake is .neglected and ideal mixing is assumed. Stahl and May (1967), however, report on the temperature distribution in 69 a l m deep rat waste lagoon and in two 1.3m dairy waste lagoons and find temperature decreases of more than 10°C in the top .5m on most summer days. Under otherwise iden- tical conditions the average temperature of a stratified lagoon will be lower than the temperature of a mixed lagoon, since a stratified lagoon has its highest tempera- ture at the surface which increases the loss of heat. Stratification will thus reduce the rate at which a lagoon warms up. After the various coefficients are determined (see Chapter 7) the equations in this chapter would make it possible to predict the warm-up of a particular lagoon under specified climatic conditions. 5.6 Scum formation After the ice cover disappears from the surface, a lagoon will be covered with scum for some time. Subse- quently the scum will appear with several variations until summer when scum formation is at a minimum. Scum has a very important influence on the proliferation of purple sulfur bacteria, since it almost completely absorbs all available solar radiation. The formation of scum needs to be understood. As a gas bubble rises in a lagoon, solid particles are entrained in the wake from the time that the bubble emerges from the sludge layer at the bottom. If the anaerobic digestion processes are close to completion, the solid 70 particles will be high in inert material and low in proteins and fats. In addition, the volatile fatty acids, which are known to reduce the surface tension, are in low concentra- tion. Arriving at the surface the liquid film of the bubble evaporates, the bubble collapses and the solids sink back to the bottom. If, however, the digestion processes are incomplete, the solid particles at the surface can stick together and form a mat until after evaporation of the water content they are dense enough to sink. A high con- centration of volatile fatty acids will promote formation of a solid mat or scum because surface tension is reduced and it therefore takes longer for a bubble to collapse. This results in a higher concentration of solid particles at the surface and consequently increases the probability that the solid particles will stick together. In addition, the longer existence of a solid mat can entrap other gas bubbles which keeps the mat floating. The liquid film of a bubble doesn't evaporate if the temperature of the air is equal to or less than the dew point temperature. On the contrary, water will be added. Bubbles will less easily collapse and will coalesce to very large sizes as can be seen from the picture (Figure 5.2) which was taken on a spring morning when such conditions occurred. Later in the day the temperature rises, the bubbles collapse and typically, a large solid mat is formed. 71 F‘ 10 cm at the center of the photo. Figure 5.2 Scum formation in Spring. 72 Unless there is a strong wind or precipitation, such a mat can cover most of the lagoon for a large portion of the day. I have considered the possibility that low tempera- tures would inhibit the acid consuming bacteria more strongly than the acid forming bacteria which would explain the same phenomena. Volatile fatty acids (VFA) concentra- tions were measured with a gas chromatograph during a day- night period when the scum cover ranged from 5 to 100% of the lagoon surface. I could not detect a change in VFA concentrations. I conclude that the influence of VFAs on the scum formation is more of a seasonal character, while diurnal variations of the scum cover are caused by the changes in relative humidity of the air. 73 mH. no. mm. mm. 5v.m om.H I) (I .z.m OmumH In (I (I II (I I) cm on .z.< omuHH HH. OO. mm. mm. mm.m hm.H OOH OO .z.4 OmuOH OH. mO. HO. hm. mm.~ hw.H OOH OOH .z.€ Omum OH. mO. Hm. mm. om.m mv.H OOH OOH .2.< Omum NH. no. Nb. om. hm.m mv.H OOH OOH .2.< omum OH. (1 OO. mm. mm.m mv.H OOH OOH .2.¢ mvuw (1 HH. (1 mm. (I mm.H OOH OOH .z.¢ mmum m 3 m 3 m 3 m 3 III E\quWII IllmE\quHll .IIImE\oHo&III .ll) w .III OHO< oHuwusm pHo< UHGOHmoum UHod UHumod Ho>oossom .hhmH .Om HHumm co mcoommq on» :H mpHo< Muumm mHHumHo>IIN.m OHQMB CHAPTER 6 THE SULFUR CYCLE AND KINETICS 6.1 Introduction In the Chapters 1, 2 and 3 a description was given of the microbial populations and their interactions with the environment. Since these interactions are very complex, it would be very time-consuming to test all variations in the lagoon situation or even in the lab. With certain simplifi— cations and therefore some loss in accuracy these inter- actions can be simulated on a computer with an appropriate model. In this way other researchers have had great success in revealing details in the behavior of the anaerobic diges- tion process. In paragraph 2.5 a short description of the models for anaerobic digestion was given. These models describe the behavior of acid and methane forming organisms. No similar models exist for the sulfur cycle. Prin- cipal differences with the anaerobic digestion model are: l. The purple sulfur bacteria and the sulfate reducing bacteria cause sulfur to move in a cyclic process, while the carbon path in the anaerobic digestion follows a sequential process from acid formers to methane formers. 74 75 2. The gaseous product H28 is an odorous pollutant and therefore unwanted, whereas the gaseous pro- ducts methane and carbon dioxide are valued pro- ducts of anaerobic digestion. 3. Sulfur is stored in the purple sulfur bacteria, while no storage products are considered in anaerobic digestion. 4. Purple sulfur bacteria are light-dependent, while light has no significance in anaerobic digestion. 5. Almost all models of anaerobic digestion consider the steady state. Because of the storage of sul- fur and the changing meteorological conditions (including the availability of light), the steady state is not applicable. In the following model of the sulfur cycle in the lagoon, I will include all these features. Before assembling the total model I will discuss the different components: a. Description and mathematical formulation of the lagoon. b. Description and mathematical formulation of gas bubbles rising in the lagoon liquid. c. Transfer of hydrogen sulfide between the gas bubbles and the liquid. d. Transfer of hydrogen sulfide from the liquid to the air. 76 e. The microbial conversion processes and growth rates. f. The influence of the pH. 9. The influence of the temperature. h. Changes in the lagoon volume. 6.2 Description and mathematical formulation of the lagoon Most lagoons are designed as trapezoidal structures as shown in Figure 6.1. The cross sectional area at depth h, measured from the bottom, becomes then: Ah = L * W + 2h (L + W ) cot o' + 4h2 cotzd' (6.1) o o o o in which: area at depth h (m2) Ah = L0 = length of the lagoon at the bottom (m) Wo = width of the lagoon at the bottom (m) d' = angle of the side slopes (deg) The volume below the depth h becomes: Vh = LO * Wo * h + h2 (LO + W0) cota' + % h3 cotzo' (6.2) The lagoon liquid is assumed to be ideally mixed. This means that the input of manure is evenly distributed over the whole volume and that the concentrations of sulfide, sulfate, sulfur and microorganisms show no spatial differences. 77 : coommq pwmmnm HmpHoNommuh one H.O ousmHm 78 Further I have assumed, that the urine enters the lagoon via the liquid phase and that the feces are part of the sediment. The feces are degraded in the sediment layer and provide a steady production of gas containing all the sulfur of the feces in the form of sulfide. Since the urinal sulfur is mainly sulfate the urine addition raises only the level of sulfates. At the bottom of the lagoon, then, the concentration of sulfide in the gas bubbles [H28]go becomes: [TS]f * F * N * w [H28]go = O (6.3) 9 in which: [TS]f = the total sulfur concentration in the feces (mole/m3) F = the rate of feces production per unit weight of animals (m3/kg s) N = the number of animals whose manure is trans- ported to the lagoon w = the average weight of the animals (kg) Q = flowrate of the gas (m3/s) 9 fPhe flowrate of the gas can be expressed as: Qg=[C]f*F*N*w*VI (6.4) 4 in which: 79 [le = the concentration of organic carbon in the feces (mole/m3) VM = molecular gas volume (m3/mole) The change of sulfate concentration caused by the intro- duction of the urine is equal to: * * * d[SO4] [TS]u U N w = (6.5) dt V in which: U = the rate of urine production per unit weight of animals [TS]u = the total sulfur concentration in the urine (mole/m3) V = the total volume of the lagoon (m3) 6.3 Description and mathematical formulation of gas bubbles riSing in the lagoon liquid For the following derivations I have assumed that: 1. gas production takes place only at the bottom surface 2. the gas bubbles have a constant equivalent diameter de of 1 cm (see equation 6.17) 3. as the gas bubbles rise, they remain equally distributed over the lagoon area 80 Knowing the equivalent diameter the Eotvas number (Eo) can be determined, which is related to the shape of a bubble: E0 = p“ e (6.6) in which: 9w = the density of water (kg/m3) g = gravitational acceleration (m/sz) de = the equivalent diameter (m.) o = surface tension of water (kg/52) For pure water 0 = 7.275 * lO-Zkg/s2 and the Eatvés number becomes 13.5, at which value the bubbles have the shape of a spherical cap as is drawn in Figure 6.2. Figure 6.2 A Spherical Cap. 81 If the Reynolds number (Re) for the bubble is larger than 800, the drag coefficient (Cd) of the gas bubbles is constant and equal to 0.95. The bubbles will quickly reach their terminal velocity, at which velocity the buoyant force, is equal to the resistance of the water: 1*3* * =.* *2* *£*2 6 de AD 9 5 pw Ub cd 4 60 (6.7) in which: Ub = rising velocity of a bubble (m/s) d0 = the actual diameter of a bubble (m) Ap = the difference in density between bubble and water (kg/m3) This equation can be rearranged to: 3 'k * U = 4 g de b 3 * C * d2 (6.8) d 0 According to Davies and Taylor (1950) and Wu gt_al. (1974): d o 3_'= 3.65 * Fr (6.9) e in which Fr is the number of Froude: Fr = —— (6.10) 82 The equations (6.8), (6.9) and (6.10) give: U: = 4 * d: = 4 * * 'k * * gde 3 Cd do 3 .95 (3.65) Fr As a result, Fr = .47. From equation (6.10) the Ub can be calculated: U = Fr * g * d I = 22 m/s (6 12) b e . . The Reynolds number becomes then: D U d Re = —E——9——§ = 2100 (6.13) u w The value of .95 for the drag coefficient is thus justified. A simple calculation shows, that the bubble will reach a velocity of 99% of the terminal velocity within 1 cm of the point of release. The velocity of the bubble can there- fore be considered constant. The surface of the sphere segment A in Figure 6.2 b is: A = 2n * Rb * hb (6.14) 1 hb=-%V4*R:-d: +Rb (6.15) 83 Davies and Taylor (1950) give for the radius Rb: Rb = % * Fr * de (6.16) By definition of the equivalent diameter the volume of each bubble is: _ 1 4 3 vb — 6 de (6.17) The time a bubble is in the water: ,4 II CID: b (6.18) Combination of the equations (6.4), (6.17) and (6.18) makes the number of bubbles in the lagoon at any time (Nb) a calculable quantity: N = —3————9 (6.19) The specific surface of the bubbles (Sh) will decrease with increasing height in the lagoon, because of the increasing cross sectional area of the lagoon: N * A Sh = H 5‘3““ (6.20) Substitution of N and A (equations (6.19) and (6.14)) b b gives: * * * * S = 99. Th Zn Rb hb h Vb *rH * Ah (6.21) 84 Rearranging this equation and substitution of-rb, Rb’ h b and Vb results in: Sh * Ah 3/2 -3/2 -1/2 Z a _—6—_—_ = 25.2 * Fr * de * g (6.22) 9 This specific surface group, further identified with the symbol Z, provides the surface through which mass exchange between the bubbles and the lagoon liquid takes place. For this mass transfer, which takes place according to an equation similar to the equations (2.5) and (5.13), it is important to know the mass transfer coefficient k This b' coefficient can be found by using the penetration theory of Higbie (1935). This theory can be used, because gas bubbles of the size assumed have an internal circulation, which keeps the concentration of gases uniform inside the bubble. D _ H 3 kb ' 2 2 (6.23) n * T in which: D = diffusivity of H S in water (mZ/s) H25 2 T = contact time of a water particle with the bubble (s) This contact time can be taken as: d r = —3 (6.24) Ub 5 .4 Transfer of hydrogen sulfide As was mentioned earlier, hydrogen sulfide is t3ransported between the gas bubbles and the liquid and from 85 the liquid to the air. Both transfer processes can be described by the equation: = * * t - NHZS kH28 A ([st]w [st]w) (6.25) in which: N = mass transfer rate of H S (mole/s) H28 2 kH S = mass transfer coefficient for H25 (m/s) 2 [H28]w = concentration of H23 in the water (mole/m3) [H28]; = concentration of H25 in the water, which would be in equilibrium with the concentration of H28 in the air or in the bubbles (mole/m3) If the concentration of hydrogen sulfide in the air is neg- lected, this last concentration becomes zero in the calcu- lation of transport of H28 to the air. For this process the transfer area A is equal to the lagoon surface area, which can be obtained from equation (6.1), assuming that the sur- face is smooth. Gloyna and Espino (1969) have determined the mass transfer rate for H S from pond water, when there 2 is no mixing and the water is under an atmosphere of carbon dioxide. They found a transfer coefficient of 2.3 * 10-3 mVs. They claimed that this coefficient increases by a factor of 20 when "slow mixing" was applied by means of air jets. In this last case the transfer of oxygen and subsequent oxidation of H28 could have interfered with their measurements, resulting in an excessive value for the 86 mass transfer coefficient, since the transfer of oxygen is faster than the transfer of H28. (The diffusion coefficient for oxygen is 1.5 times that of H S.) 2 For the transfer of hydrogen sulfide between the bubbles and the liquid the transfer coefficient is given by equation (6.23). Substitution of the values for T and DH S gives a value for k of 2.2 * 10-4m/s. b 2 The equilibrium concentration can be calculated with Henry's law: I = * [H28]w Kst PH S (6.26) This equation is identical with equation (2.4). The concentration of hydrogen sulfide in an as- cending bubble is a function of its height above the bottom of the lagoon. To obtain the total amount of H28 trans- ferred, the transfer rate has to be integrated over the height. The partial pressure in Henry's law can be replaced by concentration using the ideal gas law. As a result, the concentration of hydrogen sulfide in the bubbles at the surface of the lagoon (height H) becomes: [H S] [H 2 ._ W _ [stlgH - 21 + ([HZSlgo' 87 in which: _ * * Zl — KH S R T (6.28) 2 R = gas constant (m3Pa/mole K) T = absolute temperature (K) The value for Z is obtained from equation (6.22). The amount of H28 transferred is then: Qg ([st]go - [stlgH) (6.29) The total rate of H S leaving the lagoon as an odorous air 2 pollutant is the sum of the mass transfer rate at the sur- face and the product of gas flow rate and sulfide concen- tration in the bubbles at the surface ([HZS]gH). 6.5 Microbial conversion processes 6.5.1 Sulfide Balance.--Although in the last paragraph a relation was given for the air pollution, this relation contains still an unknown quantity: the sulfide concentration in the water phase. In order to determine this quantity a mass balance has to be made for the pro- cesses which introduce sulfide and the processes which remove sulfide. Two of these processes have been identified in the previous section (section 6.4). The others are: 1. Conversion of sulfide to internally stored sulfur by the purple sulfur bacteria in the light. 2. Conversion of internally stored sulfur to sulfide by purple sulfur bacteria in the dark. 88 3. Conversion of sulfate to sulfide by the sulfate reducing bacteria. 4. Chemical oxidation of sulfide by oxygen. (The con- centration of oxygen in an anaerobic lagoon is so low, that I will neglect this.) 5. Dissociation of hydrogen sulfide in its ionic form. 6. Precipitation of sulfides with a low solubility. This process requires a complete model for itself. Moreover I don't think that heavy metals play an important role in the treatment of animal waste. Therefore I decided to neglect it. The fifth process will be discussed in the next paragraph. This process is so fast as compared with the first three microbial processes that equilibrium can always be assumed. For the three microbial processes I will assume, that the Monod model is applicable. In many microbial treatment processes this assumption has been shown to lead to a good simulation of practical results. Analogous to the inhibition of methanogenic organ- isms by undissociated acid, mentioned in paragraph 2.5, van Gemerden (1974) reports that the purple sulfur bacteria are inhibited by undissociated hydrogen sulfide, which also serves as a source of reducing power. To express this inhibition, he introduces a function similar to the one used by Andrews. I will make use of the same function: Ll “2" in which: 89 max2 [HZSJW (K2 + [H28]W (l + [H28]W KI (6.30) “2 = the specific growth rate for purple sulfur bac- teria, using H25 as substrate. (l/s) “max2 = the theoretical maximum growth rate for this process without inhibition (l/s) K2 = the saturation constant for the oxidation of H28 (mole/m3) KI = the inhibition constant for H28 (mole/m3) The rate of sulfide oxidation becomes then: 2112:]! = - UZXZ (6 31) dt Y2 ' in which: t = time (5) X2 = concentration of purple sulfur bacteria (cells/m3) Y2 = yield coefficient for the oxidation of hydrogen sulfide (cells/mole substrate converted) The yield coefficient depends on the source of carbon as is explained in paragraph 3.3. The rate of the second microbial process, the reduction of internally stored sulfur, will be determined by the surface area (S) of the sulfur granules: 90 l 2 S * * 'k * 4 n ps NO x2) (6.32) A * [S] * 3 S = 4 * n * N * X o 2 in which: N0 = the number of sulfur globules per cell As = atomic weight of sulfur [S] = the concentration of elemental sulfur in the lagoon (mole/m3) ps = the density of sulfur in the globules (kg/m3) The specific growth rate (u4) for this process becomes then: u *S _ max4 U4 - K + S (6.33) 4 in which: umax4 = the maximum growth rate of PSB using internally stored sulfur K = the saturation constant for the use of inter- nally stored sulfur (mZ/m3) The rate of sulfide production becomes: * 922%.- u (6 34) (it Y4 ° in which Y4 is the yield coefficient for the reduction of internally stored sulfur. The reduction of sulfate depends on the Specific growth rate of the desulfurizing bacteria (pl): 91 * “1 = imafilTSOIIO4lw (6'35) 1 4 w in which: umaxl = the maximum growth rate of the desulfurizing bacteria (l/s) [SO4]w = the concentration of sulfate in the lagoon (mole/m3) Kl = the saturation constant for sulfate (mole/m3) The resulting rate of sulfide production is: * X le S] u 2 w = —l————l (6.36) dt Y1 in which Y is the yield coefficient for the reduction of l sulfate to sulfide (cells/mole of substrate converted). The equation (6.31) is only valid if sulfide is the growth rate limiting substrate. If light is limiting, the growth rate has to be calculated for the available light intensity. The light intensity as a function of the depth can be described by Beer's law: Ih = I * exp (-n * (H - h)) (6.37) with: Ih = light intensity at height h above the bottom of a lagoon with total height H (1x) I = incident light intensity (lx) extinction coefficient (l/m) 3 ll 92 This equation assumes that incident light is normal to the water surface. For the photosynthetic bacteria, let the transition from light zone to dark zone occur at a height hl (less than the lagoon depth H) at which the light inten- sity I is just sufficient to permit phosynthesis: h1 = H + n * log (ll/I) (6.38) From the measurements of van Gemerden (1968a) of the growth rate at different light intensities a relation- ship can be derived. I have put his data on a Lineweaver- Burk plot (Figure 6.3); the convincing straight line makes it possible to determine the growth rate (us) with a Monod-type equation: )4 *I = maxS h (6.39) p 5 K5 + Ih in which: umaxS = the theoretical maximum growth rate for light limited growth K5 = light saturation constant will be the growth rate from the height h (the compensa- u5 1 tion point) up to the level where the combined rate of sul- fide and sulfur oxidation becomes the limiting factor. This Monod-type equation is comparable with the equation for a rectangular hyperbola, which is discussed by Takahashi and Ichimura (1970). I chose for the Monod-type equation, because the constants in this equation are more meaningful. 93 l/u (hr) 25 I 20 ‘ 15 4 o 0 10 J . 5/ x / (klx- ) HIE-4 Figure 6.3 Lineweaver-Burk plot for light-limited growth. 94 Now all the components for a sulfide balance are described. For the equations (6.30) to (6.39) one must supply the model With values for umaxZ' K2, KI' Y2, No’ K4, Y4, u K1, Y1, I, n, I1, umaxS and K5. ps’ umax4’ maxl’ (A complete list of parameters and their values is given in Appendix A.) The values for X [S], X and [SO 2’ l 41w can be calculated with mass balances for sulfur, sulfate and the microorganisms, as will be described later in this chapter. Furthermore, a starting value for the sulfide concentration must be established. 6.5.2 Sulfate Balance.--For the sulfate balance the following components have to be considered: 1. Introduction of sulfate into the lagoon via the urine (equation 6.5). 2. The reduction of sulfate to sulfide by desul- furizing bacteria (equation 6.36). 3. The oxidation of internally stored sulfur to sulfate by the purple sulfur bacteria. The last component is a function of the sulfur surface as is given by equation (6.32). The specific growth rate (H3) for the oxidation of sulfur is: u *S = max3 . (6.40) The resulting production rate of sulfate is: d[SO] u * x __EFJLJ£ = _§____2. (5,41) t Y3 95 in which Y3 is the yield coefficient for the oxidation of sulfur. This coefficient is likely to be three times the yield coefficient for the oxidation of hydrogen sulfide as can be seen from the apparent reducing power in the equa- tions (3.1), (3.2) and (3.4), (3.5). For the sulfate balance an initial sulfate concen- tration has to be supplied. 6.5.3 Sulfur Balance.--Three processes take place with the internally stored sulfur: 1. Formation of sulfur from the oxidation of hydrogen sulfide (equation 6.31). 2. Consumption of sulfur for the oxidation to sulfate (equation 6.41). 3. Consumption of sulfur for the reduction to hydrogen sulfide (equation 6.34). All the components of the sulfur balance are given in the previous paragraphs. An initial sulfur concentration has to be supplied. 6.5.4 Balance of purple sulfur bacteria.--In order to find the growth of purple sulfur bacteria over a certain time span the contributions to the growth rate from equa- tions (6.30), (6.33), (6.39) and (6.40) have to be inte- grated over that part of the lagoon volume, in which they are active. Since ”2 and u3 represent growth based on coupled phenomena their sum is limited by u (Trfiper, max2 1964). 96 An initial concentration of bacteria has to be supplied. Over longer periods of time a death rate has to be taken into account. For the short simulations in this thesis, the death rate has been neglected. 6.5.5 Balance of desulfurizing bacteria.--In this model the growth of the desulfurizing bacteria is con- sidered to depend only on the reduction of sulfate. The specific growth rate is obtained from equation (6.35) and the death rate is neglected. An initial concen- tration of bacteria has to be supplied. 6.6 The influence of pH The pH has a strong influence on the performance of the microbial populations under consideration. In order to keep the complexity of this model within limits, the influence of the microbial activities on the pH and the influence of the pH on the maximum growth rates of the microorganisms have been neglected. The only pH effect which has been considered is in the formation of a sulfide buffer. As is described above, only undissociated hydrogen sulfide can serve as a substrate and inhibiting agent for the purple sulfur bacteria and only undissociated hydrogen sulfide is transferred from the gas phase and to the air. The total available amount of sulfide is, however, the sum of undissociated and ionic forms: 97 - 2- [H28]w + [HS ]w + [S ] (6.42) [H25] w w,total = Using known dissociation constants K' and K'b this can be written as: Kl KI * K" [HZS]w,total = [H28]w {1 + Tfi+1 + IE:T§__. (6.43) The hydrogen sulfide formed, consumed and transferred in the equations (6.25), (6.31), (6.34) and (6.36) has to be corrected by this factor. 6.7 The influence of temperature The maximum growth rates of the microorganisms are functions of the temperature. Below the optimum tempera- ture an Arrhenius type of relationship is displayed. Gloyna (1971) gives the equation: * (1.085)T ' TR “ = “R (6.44) The mass transfer processes are also influenced by the temperature. The temperature dependence of the diffu- sion coefficient, which is important for the mass transfer processes is given by the law of Nernst-Einstein: = constant (6.45) in which “w is the dynamic viscosity. The temperature dependence of the dynamic viscosity of water is: 98 b a + < c + d (e - 20) + e (e - 20)2) (6°45) u = .l * 10 w in which: 6 = temperature (°C) a,b,c,d and e are constants: a = -3.30233 b = 13.01 C = 998.333 d = 8.1855 e = .00585 For short periods of time the temperature of a waterbody can be treated as constant, but for longer time spans temperature variation plays an important role. 6.8 Changes in the lagoon volume For short periods of time the volume of a lagoon can be considered constant. Over longer periods several factors change the volume: 1. The volumetric loading with manure 2. Precipitation 3. Evaporation and condensation (see paragraph 5.3) 4. Pumping of the lagoon contents to the field The volumetric loading rate doesn't require any additional information since it uses the feces and urine production rates as were used in the equation (6.3) and (5.5): dV EE'= (F + U) * N * w (6.47) 99 This amounts to a very small percentage daily addition to total lagoon volume. The precipitation has to be obtained from climatic or meteorological data or can be simulated by a random variable apprOpriately distributed. Precipitation dilutes the lagoon contents. Although it can have an important influence on the surface roughness and therefore on the transfer of hydrogen sulfide to the air, I have neglected these effects on the grounds that the duration of preci- pitation events is not great. The pumping of the lagoon contents causes an occa- sional drastic change in the lagoon volume. For the micro- bial processes, lagoon pump out will act as a step increase in all loading rates and can therefore greatly upset the system. I have not simulated pumping of the lagoon since pumping usually occurs prior to and at the end of the lagoon's May to November operating season. Volumetric changes will thus play no role in this model. 6.9 The total model The total model can best be illustrated by a few diagrams. In figure 6.3 the different processes are represented schematically and in figure 6.4 a flow diagram of the major part of the computer program is given. 100 (#sz3 .oHo>U HDMHsm man we Hope: v.m wusmHm -Nm 2 [m I mmum... W e o... ocHNHmsmHsmop vmmx _ R/ 6%).» oumwpsm :oome mHnouump HamHsm onhso 101 Flowchart of the main loop. I‘ITER EXIT calculate : Zl , rearrange bubble properties , variables . o a. . T calculate air polln. S-an(6.32) s . 0 calculate new L J H255.504.1155 and PSB rate of rate 0‘ calculate all other S oxidation 5 oxidation balmce (merits - 0 J T 1‘ rate Of calculate H25 oxidatior. H..$ and 5 0x. hcr'o chronents 1' 11,5 6:90? V x h )0” ‘ cr ‘ Y N 0 5 0300? S 0100? Y N p H O T o , calculate N (12.0 mm activities light always limiting ( I critical coefficients balances yield light limited hl-Eqn(6 . 38) Figure 6. 5 Flowchart of the Main Loop. PART III EXPERIMENTS CHAPTER 7 HEAT BALANCE 7.1 Introduction In Chapter 5 a theoretiCal model for the heat balance of a lagoon was described. In that model there are a number of unknowns which have to be determined experi- mentally. The most important of these unknowns are the parameters in the equations for the evaporation and con- densation (a and b in equation 5.14). These parameters can be determined by supplying all the data necessary for a complete heat balance. These data are: l. The solar radiation (08) 2. The temperature of the air (Ta) 3. The temperature of the lagoon surface (Tw) 4. The surface area of the water (A) 5. The water vapor concentration in the air ([H20]a) 6. The saturated water vapor concentration in the air at Tw ([Hzolé) 7. The wind velocity (u) 8. The density of the moist air (pa) 9. The temperature distribution in the lagoon 102 103 10. The shape of the lagoon The data for solar radiation were kindly supplied by D. E. Linvill. The data for the air temperature, the wind velocity and the humidity of the air were derived from the monthly reports of the National Weather Service. These data are tabulated in Appendix C. From these data the density of the air is calculated with a psychrometric computing routine. In the following paragraphs I will describe the measurements of the shape of the lagoons and the temperature distribution in these lagoons. Thereafter the data will be applied to the model developed in Chapter 5. 7.2 The shape of the lagoons The heat content of a lagoon is determined by the temperature and the volume or the temperature at each depth and the corresponding volume at that depth. From equation (6.2) one can see, that the volume is determined by the length, the width, the angle of the side slopes and by the depth. In Chapter 4 the design values for these dimensions are presented. In order to know the actual sludge accumulation and the actual dimensions, a depth- profile of the west lagoon was measured. On February 1, 1977, when the lagoons were covered with ice, 21 holes were drilled through the ice as indicated in Figure 7.1. The water depth was measured with a sand filled bottle on a r0pe and the total depth was measured 104 C.) C. -H 9.. H I g r; < x 4‘ g - .1.) -H 3 Input pipe Legend: H Fence X Measuring point Figure 7.1 Water Depth of the West Lagoon. 105 using a metal stick. A measurement was also made of the ice thickness. The results are tabulated in Table 7.1 and the corresponding profiles for the water-depth are drawn in Figure 7.1. Also given in this figure is the shoreline, which was established by measuring distance from the fence- line on April 27, 1977. In Figure 7.2 the approximate lines of equal sludge accumulation are drawn. From these graphs one can conclude that the locations of the inlet and outlet structures of the lagoon have a marked influence on the depth profile. In general the sludge tends to accumulate in the deeper parts of the lagoon. In the immediate vicinity of the inlet and outlet, however, the flow of water scours the sludge. The depth profile of the east lagoon was not mea- sured because the ice was too weak. A single depth mea— surement for the east lagoon was made at one point corre- sponding to point A4 of the west lagoon. Since the values were the same, and the construction of the east lagoon is identical to that of the west lagoon is, it is assumed that both lagoons have similar bottom profiles. The side slopes of both lagoons have been measured in a way similar to that described above for the water depth. For both lagoons the side slope had an approximate 1:3 ratio. 106 Table 7.l--Depth Profile of the West Lagoon. Total Water Sediment Ice Point depth depth depth cover cm cm cm cm A1 191 112 79 48 A2 259 188 71 27 A3 271 216 55 20 A4 282 269 13 14 A5 284 267 l7 17 A6 198 132 56 25 A7 79 38 41 23 Bl 224 165 59 30 B2 272 178 94 32 BB = A4 B4 281 226 55 15 B5 267 254 13 18 BG 272 244 28 22 B7 244 224 20 24 BB 119 102 17 33 Cl 236 188 48 ' 4 C2 = B7 C3 224 178 46 20 D1 271 211 60 20 D2 = B5 D3 263 204 59 20 El 152 124 28 34 E2 = El E3 211 160 51 36 E4 216 168 48 46 107 25cm fllcm 25cm 25cm Figure 7.2 Sediment Depth of West Lagoon. 108 7.3 Temperature distribution At point A4 in both lagoons a float with c0pper- constatan thermocouples was installed. At each float two thermocouples measured the air temperature. From the float a line of thermocouples hung down into the water with a distance of .3m between thermocouples. The thermocouples' millivolt outputs were recorded hourly by an Esterline Angus Data Collection System. These mV recordings con- verted into degrees Celsius with the following equation: °c = 25.86 * (mV - c) - .651 * (mV - c)2 (7.1) in which c is a correction faction. A comparison with the accepted tables (Omega, 1974) shows that this equation gives the temperature within .03°C for temperatures between -20 and 40°C. During the period in which measurements were made, the most reliable results were obtained during the first 20 days of May 1977. This period happens to be the critical period for odor production. With the aid of a computer plotting routine a series of bi-hourly measurements for air, surface and bottom temperatures are presented in Figures 7.5 - 7.8. Figure 7.5 shows that in the beginning of May the water temperature of the east lagoon has a value equal to the average daily air temperature. One can also deduce that little or no stratification occurs in the east lagoon and that the bottom temperature is almost constant. Figure 7.3 Installation of Temperature Measurement System Figure 7.4 Temperature Measurement System in Operation. ('3 ZH NWCHIPFUNWEMH 4O 30 20 10 00 Figure. 7.5 Temperatures in the East Lagoon from 110 1 : AIRsoo*oo . SURFACE= --#-- : BOTTOM= --+-- "f .* O *.. 4° . : #3 .° ’3 . . °. : .# ,. *H 5”,, ..# #2" #. 9‘. ‘+ :. .. #.1.## . ##°## ++~+1 2+: -h+dfifi+4HP+ -tf+zv+4f{+4u++4wr ' 3 3 it; :*a k. . . *0 0* * TIME IN HOURS May 1, 9 P.M. to May 4, 8 A.M. ZH NWCwaMWZMF-i O 111 i 40 -) 4:. ‘4: :*. o - "5 : :1 :6:- o*.* .* : .0. 0 °*. '. .0 0. *. 30 - : o. * '. f . . 0": : z . O. >1 .##... ’ #5“. ' *3. # - #.- - j} if! ,4 . 4; # 1, (I .# #'# 3 ' ’4: ’9' # if!” '. 3' ##i“#£ # 3 I * 20 - {I} ' a .0 ° 2 ; + ' :++ ++ &. A - -F H~+$ * + . a o+++ #8 +++ +.++ + ++ +.°:.'t :57‘ .0 2;. at i: 10 - 1 """""""""""" : : AIR: 0 0*. a ' 1 SURFACE= H .5” z i : BOITOM= ~+-- . p _________________ 7 00 r."--"“'. """"" . ------ r ""“-7"""". """""" . 0 12 24 36 48 6O 72 TIME IN HOL’RS Figure 7.6 Temperatures in the East Lagoon from May 18, 9 A.M. to May 20, ll P.M., with a Three Hour Interruption at the Thirtieth Hour. ZH MWCH>WWVZNH O 40 30 20 10 112 l : AIRgaa*aa : . SURFACE: --#-- ; : BOTTOM-t --+-- . 7 0*.*. a k . 10#..# .0 # ' -# f’ 1" .# . .# #1” ', #. y). # g £.# #0. .#O.;. * # #.# #. ' o# a # # o# *. z # .0 0. #, 'o# °*.+ ++++E+°+41++.+.++E++++'+’+-.t++'+++‘°..:+‘H’°+ ' 3 ° 5 *-. * a o. . a. . 0*. TIME IN HOURS Figure 7.7 Temperatures in the West Lagoon from May 1, 9 P.M. till May 4, 8 A.M. 0 2H MWCH>WNWEWH 40 30 20 10 00 113 4 1 4'. a *. '5 .* * -' ° , ** 3 . i f 4' , a: °. ; °. *.' 3 .: 1 3 E 3 * * ? {I 5 #. 3 o. : a. 3.- : #° 1... .1“ a #. ' o ' ° # . j 0*. 1 * '# # g # # #. #°# .# £.t. # # #.:#'# i“ 1} * 7 fl 2 {a .# fl ' J' ', : :P’++§ + . +. .+++ # .$++£h+fi+fi ¥ 3 .++ + .434"+ '{ . +° '5 + k I _ o; g ----------------- I : AIR==--*-- : v SURFACE = ~-#-- g 3 BOTTOM = - -+- - . ) p _________________ ' ‘T"""'1 """" F"""'T"""’T"”"’1 """" 1 0 12 24 36 48 60 72 TIME IN HOUR Figure 7.8 Temperatures in the West Lagoon from May 18, 9 A.M. till May 20, ll P.M. with a Three Hour Interruption at the Thirtieth Hour. 114 It is important to note that the bottom temperature is high enough to allow substantial microbial activity. Com- parison of Figure 7.6 with Figure 7.5 shows that during May 1977, the bottom temperature increased only slightly, while the surface temperature remained close to the daily average air temperature. Consequently temperature differ- ences of five to ten degrees developed between the surface and the bottom. In the beginning of May, the west lagoon (Figure 7.7) has the same surface temperature as the east lagoon, but a lower bottom temperature. The west lagoon thus shows a more stable microstratification. This difference can be explained by the higher loading rate which was applied to the east lagoon, causing greater mixing, and consequently a reduced heat loss, as is explained in Chapter 5. At the end of May this difference was no longer as pronounced. The temperatures in the west lagoon were then slightly higher than those in the east lagoon. Because of the above mentioned higher loading rate, the east lagoon developed a more extensive scum cover than the west lagoon during the month of May. Consequently, temperature profiles can possibly be explained by a larger heat loss in the east lagoon as result of the greater sur- face roughness. Scum thus seems to increase the heat loss or reduce the heat gain or both. 115 To further verify this I calculated the heat input and output of the two lagoons, except the heat lost by eva- poration. I then set the heat lost by evaporation equal to the difference between the net heat input to the lagoons and the change in heat content. The heat content of the two lagoons is plotted in the Figures 7.9 and 7.10. A linear regression, which I subsequently performed on log (Sh) against log (Re), resulted in the following values for the parameters b and c: (equation 5.14) b c East 1070 -.65 West 1.29 .32 The two linear equations are different with a 90% confidence (N = 32). The differences in b and c show, that not only are heat losses greater for the east lagoon, but also the heat loss increases with decreasing wind velocity. This phenomenon can be explained by the scum removal action of the wind. 23H HZMHZOO HCDF'JL'C CO\CH,'E: "7 H 100 80 60 40 20 00 116 0* *. * .* * *0*0* * * * *0 * *. .* *9 . *. 4; #°#'. .#.#.*'* * * *'*..# # if #1; 1; 7 7.; **** “i; ###. ~#"#‘ # . . # f# #)¥#. {f # TIME IN HOURS Figure 7.9 Heat Content of the Two Lagoons From May 1, 9 P.M. to May 4, 8 A.M. D--------------------------------------—------- EH HS'St'ft-‘JLSOD H3>F1CS nxu: 2C: 100 80 60 40 20 00 117 d :1!“ # .* .*.* _:.*#;;* n .n #‘*°-.*5.# * ° . .*; ' #i. .3 * ° *3 w *' *- H # .* ' # 3 '11:?! fr: ' . J; # .. . . " * fl. # .# Ofl.‘ # # q ---------------- ! : EAST= * i . WEST= #o- ; _ : ' '7"’-"-'1 """"" r“'---'r-"-'--7----'-'1 ------ . o 12 24 36 48 60 72 Figure 7.10 TIME IN HOURS Heat Content of the Two Lagoons from May 18, 9 A.M. to May 20, 11 P.M., with an Interruption at the Thirtieth Hour. CHAPTER 8 KINETICS 8.1 Introduction In support of the model which I described in Chapter 6, I have conducted a series of experiments. In the following paragraphs I will describe the experimental procedures, the results obtained, and the simulation of these experiments with the model. The goal of these experiments is to determine at what rate the purple sulfur bacteria perform the conversion of hydrogen sulfide to sulfate with the intermediate storage of elemental sulfur. They are intended to show the influence of various tempera- tures and light intensities in the chemical and microbial environment of an anaerobic swine waste lagoon. 8.2 Experimental Procedures Late in the afternoon, test tubes were filled with liquid from the purple west lagoon, which is described previously. In order to avoid contact of the lagoon liquid with air, the tubes were filled and closed with rubber stoppers under the surface at about the center of the lagoon. The outsides of the tubes were cleaned with distilled water to prevent formation of light-absorbing 118 119 spots and the tubes were placed in the dark as soon as possible. One hundred of such tubes were divided into four groups and exposed to light intensities of 11, 75, 205 and 540 lux after an incubation period of about 14 hours in the dark. The light and temperature regime were provided by a Sherer environmental chamber (model CEL25- 7HL), which is equipped with 12 frosted incandescent light bulbs of 25 W each. The spectral distribution of the incandescent light source is given in Figure 8.1. This spectral distri- bution can be approximately described by the black body relationship: 2 2 E = 2n h'c' n' (8.1) A5 (exp(h'c'/k T)-l) in which: E = Radiant energy at a wavelength (W/m2 - m) h' = Planck's constant (J-s) k = Bolzmann constant (J/K) c' = Velocity of light in vacuo (m/s) n1 = Refactive index of an emitter T = Absolute temperature of the body (K) A = Wavelength (m) The spectral distribution of Figure 8.1 results in a value of 2.89 - 1034 (including an equipment correction factor) for the group 2n hpc.2n,2 and a value of 5.86 - 10-7 for the group h'c'/kT. These values go into equation 8.1 to 120 .mousom ucmflq uncommpcmocH wcu mo :oflusnfluuman Hmuuommm H.m wusmwm zo— rL P.— ».LLLL.IrIFL .P {P Dem? PL comm J J1-— a 1.1 « \II\|\|\‘ \\ h FF LL 5 mqu »Ic_; ppcz on .t «In _H|IHLLIU|rVHnw\..L.4 ‘I‘I‘ o. 121 provide a value for E which can deviate 25% for wavelengths of about 4 - 10'-7 5 - 10-7m. Thus, the bulbs have a black body temperature m and 0.3% or less at wavelengths about of 2455 K. Since the 12 light sources are equally distributed over the ceiling of the 0.55m by 1.25m environmental chamber, the test tubes were placed in a horizontal position at a distance of 0.9m from the light sources. The various light intensities were obtained by covering the test tubes with stretched layers of black nylon netting, which were fixed on a frame. For the light intensities of 540, 205, 75 and 11 lx, I used respectively 0, 8, 16 and 32 layers of netting. Three temperatures were used: 10, 20 and 25°C. Every four hours, the series were sampled for sulfide and sulfate analysis and occasionally for sulfur analysis, microscopic examination and microorganism counts. Total sulfide was analyzed with a specific ion electrode as described in Appendix B.l. Sulfate was analyzed gravimetrically as described in Appendix 3.2. For the analysis of sulfur the cells were centrifuged and extracted with ethanol. The extract was transferred in the dark and the absorption spectrum was measured with a Cary double beam spectrophotometer. As described in van Gemerden (1968a), the solution of sulfur in ethanol absorbs at 260 nm. Since the absorption peak for sulfur ‘ 122 has to be corrected for the presence of bacteriochlorophyll and since organic contaminants from the lagoon liquid con- tribute to the absorption the analysis of sulfur is not very accurate in my situation. The results are therefore only used as an indication. Occasionally, I made counts of purple sulfur bacteria with a Petroff Hauser counting chamber, but since the lagoon liquid was often contaminated with particles, the results are somewhat inaccurate. I counted desulfurizing bacteria with a most- probable-number technique, using the medium of Baars as described by Pankhurst (1971). As reducing agent, I used titanium (III) citrate instead of sodium thioglycolate (Zehnder and Wuhrmann, 1976). In order to obtain a more accurate description of the oxidation of sulfide I made duplicate measurements at 25°C and 540 1x, sampling the cultures every two hours. The data from these measurements were later used to support a computer simulation. At the end of Fall, 1977, I set up an experiment to determine the maximum sulfide removal rate at 540 1x, since none of the previous experiments had sulfide con- centrations which were high enough to provide this infor- mation. For this experiment, the test tubes were filled with lagoon water from a bucket, to which various quanti- ties of a sodium sulfide solution were added. 123 8.3 Results and Discussion Microscopic examination indicated the presence of Thiocapsa rose0persicina as the dominant species: The cells contained globules which were light in the center and a dark ring around them, characteri- stic for sulfur globules. The cells were round, often occurring as diplococci. I could not detect any motility. India ink stain indicated the typical capsulated structure. Supporting evidence was found in the spectral measurements: Ethanol extracts gave a peak at 775 nm, which is characteristic for bacteriochlorophyll a. Cells suspended in a sucrose solution gave peaks at 370, 480, 511, 547, 587, 670, 798, 855 and 895 nm. Takacs (1971) gives for Thiocapsa roseOpersi- cina cells absorption maxima at 375, 495, 515, 550, 590, 800, 850 and 890 nm. The lagoon samples showed a sharp decrease of the H28 concentration in the light, while the concentration increased slightly in the dark. This shows the presence of a photosynthetic sulfur bacterium. The concentration of 8 these bacteria was about 10 cells/ml at the height of the bloom (September 1977). 124 T. roseopersicina are immotile. Turbulence such as that generated by the wind keeps these bacteria in suspen- sion in a lagoon. In test tubes, no such mixing is avail- able; as a result, cells tended to settle during the course of a day. Some initial longer experiments (not reported) showed a compaction of cells at the bottom of the tubes. Most of these experiments were carried out during Fall 1977. At the end of this testing program, the cells taken from the lagoon showed a tendency to settle faster in test tubes. Although cell counts in the lagoon were similar to the counts at the beginning of Fall, the activity of the cells was much lower at the end of Fall. This is probably caused by a reduction in the number of viable cells as a result of the rapidly decreasing temperatures (see also Figure 8.8). The results of the sulfide and sulfate measurements for the experiments with various light intensities were presented in Tables (8.1, 8.2, 8.3) and Figures (8.2) through (8.7). At 11 lx, no significant removal of hydrogen sulfide occurred at any of the three temperatures. This is in agreement with a previously-mentioned compensation point figure of about 10 1x as reported by Takahashi gt El- (1972). At 75 1x the rate of sulfide oxidation seems to be almost independent of the temperature: the light intensity appears to be the rate limiting factor. For an unknown reason, the rate at 10°C, as measured, was higher than at the other two temperatures. All three temperatures show an mm. «a. mm. mm. cam Hm. mm. mm. mm. mom am. pm. me. an. mm ma. . so. me. so. as .z.m m .z.m v 2002 .2.m m muflmcmucH usmfiq 125 AmE\mHOEV mafia powwowpcfl um Guamasm mo mcoflumuucmocoo om. mm. mv. hm. ovm om. Nv. am. pm. mom om. mv. mm. mm. mm mm. mm. cm. om. Ha .z.m m .z.m v cooz .2.4 m mufimcwucH named AmE\mHoEv mafia cmumoflch um mcwmaom mansaom mo coflumuucmocou .mmfluflmamucH unmflq nsom cam oooa um wcflmasm cmmoucmm mo coaumowxonna.m wanna 126 vm. hm. om. mm. ovm mm. as. mm. we. mom Ne. ma. mm. om. mu am. mm. av. mm. Ha .z.m m .z.m a 2002 .z.m m muflmcmucH unmfla AmE\mHoEv mafia pmumoflpcfi um mummasm mo mcoflumuucmocou NH. 5H. mm. «m. cam am. mm. ms. mm. mom as. me. am. am. ms mm. mm. mm. hm. dd .2.8 m .2.m v cooz .z.¢ m muflmcmucH unwed AmE\wHOEV mafia wmumoflpcfl um mcwmasm mo mcoflumuucmocoo .mmHUHmsmch ucmfiq Hsom can Doom um mpflmasm cmmouoam mo coflumpflxOII~.m wanna 127 mm. mv. mN. ovm mh. NV. 5N. mON mw. 0%. EN. m5 Nv. mm. mm. HH .z.m omum .z.< canfifl .2.< m suflmcmucH unoflq AmE\wHoEV mafia Uwumoflpcfl may um mummasm mo mcofiumuucwocou MH. mm. mm. Ovm vm. mv. vm. mON me. am. am. m5 Nm. mm. mm. HH .z.m omum .2.¢ oauaa .z.< m muflmcmucH unmflq Am E\0HOEV mafia pmuMOflocfl mnu um prmHsm mo mcoflumuucmocou .mwfluflmcmucH unmflq usom 0cm comm um onwmasm cmmoupwm mo cowumvflxOIlm.m wanna SULFIDE CONCENTRATION (mole/m3) 1. .128 H 11 1x H 75 1x H 205 lx H 540 1x TIME (HOURS) 04 .81 .5 f + \\. .4: .2i 0 v . fi 12 16 20 Figure 8.2 Oxidation of Hydrogen Sulfide at 10°C and Four Light Intensities. SULFATE CONCENTRATION (mole/m3) 129 H 11 1x A——I ‘75 1x H 205 1x H 540 1x .o' X x .81 . .6‘ A O .44 .21 0 T I u 8 12 16 20 TIME (HOURS) Figure 8.3 Formation of Sulfate at 10°C and Four Light Intensities. SULFIDE CONCENTRATION (mole/m3) o (D o 0‘ 130 H 11 1x ‘. " 75 1x )9—1K 205 1X H 540 1x 12 16 TIME (HOURS) Figure 8.4 Oxidation of Hydrogen Sulfide at 20°C and Four Light Intensities. SULFATE CONCENTRATION (mole/m3) 131 . . 11 1x ‘-1I 75 1X X-—X 205 lx '_.. 540 1x 1.0‘ .81 .6 ‘ _“_____¥ «1 N TIME YOURS) Figure 8.5 Formation of Sulfate at 20°C and Four Light Intensities. SULFIDE CONCENTRATION (mole/m3) 132 . . 11 1x H 75 1X H 205 1X H 540 1x .0‘ .8d .61 —fi .41 .2‘ 0 v . h 8 12 16 20 TIME (HOURS) Figure 8.6 Oxidation of Sulfide at 25°C and Four Light Intensities. 3) (mole/m SULFATE CONCENTRAT I ON 133 O—-O 11 1x ‘-I. 75 1x X——X 205 1x H 540 1x .0‘ .81 .6‘ .41 ///”,fl¥ .2‘ 04 , _ . 8 12 16 20 TIME (HOURS) Figure 8.7 Formation of sulfate at 25°C and Four Light Intensities. 134 .A.z.m mum um amusmwmev puma «0 Hana may ocwusn coommq ummz mcu :fi mvflmasw cmwouvxu mo aofiumuucwocou m.w musmflm umnouoo HmnEmuQmm umnms< Hm ma H ma H mH H a E A V .A 1 P I. u T: D. 1 I I co 0 (Em/atom) NOILVHLNHDNOD 3013103 .135 increasing rate of sulfide oxidation with time. This is especially clear for the 20°C measurements (Figure 8.4). This can likely be explained by an adaptation of the purple sulfur bacteria to the low light intensity, resulting in an increase in concentration of bacteriochlorOphyll. At 205 1x, the rate of sulfide oxidation shows a dependence on the temperature between 20 and 25°C, re- sulting in a higher rate at 25°C. Between 10 and 20°C no significant change is apparent. As at 75 1x, so also at 205 1x a slight increase with time in the rate of sulfide oxidation appears. The increase in rate is with time, however, less pronounced at 205 1x. At 540 1x a strong dependence of the rate of sul- fide oxidation on temperature shows up. Comparison of the three figures (Figure 8.2, 8.4, and 8.6) reveals that below 0.3 mole/m3 the sulfide concentration is a limiting factor in the oxidation of sulfide. No influence of desulfurizing bacteria is apparent from the sulfide measurements. The increasing rate of sulfate formation at 540 1x, while the rate of sulfide oxidation decreases, can probably be ex- plained by the increasing reliance of the PSB on the oxi- dation of internally stored sulfur. The concentration of sulfate at 8 A.M. is surprisingly low for the 25°C measure- ments, but almost exactly the same for the different samples. The variation in samples as obtained from the lagoon is not likely to be the cause of this. In Figure .136 8.8, I have plotted the sulfide concentrations in the lagoon during the period that these measurements were made. All samples were taken between 5 and 6 P.M. The average values of duplicate measurements of sulfide and sulfate concentrations for 25°C and 540 1x are presented in Table 8.4 and Figure 8.9. The slight increase in the sum of sulfide and sulfate concentration could have been caused by oxidation of internally stored sulfur. At the end of the afternoon, the sulfate concen- tration shows a decrease, possibly caused by an increase in the number of desulfurizing bacteria as a result of the high sulfate concentrations. As I mentioned previously, these data were used for a computer simulation. In order to permit an accurate comparison between data and computer simulation, I have plotted the data with the computer plotting subroutine (see Figure 8.10). For the simulation of this experiment, I simplified the model presented in Chapter 6 by removing all terms which involved the addition of feces and urine, the action of gas bubbles and the release of hydrogen sulfide to the air. Basically, the data as supplied by Appendix A were used. The initial measured values for sulfide, sulfate and microorganisms were supplied. Based on the change in the sum of sulfide and sulfate, I estimated the concentration of internally stored sulfur as .05 3 . mole/m . Assumed values for “max4 and Y4 were supplied, 137 Table 8.4--Oxidation of Hydrogen Sulfide at 25°C and 540 1x. Time [H28] [S04] [H28 + 504] 8 .47 .16 .63 9 .27 .28 .55 10 .22 .46 .68 12 .11 .58 .69 14 .05 .66 .70 16 .04 .68 .72 18 .04 .65 .69 20 .03 .62 .65 CONCENTRATION (mole/m3) 138 H = sulfide H = sulfate 0.8+ . I = sulfide + sulfate If /I/(' \:\ h 0'61 ll 0.44 O 0.2j O 2.;1 1__ O o ' ‘ - 3 12 16 20 TIME (HOURS) Figure 8.9 Oxidation of Hydrogen Sulfide at 25°C and 540 lx. 2H ZOHH>wHZmozon Z Gn\mfioz 139 1.0- , ------------------- ' : 325: ..*.. : ; $04=~#-- ; ' H28 + 8042...}... ' g ___________________ 1 0.8- ........... +. 00.... . 4' ...... +00’° T 0004;0’0. +.. 0 -'° "°-#--.. + In ..... 0.6~ ., 1;, i, + .' 1'. if" 0.4-1'3 0"... O 0* 0.2-.o ' fl 0*. * °°°°°°° m ....... i . a 0.0-' """"""""" r"""'-'------7 ---------------- , 8 12 16 20 TIME IN HOURS Figure 8.10 Oxidation of Hydrogen Sulfide at 25°C and 540 lx (plotted by computer subroutine). 140 although they appeared not to be important, since at a depth of 2 cm the light intensity is still above the compensation point (for I = 540 lux). The computer simulation calculated rates of micro- bial processes which were many times too high. So I ad- justed a few parameters based on the following assumptions. 1. The number of viable cells of purple sulfur bacteria is less than the direct count. 2. Under natural conditions a certain substrate is likely to be in a less accessible form as it is in a laboratory medium. This will result in an in- crease of the saturation constants. 3. The value for Y2 (the yield coefficient for the oxidation of sulfide) which is unknown, is likely to be close to 1/4 of the yield coefficient for the desulfurizing bacteria, since l/4 of the electro- transfer takes place. The formation of storage carbohydrates will temporarily lower the yield coefficient. The net effect is probably negligible since the yield coefficient is higher when storage carbohydrates are converted to structural cell material. The best fitting simulation, as is plotted in Figure 8.11, was obtained by substituting the initial microbial concen- trations of 1012 and 1010 for respectively purple sulfur and desulfurizing bacteria and K2 = .1 and K1 = .15 mole/m3. 3 CO\mHoz ZH ZOHH>szmozoo 141 1.0- , ------------------- , : 325: ..9:.. : t 804=:-'#-- : 1 H28 + 304= ..+. . ' ___________________ I 0.8-1 + + +o..+.oo+oooa_...+...+. + + (,, . - ...)... ... ... 0 61 + 4' ..#° .7} 1 1; 1; #o .1}. O‘ a # .01r i A)" 0.4- . .#.. OK . ..# 0*. 0.2-..' . i) *- .*. k . * * * -*- k * '* 0.0.; ----------------- . ---------------- , 8 12 16 20 Figure 8.11 TIME IN HOURS Computer Simulation of the Oxidation of Hydrogen Sulfide at 25°C and 540 1x. 142 As was noted above, the only effect apparent on the graph (Figure 8.9), which could possibly result from the desul- furizing bacteria would be the lowering of the sulfate concentration at the end of the afternoon. In addition to this, the desulfurizing bacterial processes probably affect the exact shape of the curves, but their influence is minor. As far as the computer simulation is concerned, the cell concentration of 1010 and the K1 of .15 need not be very accurate. The K2 of .1, however, appears to be quite critical. The results of the measurements made to obtain the maximum hydrogen sulfide oxidation rate at 25°C and 540 1x are presented in Table 8.5 and Figures 8.12 and 8.13. A comparison with Figure 8.9 or 8.6 shows, that the maximum rate of sulfide oxidation must be decreased. This could possibly be caused by a reduction in the number of viable cells which I postulated previously. The low point in the concentration of sulfate (Figure 8.13) is a bit strange. A possible but unlikely explanation is that the temperature in the lagoon liquid is very low, resulting in a reduced microbial activity. By introducing the liquid into a 25°C environment the desulfurizing bacteria could have become much more active, resulting in a decrease of sulfate and a slight increase in hydrogen sulfide. This is however, not in agreement with the rest of the curves. Therefore, it remains an unexplained phenomenon. 143 mm. Hv. mH. cm. hm. m mm. mm. mm. mo. mm. v mm. 5w. vm. mH. mm. m we. mv. om. mo. Hm. N we. mq. om. mo. em. H .2.m m .2.m v COOZ .2.¢ omum .Z.¢ w wHHmm AmE\wHOEV meHB UmumoHUcH um ouMMHsm mo mcoHumuucmocoo mH. mH. ow. mm. mm. m om. mm. vm. mm. mm. v mm. mm. mm. mm. mm. m mm. mm. mm. No.H MH.H N me. vs. oo.H mo.H m~.H H .S.m m .2.m v :002 .Z.¢ omum .S.< m mHHmm AmE\oHOEV mmEHB UmumoHocH um mUHMHsm mo mcoHumuucwocoo .mcoHumuucmocoo mcHuHmum ommmmuocH nufiz xH own new xH oomm um mcflmasm cmmounmm mo coflumcflxonum.m mHnme 144 m.m mMSOHm mom « Rome on. Amos mm. oooH AmHC so. isms ma. oomH Ammo NH. lame pH. ooom Ammo ma. n oomm 103 MO. Ame so. Ame «o. AHHV mo. 8 oomm .z.m m .z.m o .2.8 a .z.m omum .2.m m laws we. Ammo hm. AOOHV om. oooH ~H\OH Rome em. Amsv mm. local as. oomH «\m lam. mm. AOOHV em. ooom m\oH Ammv mm. AOOHV mm. n oomm -\m Ammv HH. Ahab mm. isms em. iooav sq. m oomm smxm cooz .2.¢ oHHHH .2.« 0H .2.4 m .z.< m musumummsme «mama AHMHuHcH mnu mo w on» mHmmxucwumm cmmzumn CHV mmEHu omumoHUGH um wwHstw mo coHumuucmocou .mmusumummEmB unom can xH ovm um mpHMHsm cmmouoxm mo :OHHMpHxOIIo.m 83$ SULFIDE CONCENTRATION (mole/m3) 145 . .0 u .8 . I O .61 . C3 .4 I o .2- J D I: O u 8 12 16 20 TIME (HOURS) Figure 8.12 Oxidation of Hydrogen Sulfide at 25°C and 540 1x with Increased Starting Concentrations. SULFATE CONCENTRATION (mole/m3) O (I) L O 0‘ A 0 sh .21 '1 146 Figure 8.13 12 16 20 TIME (HOURS) Formation of Sulfate at 25°C and 540 lx with Increased Starting Concentrations of Hydrogen Sulfide. 147 Figure 8.14 Oxidation of Hydrogen Sulfide at 540 1x TIME (HOURS) and Four Temperatures. H 10°C 10/12 H 15°C 9/4 H 20C 10/5 l—l 25c 8/27 g—a 25c 9/22 100 f: (U .H \ :31 80-1 \ c -H “-1 0 up V 60‘ A g D H E S 40‘ m U 8 o a g 20" H In H D U) o T ' 8 12 16 20 PART IV DISCUSSION CHAPTER 9 MANAGEMENT IMPL I CAT IONS In this chapter I will discuss various management criteria which are important in the proper operation of an anaerobic lagoon and how these management criteria are influenced by the purple sulfur bacteria. Some of these management criteria will be the same as the ones for anaerobic digestion since the basic anaerobic treatment processes, as described in Chapter 2, are the same. Some other management criteria are related to the specific properties of an anaerobic lagoon and to the specific needs of a farmer. Lagoon design criteria will also be affected by the decision to foster purple sulfur bacteria in the lagoon. The loading rate is an important factor for lagoons as well as anaerobic digesters. Since a lagoon is not equipped with specific devices which control theenviron- ment and since a lagoon is a semi-batch system, from which the contents are removed about twice a year, the loading rate, as based on volume and quality of organic material per unit lagoon volume, has to be much lower than for an anaerobic digester. Similar to the anaerobic digestion 148 14'9 loading has to occur on a regular and frequent basis, (i.e., once or twice daily) to prevent conditions which upset the microorganisms. A farmer, who empties the contents of a pit into a lagoon, asks for problems. In Chapter 4, I reported that the lagoons at MSU were designed using the figure of 0.062 m3/kg of animal weight. This is equal to the loading rate recommended by Smith and Miner (1975) and equivalent to 0.077 kg volatile solids (VS) per cubic meter (0.048 kg VS/kg animal daily manure production). In the presence of purple sulfur bacteria this loading rate does not lead to malodorous conditions. In the design of the lagoon system at MSU, 200 m3 were included to account for a three years' sludge accumu- lation. In Table 7.1 the sludge accumulation is given as measured at the beginning of 1977. From this table I esti- mate that the total sludge volume is slightly more than 200 m3. The design value for sludge accumulation appears to be correct. This value is twice the volume given by Smith and Miner (1975) (i.e., 20% of the design volume). The lagoon system at MSU consists of two identical lagoons. In Chapter 4, I described some management features which I tried out on these lagoons and which proved to be successful. I took advantage of the existence of the two lagoons by using one of them to preserve a favorable bacterial culture during the winter. Since the purple sulfur bacteria were able to survive anaerobically in the 150 refrigerator for more than two years, it seemed possible to obtain winter survival at culture densities that would be immediately beneficial for Spring odor control. I reduced the amount of organic material going to the purple sulfur bacteria by diverting the winter accumulation mostly to one lagoon. By pumping the most heavily loaded lagoon in the Spring, the purple sulfur bacteria in the other lagoon were preserved in a condition in which they could immediately photosynthesize in the Spring. Thus the odorous Spring episode could be avoided. The reduction of organic material can also be obtained with a single lagoon system by pumping the lagoon down to a low level in Spring and filling it partly with fresh water. The latter procedure results, however, in a larger loss of the bacterial popu- lation. The preservation of the bacterial population in the two-lagoon system can be compared with the increase in solids retention time (i.e., microorganism retention time) which is obtained in two-stage digestion and the activated sludge process by separation and recycling of solids (i.e., microorganisms). In my opinion, the best time to pump a lagoon is the time that a noticeable increase in odor occurs. At this time the bacterial activity has substantially increased and as a result the concentration of suspended solids will be very high as compared with other times. The operator can select his/her pumpout time to take advantage of wind velocity and direction. 151 Figure 9.1 Pumping the East Lagoon in Fall 1977. 152 The purple sulfur bacteria are considered ubiqui- tous. In principle, therefore, inoculation is unnecessary. As described in Chapter 4, the purple sulfur bacteria don't seem to be able to cope with an increased loading rate if their number is limited. This means that a bloom of purple sulfur bacteria will only survive in a lagoon when they get a chance to develop in sufficient numbers. In marginal cases, inoculation of a lagoon with a culture can make the difference between establishment or failure of a population of purple sulfur bacteria. 153 Conclusions 1. Once purple sulfur bacteria have established them- selves in large numbers in a lagoon it is possible to increase the loading rate of this lagoon to a certain extent without causing malodorous conditions. A method of increasing the microorganism retention time has been successfully applied to an anaerobic lagoon system. The MSU swine waste lagoon system appears to be working according to the design; the 1972 design criteria are satisfactory for a relatively odor— free anaerobic lagoon. It is possible to operate an anaerobic lagoon in Michigan's cool-wet climate, without odor problems by following simple management guidelines and proper design criteria. A froth-type scumcover on a lagoon is likely to increase the heat losses. Changes in sulfide and sulfate concentrations in a mixed lagoon culture under laboratory environ- mental conditions are a repeatable phenomenon which can be modeled and predicted. In Fall the rate of sulfide oxidation decreases resulting in rising sulfide concentrations in a lagoon. This is considered to be caused by a 154 decrease in the number of viable cells of purple sulfur bacteria. Diurnal temperature fluctuations are not seriously affecting bacterial activity. APPENDICES APPENDIX A CONSTANTS AND PARAMETER VALUES APPENDIX A-l Table A-l--Conversion of Units (Weast, 1976L From To Multiplication Factor Acres m2 4046.8564 Atmospheres Pa l.01325*105 Btu J 1054.18 Btu/lb J/kg 2324.44 Calories J 4.184 Centipoise kg/ms 10"3 Cubic feet m3 0.028316847 Feet m 0.3048 Foot-candles 1x 10.763910 Gallons m3 0.0037854118 Inches of Hg Pa 3386.39 Knots m/s 0.514444 Lamberts 1x 104 mm of Hg Pa 133.3224 Phots 1x 104 Pounds kg .45359237 Square feet m2 .09290304 Stilbs 1x 3.1415927*1o4 .155 APPENDIX A-Z Table A-2--Physical Constants (Weast, 1976). Constant Value Units AC 0.012011 kg/at As 0.03206 kg/at c 2.9979*107 m/s cpa (300K) 1004.6 J/kg K cpw (273K) 4217.7 J/kg K DH20 (281K) 0.239*10‘4 mz/s DH 5 (289K) 1.77*10'9 mz/s 2 g 9.807 m/s2 h' 6.6256*10-34 J.s K1 (291K) 9.1*10‘5 mole/m3 K2 (291K) 1.1:»10'9 mole/m3 k 1.3805*10-23 J/K Le (273K) 2.50*106 J/kg MW 0.01801534 kg/mole P 1.01325*105 Pa R 8.314 m3.Pa/mole.K 1a (300K) 2.64*lO-2 W/m.K 1w (280K) 0.574 W/m.K 156 157 Table A-2 Continued. Constant Value Units “a (300K) 1.84*10-5 kg/ms uw (293K) 1.002910'3 kg/ms pa (273K) 1.2929 kg/m3 ow (273K) 1000 kg/m3 8 2 4 0' 5.67032*10" W/m K APPENDIX A-3 Table A-3--Waste Characteristics. Parameter Units Reference P 4.1410‘10 m3/s Pratt et al. (1975) 0 3.4910"10 m3/s Pratt et a1. (1975) [TS]u 35 mole/m3 Ngoddy et a1. (1971) [TS]f 33 mole/m3 Ngoddy et a1. (1971) [c1f 4500 at/m3 Humenik (1977) N 125 -- 1) w 60 kg (Personal communication M. G. Hogberg, MSU) 1) This is a principle variable in the process of establishing an optimal loading rate for the anaerobic swine waste lagoon. 158 APPENDIX A-4 Table A-4--Lagoon Characteristics. Parameter Value Units L 10 m o W 5 m o cota 3 ’- H 2.7 m T 15-20 oC [H+] 2.5*10-5 eq/m3 All these parameters are measured. The description of the shape parameters is given in Chapter 7. .159 APPENDIX A-S Table A-5--Microbia1 Characteristics 1) Parameter Value Units Source 0 1 4410"4 l/s Senez (1962)2) maxl ' -5 * “max2 3.6 10 l/s van Gemerden (1974) “max3 ”max2 l/s Truper (1964b) “max4 not known 1/s -5 . * _ “maxS 4.22 10 l/s Figure 6 3 Y1 l.4*10lo cells/mole Senez (l962)3) Y2 ”.25*Yl cells/mole 4) Y3 = 3*Y2 cells/mole Eqns. (3-1), (3-2) and (3'4) I (3'5) Y4 not known cells/mole Kl .15 mole/m3 From computer simula- tion K2 .007 mole/m3 van Gemerden (1974) K3 55 m2/m3 van Gemerden (1971) _ 2 3 K4 —K3 m /m 5) K5 200 1x Figure 6-3 KI 2.5 mole/m3 van Gemerden (1974) I1 10 1x Takahashi et al. (1972) N 2 -— Microscopical obser- ° vation p 2000 kg/m3 van Gemerden (1968a, 160 1971) and Weast(1976) APPENDIX A-6 Comments on Table A-5. 1. Since Thiocapsa roseopersicina and Chromatium vinosum show in many aspects an identical behaviour I have used the reported values for g; vinosum, if none were available to me for T; roseopersicina. Value is given for Desulfovibrio desulfuricans. I deduced this value from the yield coefficient (in grams of cellular material per mole of sub- strate) which is given by Senez (1962) for the reduction of sulfate with pyruvate and with acetate as source of carbon. This is just a first estimate. It is likely, that both sulfur consuming processes are governed by the same saturation constant, since not the metabolic process itself, but the transport from the globules is rate limiting, as far as these two processes can be considered separately. 161 APPENDIX B ANALYTICAL METHODS APPENDIX B ANALYTICAL METHODS 8.1 The sulfide analysis Many procedures have been developed for the analysis of hydrogen sulfide. Benthge (1953) describes three groups of analytical methods which are based on the oxidation of the sulfide. He concludes that the iodine method is the most accurate, but relatively insensitive and that the alkaline hypochloride method is the most sensitive, but relatively inaccurate. For all methods the precision depends on suitable oxidation conditions. In the analysis of sulfide in lagoon water many interferences for these oxidative methods can be expected, since the anaerobic environment contains many compounds in the reduced form. The analytical methods discussed by Bethea (1973) concern the analysis of hydrogen sulfide as an odorous pollutant. All these methods are quite elaborate and are influenced by the many possible interferences. In the following I will describe the use of a specific ion elec- trode which I have used for my measurements. The advantages of the specific ion electrode are the simplicity of the procedure and the low number of interfering compounds. 162 -163 (Mercury is known to interfere with the electrode per- formance. Free mercury is, however, not present in sulfide containing samples.) The sulfide ion electrode used is the model 94-16 solid state electrode of Orion (1974). In addition to the electrode the instrumentation consists of a double junction reference electrode, a digital pH/mV meter, an electrode holder, a magnetic stirrer and stirring bar, a buret and sample beakers. Chemicals which were used are: - sodium hydroxide - sodium salicylate - ascorbic acid 1 2 3 - lead perchlorate (10’ M, 10’ M, 10’ M and 10'4M) - potassium nitrate (1M) - nitroqen For the preservation of samples a sulfide anti-oxidant buffer (SAOB) was prepared by adding 40 g sodium hydroxide, 160 g sodium salicylate and 36 g L-ascorbic acid to 500 ml distilled water (flushed with nitrogen). The final volume was made up to 1 liter with distilled water (flushed with nitrogen). Sample bottles were filled for half of the volume (i.e., 50-60 ml) with the SAOB solution and flushed with nitrogen. At the time of sampling the content of two test tubes with sample of a combined volume of 50 ml was added to the sample bottle with SAOB and the tubes were once rinsed with a small quantity of distilled water. 164 Samples were titrated with a lead perchlorate solu- tion. After each addition of lead perchlorate, a millivolt reading with the Specific ion electrode was made. Small additions near the endpoint resulted in an accurate titra- tion curve from which the endpoint was graphically determined. After each addition of lead perchlorate, it is necessary to reach equilibrium before a millivolt reading is made. The time-required for this process appeared to be a function of the age of the sample. While no significant change in the endpoint was observed, the time required for the titration could be reduced considerably by storing the sample for about 24 hours. A possible explanation for this phenomenon might be the breakdown of organic complexes. The analysis of sulfide with a specific ion elec- trode gave generally values within 1% accurate. A few values will deviate 5-10% as result of a choice for a too concentrated lead perchlorate solution. B.2 The sulfate analysis Sulfate was determined gravimetrically with barium chloride. The content of two test tubes (50 ml) was added to a sample bottle, which contained a small quantity of activated carbon powder. The sample was subsequently acidified with a few drops concentrated hydrochloric acid and stored in a refrigerator. After centrifugation, the sample was filtered through a previously washed millipore 165 filter (.45 um) and the filtercake was washed with distilled water. The filtrate was heated till the boiling point and a filtered warm solution of barium chloride was slowly added till an excess barium chloride was present. After 2 hours, the filtrate was cooled down and filtered through a washed, dried and weighed millipore filter (.45 um). The resulting filter and filtercake were washed with warm distilled water (with a drop of ethylene glycol to reduce the surface tension) till no precipitate was formed (in the filtrate) with a silver nitrate solution. As a final step, the filter was dried till constant weight. The experimental error in the sulfate analysis is estimated to be about 15 to 20%. APPENDIX C ENVIRONMENTAL DATA APPENDIX C-l q.— «.0 n.~ w.HH m.~ 0.0” n.m m.0H 0am 0.0 0.H N.0H 0.H m.HH 0.H N.HH 0.H m.Hn v.~ 0.0” w w ~.H 0.0 0.0 m. 5.0 0.v ~.H v.0 0.H N.0 Hmlcm 0N10H 0.HN H.vm 0.vm n.5m N.QN 0.0m 0.0m 0.0m H.NN v.vm 0.w~ H.5m m.m~ 0.0m N.0N H.0m 0.0m v.00 m.mN w w w 0.NN N.vm 0.m N.0 n.0H 0.0N 0.0m n.0m m.0 0.0M 0HI0H manna 0.mv 0.0V H.0v H.0v 0.Hv 0.0V m.~m swam m.~m w e 0.5V 0.0a w.mv H.0v m.mm BHI0H N.vm v.~0 v.0m 0.0m H.wv 0.N0 m.N0 0.00 0.N0 $0 5. 0.5a H.00 H.0H N.0m NHvHH m.va 0.0m m.0v m.mH 0.5m Haaofi N.Nm 0.0m 0.0m 0.wm w.mH 0.0m N.0m v.00 N.nm (s. 60 v.0H H.n v.0m 0.5 0.mm 0H10 H.0H 0.?N m.0~ N.H~ H.NH 0.0m H.v~ H.0N 0.0m 6- o. 0.H v.0N 0.v N.0N 0.:0 m.n 0.0H 0.0 0.0 m.m 0.0m 0.0 N.0H 5. (x. 0.N 5.0 0.5 v.H n.“ wlu nl0 95H 0.0 0.0 «.0 0.0 H.0 0H\m mH\m 4 10. 40 1. 00 0 . . .(_ c 1 _ 7 ' ' 7 l 40 40 :0 0 "u~ 00 10 . 1 ,. . 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CL IHQVIC CENTER co" 21.0. m FEDERAL BUILDING “SFEVILLE- N-C- 20801 I FIRST CLRSS BIBLIOGRAPHY BIBLIOGRAPHY Abeliovich, A. and Azov, Y. 1976. Toxicity of Ammonia to Algae in Sewage Oxidation Ponds. Appl. Environ. Microbiol. 31(6):801-806. Albertson, O. E. 1961. Ammonia Nitrogen and the Anaerobic Environment. J. Water Pollut. Control Fed. 31(9): 978-995. Albone, E. S. 1972. Formation of Bis-(p-chlorophenyl)- acetonitrile (p.p' - DDCN) from p.p'-DDT in Anaero- bic Sewage Sludge. Nature (Lond) 240:420- Amir, I., and Ogilvie, J. R. 1977. A mixed Integer Pro- gramming Model for Choosing an Optimal Swine Manure Handling System. Paper No. 77-4029 American Society of Agricultural Engineers, St. Joseph, MI, 20 pp. Anderson, E. R. 1952. Energy Budget Studies Water-Loss Investigations: I, Lake Hefner Studies. U. S. Geol. Surv. Circ., 229, 71-119. Andrews, J. F. 1965. Kinetics and Characteristics of Multi-stage Methane Fermentations. Ph.D. Thesis, Univ. of CalifOrnia, Berkeley. Diss. 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