This is to certify that the ' thesis entitled THE REMOVAL OF NITROGEN FROM NASTEWATER BY CORN AND RYE: A GREENHOUSE STUDY presented by Michael Eric Sevey has been accepted towards fulfillment of the requirements for M. S. degreeinCrop and Soil Science {221%} Major professor £//7’,/7,7 0-7639 THE REMOVAL OF NITROGEN FROM NASTEWATER BY CORN AND RYE: A GREENHOUSE STUDY By Michael Eric Sevey A THESIS Submitted to Michigan State University in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Department of Crop and Soil Sciences 1979 ABSTRACT THE REMOVAL OF NITROGEN FROM NASTEWATER BY CORN AND RYE: A GREENHOUSE STUDY By Michael Eric Sevey Nitrate losses to ground water must be minimized in a land application site for sewage effluent. Crop sel- ection is an important consideration in the management of nitrate losses from the application of sewage effluents to the land. The purpose of this experiment was to study the effect of crop selection on the loss of nitrate in drainage water from the soil profile of a simulated land application system under greenhouse conditions. There were three crop systems: corn, rye, and corn intercropped with rye. Three rates of simulated ef- fluent were applied: 5 cm/wk, l0 cm/wk, and 15 cm/wk. The simulated effluent contained an average of 6.6 ppm nitrogen, which is the average for the Muskegon County Wastewater Project. Corn intercropped with rye minimized nitrate loss- es in drainage water. Rye did nearly as well by itself. Corn was not very effective in controlling nitrate losses. Generally less total nitrate was lost in all cropping systems when treated with the lowest simulated effluent rate (5 cm/wk), but moisture and nitrogen were probably limiting crop growth. The l5 cm/wk simulated effluent treatment provided the best growth and minimized nitrate loss for rye and corn intercropped with rye. Both of these crops reduced the nitrate nitrogen concentration of the drainage water to less than 0.05 ppm by the end of the experiment. TO MY WIFE AND CHILDREN ACKNOWLEDGMENTS I AM THANKFUL FOR THOSE MEN AT MICHIGAN STATE NHO INTRODUCED ME TO AGRICULTURE. ii TABLE OF CONTENTS LIST OF TABLES LIST OF FIGURES INTRODUCTION LITERATURE REVIEW History of Land Application of Waste . The Importance of Land Application of Waste Nitrogen in Wastewater . . Nitrogen Cycle . Crop Removal of Nitrogen MATERIALS AND METHODS Greenhouse Apparatus Treatments . Experiment Initiation Sampling and Analytical Methods RESULTS AND DISCUSSION The Greenhouse Environment Water Balance Plant Growth Nutrient Balance CONCLUSIONS APPENDIX Table l - Analysis of variance for volume of drainage, nitrate nitrogen concen- tration of drainage, and total amount of nitrate nitrogen lost in drainage. Table 2 - Analysis of variance for dry weight of plants. . . . BIBLIOGRAPHY iii —l _.a ._a -‘OKO\I \1 000305014) 45 NN—l—J 60 6l 62 TABLE I TABLE 2 TABLE 3 TABLE 4 TABLE 5 TABLE 6 TABLE 7 TABLE 8 TABLE 9 TABLElO TABLEll TABLETZ LIST OF TABLES SCHEDULE OF EFFLUENT APPLICATIONS. VOLUME 0F SIMULATED EFFLUENT AND DISTILLED WATER ADDED TO SOIL COLUMNS. AVERAGE VOLUME OF LEACHATE COLLECTED AS DRAINAGE FOR EACH TREATMENT COMBINATION. APPROXIMATE VOLUME OF WATER EVAPOTRANSPIRED. HEIGHT OF CORN PLANTS. AVERAGE DRY WEIGHT OF PLANT MATERIALS. NITROGEN APPLIED IN SIMULATED EFFLUENT. AVERAGE CONCENTRATION OF NITRATE NITROGEN IN DRAINAGE WATER. AVERAGE AMOUNT OF NITRATE NITROGEN LOST IN DRAINAGE WATER. AVERAGE NITROGEN CONTENT OF PLANTS. AVERAGE PERCENT NITROGEN OF SOIL SAMPLES FROM THREE DEPTHS. APPROXIMATE NITROGEN BALANCE. iv 20 27 29 35 38 39 4T 42 46 49 52 54 FIGURE FIGURE FIGURE FIGURE FIGURE LIST OF FIGURES AVERAGE DAYLIGHT PERIOD TEMPERATURE IN THE GREENHOUSE.' AVERAGE PERCENT OF POSSIBLE SUNSHINE FOR LANSING, MICHIGAN FROM FEBRUARY 8, 1976 TO APRIL 24, l976. PERCENT OF APPLIED WATER THAT DRAINED FOR EACH CROP (AVERAGED ACROSS IRRIGATION TREATMENTS). WATER DRAINED BY EACH IRRIGATION TREATMENT (AVERAGED ACROSS CROPS). CALCULATED VOLUME OF WATER EVAPOTRANSPIRED BY EACH CROP (AVERAGED ACROSS IRRIGATION TREATMENTS). 24 26 32 34 37 INTRODUCTION Land treatment and disposal of wastewater in the past decade has risen from the status of "Ancient Practice" to that of a "Scientific Technology". Waste treatment and disposal facilities, both municipal and industrial, must be designed for maximum water renovation with minimum cost. Strict water quality criteria and rising cost of tertiary treatment require the development of cheaper and more efficient methods of waste treatment. Municipal and indus- trial wastewaters have been successfully renovated by land application. Many land treatment and disposal facilities use a vegetative cover to remove the nutrients from the waste- water. Forest and grass cover crops have been popular be- cause they require very little maintenance. Federal fund- ing agencies are encouraging land treatment facilities that will become self supporting. One method of reducing costs at a land treatment facility is to harvest the nutrients from the wastewater by growing a crop which can be sold. The Muskegon County Wastewater Management system irrigates about 2,400 hectares of corn with its sewage efflu- ent. Corn is an attractive crop because of its cash value and ready market. But it is limited by a growing seaSon of about l00 days. Corn actively removes nitrogen from the soil water system for about half this time. It seems un- likely that corn can be effectively used to renovate wastewater at Muskegon because it uses nitrogen at an appreciable rate for only about one third or less of the l45 day wastewater irrigation season. The soils at the Muskegon project are infertile sands. Water percolates through the soil quickly. The wastewater applied at Muskegon has a nitrogen concentration of about 6.6 ppm nitrogen which is low in comparison to the nitrogen needs of the corn plant. For maximum nitrogen removal the crop should have a dense root growth reaching into a large volume of the soil profile, a high nitrogen requirement and a long growth period. With these conditions the crop could quickly use the nitrogen before it is lost to the ground water. Many questions must be answered before we can make good decisions concerning each specific use of the land application concept. We need to know how well specific crops can use the nitrogen in wastewater. The diverse con- tents of wastewaters make experience at one site difficult to use at other sites. The impact of toxic compounds and heavy' metals in wastewater spread on the land is far from defined. Each site presents a new set of physical, chemi- cal and biological parameters. This study seeks to point out some of the limita- tions of corn as a crop for land treatment and disposal of wastewater. At Muskegon a cover crop such as rye could be used to compliment corn. If corn were no-till planted into es- tablished rye it would extend the nitrogen stripping period because of the different growing seasons of the two crops. This could provide better water renovation for a longer period of time. If the intercropping were managed properly it might be possible to maintain corn yields at a high enough level to reduce land treatment costs. This study seeks to determine the limitations of corn as a crop for land treatment and disposal of waste- water. It compares the nitrogen stripping capabilities of corn, rye and corn intercropped with rye, using three sim- ulated effluent application rates. LITERATURE REVIEW HISTORY OF LAND APPLICATION OF WASTE Treatment and disposal of wastewater by land application is not a new concept in waste handling. This simple technology has been employed for at least four hun- dred years (Thomas, 1973). The applications of waste to the land was the simplest and most logical method of treat- ment and disposal. As early as l875 methods such as sedi- mentation and chemical precipitation had been successfully used to purify wastewater (Egeland, l973). But land appli- cation remained the only widely accepted method of waste treatment until after l900. In Germany during the 16th century, wastewater was applied to cropland because it seemed to have some value as a fertilizer or soil amendment (Deturk, 1935). The yield from an early European sewage farm would often be double that from a conventional farm in the same area (Pound and Crites, l973b). Historically man has had a very narrow concept of land application as simply a method of waste disposal. Recently this concept has enlarged to view land application as a method of water treatment and water recycling (Thomas, l973). There is also renewed interest in the valuable nutrients contained in these waste materials. The first "land disposal" operations in the United States began late in the 19th century. Historical data on these facilities can be found in Pound and Crites (l973b). Land application of wastewater has been practiced for a variety of reasons including economical alternatives to waste treatment, to provide supplemental irrigation water, as a method of disposal in situations lacking suit— able receiving waters, and ground water augmentation (Sullivan et al., l973). THE IMPORTANCE OF LAND APPLICATION OF WASTE The Federal Water Pollution Control Act Amendments of l972 recognized the fact that our nations waters are in a degraded state. These amendments state that as a nation we must "restore and maintain the chemical, physical, and biological integrity of the nations waters" and that "it is the national goal that the discharge of pollutants into navigable waters be eliminated by l985" (Section 101, Public Law 92-500). In addition to the ”zero discharge" goal, several other portions of the amendments attempt to create a re- newed interest in land application of wastewaters. Section 20l of Public Law 92-500 requires that in order to receive federal assistance, a project must study "alternative waste management techniques" and the technique chosen must be the most effective on a cost basis. This is significant in that land application is very economical. Section 212 of Public Law 92-500 also allows that land used in wastewater treatment (such as land application, ponds, etc.) may be purchased with federal money. Section 201 also encourages revenue producing facilities which recycle waste materials into salable agricultural, silvicultural and aquacultural products, and facilities which provide recreational poten- tial. NITROGEN IN WASTEWATER In 1972 our nation treated and disposed of 7.5 billion gallons of wastewater per day (National Association of State Universities and Land Grant Colleges, 1973). The approximate concentration of total nitrogen in an average municipal secondary effluent may be 20 - 25 mg/liter (Etzel and Steffan, 1974; Lance, l972; Pound and Crites, 1973b). Most municipal treatment plants have been discharging their wastewater after secondary treatment. Combining these figures for 1972 we could estimate that 207,000 to 259,000 metric tons of nitrogen were discharged into our water re- sources by municipal treatment plants. The Hazardous Materials Advisory Committee (1973) has estimated that the wastewater discharged by our nations domestic wastewater treatment plants could contain as much as 840,000 metric tons of nitrogen. Human waste contains about 5.4 kg of nitrogen per person per year. Thus in 1972 about 1,100,000 metric tons of nitrogen entered our sewage and septic sys- tems (Committee on Nitrate Accumulation, 1972). The quality and content of municipal and industrial wastewaters at various stages of treatment are well documen- ted (Hazardous Materials Advisory Committee, 1973; Pound and Crites, 1973a, 1973c; Reeves, 1972). The composition of municipal wastewater varies greatly and industrial efflu- ents are even more diverse. The characteristics of waste- waters are so different depending on the source that each must be evaluated before good decisions concerning any treatment method can be made. Organic and ammonium nitrogen are the main forms of nitrogen in municipal effluents applied to land (Lance et al., 1976; Adams, 1973; Pound and Crites, 1973b). The ammonium ion may account for 90% of the total nitrogen in secondary effluents. Nitrate and nitrite may also be present. Nitrate concentrations are usually less than 10 mg nitrogen per liter. Nitrite is rapidly oxidized to nitrate under areobic conditions so concentrations above 1 mg/liter are unusual (Pound and Crites, l973b). Nitrite is generally not a problem in secondary municipal effluents. The control and removal of nitrogen from waste- water has been extensively studied. Several reviews give good descriptions of methods such as air stripping, ion exchange, biological nitrification-denitrification, and breakpoing chlorination (Atkins and Scherger, 1977; DeRenzo, 1978; Reeves, 1972 and Ripley et al., 1974). NITROGEN CYCLE There exists within the soil a very complex net- work of biochemical and chemical reactions which use and recycle nitrogen. A complete understanding of these mech- anisms is essential in attempting to develop a land appli- cation system which will maximize nitrogen removal from wastewater. There is an abundance of information available on soil nitrogen (Buckman and Brady, 1969; Bartholomew and Clark, 1965; Lance, 1972; Pound and Crites, 1973b). Sev- eral of these references include a detailed diagram illus- trating the nitrogen transformations and pathways in the soil. A mineral soil will normally contain 0.02 - 0.5 percent nitrogen in the "plow layer" (Buckman and Brady, 1969). This is equivalent to about 0.18 to 4.5 metric tons of nitrogen per acre furrow slice. Nitrogen occurs in the soil mainly in the organic form. Nitrogen also occurs as ammonium fixed by the soils exchange complex and as soluble ammonium and nitrate. Organic nitrogen suspended in wastewater is physi- cally filtered out as it is applied to the soil. This organic nitrogen is subject to mineralization (microbial decomposition) to ammonium. Many microorganisums are capa- ble of mineralizing organic nitrogen. Mineralization is favored by well drained and aerated soil conditions. Soil temperature, pH, and concentration of soluble mineral nit- rogen also affect the rate and occurrence of nitrogen mineralization (Harmsen and Kolenbrander, 1965). It may be possible to predict nitrogen mineralization rate as the soil moisture conditions change (Stanford and Epstein, 1974). Ammonium nitrogen, the major inorganic form of nitrogen in wastewater, may be volitalized, immobilized, adsorbed or absorbed by soil, or nitrified as wastewater is applied to land. Harmsen and Kolenbrander (1965) have reviewed the study of ammonia losses from soil. Several factors which can increase ammonia volitalization from soil are pH above 7.0, increased aeration, and ammonia concentrations which exceed the capacity of the soil to "sorb" ammonia. Ammonia losses from the wastewater before it comes into contact with the soil can be significant if the wastewater has a pH of 8.0 or greater and if adequate air-water contact is maintained such as in a spray irrigation system (Lance, 1972; Lance et al., 1977). Nitrogen immobilization is best described as the "tying-up" of inorganic forms of nitrogen in microbial cell tissue during the decomposition of organic residues that contain nitrogen. Ammonium nitrogen is more readily used by most soil microbes. In a land application system the amount of wastewater nitrogen immobilized is dependent on the carbon to nitrogen ratio of the wastewater (Lance, l972; Lance et al., 1977). Low temperatures slow down the immobi- lization process. Aerobic conditions promote greater lO immobilization than anerobic conditions. Immobilization rate varies with soil depth due to differences in moisture, temperature, pH, and oxygen availability (Bartholomew, 1965). Nitrogen is fairly stable after it is immobilized in the soil. Negatively charged mineral and organic fractions or colloids of the soil can adsorb ammonium nitrogen (Lance 1972; Pound and Crites, 1973b; Scarsbrook, 1965; Stevenson, 1965; Tilstra et al., 1972). The amount of nitrogen ad- sorbed depends on the cation exchange capacity of the soil and the concentration of other cations in the soil. It is possible to estimate the amount of nitrogen adsorbed in this way (Lance, 1972). Ammonium nitrogen which is adsorb- ed on the exchange complex is generally unstable. It is subject to microbial oxidation (nitrification), leaching, immobilization, and plant uptake if the environmental conditions are favorable. Ammonium adsorbed in an anaer- obic zone is relatively stable except when conditions favoring displacement from the exchange complex and leach- ing are in effect (Avnimelech and Raveh, 1976; Lance, 1972; Pound and Crites, 1973b). Ammonium nitrogen can be fixed by clay and organic fractions in the soil so that it is very stable compared to the ammonium nitrogen adsorbed on the exchange complex. The forces which hold this nitrogen are much stronger than those which hold ammonium in its readily available state on the soils exchange complex. The fixation of ammonium 11 by certain clays involves the incorporation of the ammonium ion into their crystal lattice (Lance, 1972; Buckman and Brady, 1969). There are several detailed discussions of this complex subject and those factors which affect it (Mortland and Wolcott, 1965; Nommik, 1965). The organic fixation of ammonium is not well understood. Compounds resulting from these soil reactions are fairly stable. The proposed nature and significance of this form of ammon- ium fixation have been reviewed by Lance (1972), Mortland and Wolcott (1965), and Nommik (1965). The soluble and readily available fractions of ammonium nitrogen in the soil may be biologically oxidized to nitrate nitrogen under aerobic conditions. This micro- bial conversion of ammonium nitrogen occurs rapidly in soils when aeration, acidity and temperature are favorable to plant growth (Committee on Nitrate Accumulation, 1972). Nitrification has been thoroughly studied and is very well understood (Bear, 1953; Buckman and Brady, 1969; Alexander, 1965). Nitrate nitrogen accumulation in soils is a major ecological concern because nitrate is so easily leached to the ground water. Nitrate nitrogen is a substantial con- tributor to the eutrophication of our surface waters and the contamination of our ground waters. The United States Department of Health, Education and Welfare has set 10 mg N/l as the maximum concentration of nitrate allowed in water for human consumption. Nitrate 12 in excess of this level may cause methemoglobinemia, a blood disorder which affects infants less than three months old. Nitrate consumed in the water is reduced to nitrite in the infants stomach. Nitrites then enter the bloodstream and oxidize hemoglobin to methemoglobin, which does not function properly as an oxygen carrier (Bailey and Thomas, 1975; DeRenzo, 1978; Lehninger, 1970; Steel, 1960). In the soil nitrites are very unstable. They are denitrified under anerobic conditions or oxidized to nitrate in the presence of oxygen and the appropriate microbes. Many studies have been conducted on denitrification (Ardakani et al., 1975; Lance et al., 1976; Volz et al., 1975). Denitrification or the microbial reduction of nitrate to nitrogen gas and its oxides requires an environ- ment which provides reducing substrates (Lance and Gerba, 1977) and anerobic conditions. Many kinds of microorgan- isms can serve as denitrifiers so the presence of the proper microflora is seldom limiting to denitrification (Buckman and Brady, 1969). Environmental factors affect- ing denitrification have been reviewed by Bear (1953) and Harmsen and Kolenbrander (1965). In well aerated soils, such as those used for crop irrigation with sewage effluents, denitrification may occur in anerobic microenvironments (Nommik, 1965; Volz, 1975). Denitrification and nitrifi- cation can occur in the same syStem by alternate flooding and drying cycles (Bouwer et al., 1974; Lance and Whisler, 1972). Nitrogen gas or nitrogen oxides evolved from the 13 denitrification process are completely removed from the soil system. This makes denitrification an important fac- tor in the removal of nitrogen from wastes with land appli- cation. Denitrification may also occur as the result of a chemical reaction between soil organic matter and nitrate. This reaction generally does not remove a significant amount of nitrogen from the soil system (Pound and Crites, 1973b). CROP REMOVAL OF NITROGEN A crops need for nitrogen is tremendous. More atoms of nitrogen are needed than any other nutrient sup- plied by soil or fertilizers (Viets, 1965). Knowledge of this fact makes cropping a prime candidate for nitrogen re- moval in a wastewater treated soil. A crop performs a very important form of "immobilization" in the soil nitro- gen system. There are several good discussions on the ability of various crops to remove nitrogen and the manage- ment practices and decisions which must be considered be- fore implementing crop removal of nitrogen by land applica- tion (Day, 1973; Erickson, 1974; Pound and Crites, l973b; Sopper, 1973; Sullivan et al., 1973). Plants can remove nitrogen from the soil in either the nitrate or ammonium form. There have been many studies to determine which ion perfbrms best in supplying nitrogen to plants. Viets (1965) and Dibb and Welch (1976) have summarized some of these investigations. These studies have pointed out that either ion may be a better nitrogen 14 source depending on specific soil environmental conditions. Alessi and Power (1973) approached this question by study- ing nitrogen recovery with various fertilizer nitrogen materials. They found the percent recovery of ammonium and nitrate to be very similar. It is difficult to study the uptake of ammonium by plants growing in soil because of nitrification. Many investigators have used solution eul- tures because there nitrification is inhibited. Chemical nitrification inhibitors have been used in the soil to maintain ammonium concentrations during uptake studies (Warncke and Barber, 1973). Uptake of other nutrients may be affected by the nitrogen source coupled with soil and plant response (Viets, 1965). The nitrogen uptake efficiency of crops vary. Also the fertility levels required to maintain maximum nitrogen uptake are different for each crop and its soil environment- al conditions. Warncke and Barber (1974a) have described the nitrate uptake effectiveness of corn, soybeans, sorghum and bromegrass. Corn was found to absorb nitrate more efficien- tly than the other crops. It stripped nitrate from a 1000‘”,M solution, reducing the nitrate concentration to as low as 2 ,LM. At this low level plants yellowed severely and nitrate uptake ceased. A solution of 1000 p.M was ad— equate for optimum growth of corn, sorghum and bromegrass. Bole and Bell (1978) studied the effect of waste- water on yield and composition of several forages grown in 15 the field. All forages except tall wheatgrass took up more nitrogen than was applied. Fertilizer nitrogen was added to some treatments. Effect of wastewater nitrogen on yield was equal that of fertilizer nitrogen. Day and Kirkpatrick (1973) found that treated wastewater can be used to produce oats with grain yield and protein content approximately equal to that of oats grown with fertilizer supplemented well water. Hanway (1963) has sought to describe the identi- fying characteristics of each stage of physiblbgical and morphological development of the corn plant. According to Hanway, nutrient uptake in corn begins to occur at a rapid rate during the second stage of growth. The second stage is characterized by the appearance of the collar of the eighth leaf, about 28 days after emergence. Maximum nutri- ent uptake generally occurs during growth stage 5 or about 66 days after emergence when 75% of plants have visible silks. Edwards and Barber (1976) studied the influenCe of age on nitrogen uptake of corn roots at low nitrogen concentrations between 01p.M and 150 fLM. In this solution culture study maximum influx of nitrogen occurred above 21 #.M. Greatest influx occurred with plants 18—24 days old. Under field conditions Mengel and Barber (1974b) found nutrient uptake of nitrogen in corn to reach a max- imum of 1200 ,aM/day at a plant age of about 50 days. This 16 nitrogen uptake decreased to a minimum of 113,“.M/day per plant at silking (about 70 days). Warncke and Barber (1974b) measured a maximum nitrogen uptake per corn plant of 8.1 mg atoms/plant/day, for plants in the 60 - 67 day age range. This rate dropped dramatically during the transition from the vegetative to the reproductive stage (74 - 81 days). Corn root development under field conditions has been studied by Mengel and Barber (1974a). They found that for the first 75 days root length increased rapidly. Root length then leveled off and began to decrease rapidly at about 90 days. Nutrient uptake increased in a similar manner but reached a maximum at 50 days and then began to decrease. A maximum nitrogen uptake of 1200’u.M/day was reported for 50 day old plants. Nitrogen uptake dropped to a minimum of 113’u.M day at 80 days. MATERIALS AND METHODS This study was conducted in the greenhouse from February 8, 1976 to April 24, 1976. The effects of various crop and irrigation treatments on nitrogen removal from an applied simulated effluent solution were studied using the following techniques. GREENHOUSE APPARATUS Columns were constructed from 1.5 m lengths of 10 cm diameter P.V.C. pipe. A plexiglass base was cemented to each column. A .95 cm drainage hole was bored in the wall of each column at the base and fitted with a drainage tube. Drainage samples were collected from the tube in 1 liter plastic bottles wrapped with aluminum foil. The 36 columns were supported in wooden racks, each rack holding 18 columns. The racks were oriented north to south on the greenhouse bench and were 75 cm apart. Columns were spaced 14.6 cm from center to center in the racks. The "C" horizon of a Rubicon sand was used as a growth medium. The soil was obtained from an uncultivated area near circle #3 of the Muskegon County Wastewater Treatment Facility. The parent material of the soil profile was used because the surface horizons of these soils are 17 18 often absent and the "C" horizon has the least organic matter and nitrogen to confound the experiment. The soil was air dried, passed through a 5 mesh sieve, and then mixed. A plug of glass wool was placed over the drainage hole on the inside of the column to prevent the soil from washing out. A large plastic funnel attached to a 1.5 m length of plastic pipe with a diameter of 2.5 cm was used to distribute the soil evenly in the columns. The soil was poured into the funnel and gently deposited with a slow circular motion. The columns were gently tapped during the filling process to eliminate air pockets and to distribute the soil as uniformly as possible. The length of the natural daylight period during the experiment ranged from 10 hours and 15 minutes on February 8, 1976 to 13 hours and 46 minutes on April 24, 1976. Supplemental lighting was used to extend the day- light period to 14 hours per day for the duration of the experiment. Two light fixtures were used, each with 4, eight foot long cool white florescent bulbs. They were made from 1.59 cm diameter pipe frames with no reflectors, so that plants would not be shaded from the sun. No incandescent bulbs were used. Temperature measurements in the greenhouse were recorded with a seven day recording thermometer. The temperature at night was kept at 13°C with steam heat. Windows were opened daily if temperatures reached 24°C or 19 more. TREATMENTS Nine treatment combinations were used, a 3 X 3 factorial with 3 cropping systems and 3 irrigation treat- ments. Each treatment combination was replicated 4 times. The cropping systems used were corn (Pioneer #8780), rye (Balbo rye), and corn intercropped with rye referred to as corn-rye in the remainder of this thesis. At the Muskegon Wastewater Project, corn is the main crop- ping system. Rye was included in this experiment because it actively removes nitrogen for a longer period of time. A simulated effluent was made up daily to match the nitrogen and phosphorus content of the effluent applied to the soil at the Muskegon County Wastewater Project. The solution contained an average of 6.6 ppm nitrogen from NH4N03 and 7 ppm phosphorus from KH2P04. The irrigation treatments were 5 cm/wk, 10 cm/wk and 15 cm/wk of simulated effluent. Starting with the fourth week supplemental applications of distilled water were given to each column to approximate what was evaporated from the surface of a pan evaporimeter with the same diameter as the columns. The distilled water was applied between irrigation treatments. During the eighth week the 5 cm/wk treatment was changed to 5 cm/wk of simulated effluent plus 5 cm/wk of distilled water. This altera- tion was necessary to maintain a measurable drainage rate. The total amount of nitrogen applied did not change. 20 Simulated effluent was applied in units of 2.5 cm per application. No more than one application of simulated effluent was made in any 24 hour period. Table 1 shows a schedule of weekly effluent applications. TABLE I - SCHEDULE OF EFFLUENT APPLICATIONS RATE OF APPLICATION SUN. MON. TUE. WED. THUR. FRI. SAT. cm/wk 5 - X - - - X - 10 - X - X - X X 15 - X X X X X X X - denotes application of 2.5 cm of simulated effluent. EXPERIMENT INITIATION The packed soil columns were brought to near saturation by flooding with distilled water. They were allowed to drain for 48 hours to bring them to approximate field capacity. Rye was planted on February 2, 1976. The seeds were soaked in aerated water for 24 hours prior to planting to hasten emergence. Twelve rye seeds were planted in each column at a depth of about 1.3 cm. The rye popula- tion was thinned to 8 plants per column after one week. Corn seed was soaked 24 hours and planted about 2.5 cm deep. Two corn seeds were planted in each column and later thinned to one plant. All columns were watered with 100 mls of distilled water per day until all seedlings had emerged. Irrigation treatments were started on February 9, 1976. 21 SAMPLING AND ANALYTICAL METHODS Drainage samples were usually collected every other day. They were stored in a cooler at 4°C. Drainage sample volumes were determined by weight. Fresh drainage samples, which had not been in the cooler more than 48 hours, were analyzed for nitrate using the colorimetric cadmium reduction method (HenrikSen and Selmer-Olsen, 1970) on the Technicon Auto-Analyzer. Drainage samples were occasionally analyzed for phosphorus, ammonium and nitrite to determine if these nutrients were being lost in the leachate. Phosphorus was measured using the ammonium molybdate-ascorbic acid method (Watanabe and Olsen, 1965). Ammonium was measured with the colormetric phenate method (U. S. Environmental Protection Agency, 1974). Nitrite concentrations were determined with the method of Henriksen and Selmer-Olsen, (1970). All of these analYSes were done with the Technicon Auto-Analyzer. The rye plants in the corn-rye treatments were cut back to about 5 cm during the eighth week to reduce the stress on the corn plants in this treatment. The harvested rye was saved and added to final yields. The experiment was terminated after 11 weeks. At this time all plant materials were harvested and oven dried at 60°C. Dry weights were determined. Plants were then ground to pass a 40 mesh screen and analyzed for total nit- rogen using the Micro—Kjeldhal procedure. After all vegetation was removed, the columns were 22 flushed with 3 liters (about 25% of the volume of the column) of distilled water. The first 3 liters of drainage water were collected and analyzed for nitrate to determine how much nitrate accumulated in the column. After the soil columns were flushed they were allowed to drain for two days. The soil—root plugs were then removed from the P.V.C. pipe by inverting the column and sliding its contents out. A 2 cm slice of soil was taken from a depth of 15 cm, 76 cm and 140 cm of each soil column. The soil samples were air dried for 5 days on the greenhouse bench, put into plastic bags, and later analyzed for total nitrogen using the Micro-Kjeldahl method. Soil samples were very sandy so they were not ground before nitrogen analysis. It was not possible to include a repre- sentative sample of roots in each 1 g sample digested for nitrogen analysis, so roots were screened out. Analysis of variance was conducted on all experi— mental data using an ADV computer program. AOV tables are in the appendix. RESULTS AND DISCUSSION THE GREENHOUSE ENVIRONMENT The average daylight period temperature in the greenhouse was calculated for each week by averaging the recorded temperatures every third hour from 6:00 am to 6:00 pm each day of the week. The average daylight period temperatures are represented in Figure 1. As expected, there was a trend toward increased average temperature. The daily average greenhouse temperature ranged from 15°C to 36°C. This would be a normal temperature range that might be encountered in the field during the corn growing season. Plants in this experiment were exposed to greater stress than might be encountered in the field even though temperatures were not abnormal. Low humidity of the green- house air, rapid temperature fluctuations of air and soil, and small pot size contributed to greater stress. Because pots were small, there was very little soil mass so that soil temperature would quickly equilibrate with ambient air temperature. Pot size also contributed to increased stress because each pot allowed only 78 square cm of soil area. If corn in the field were planted at a population of 28,000 plants per acre each corn plant would have 1445 square cm or over eighteen times more soil area. Corn in the corn-rye 23 24 .mmnozzmmmw mIF zF mash< FF b oF P gum: om F mmzoFu u mF I I. cm m d 3 “O V I. n no 3 0 .. 3 mm 25 pots shared 78 square cm with eight rye plants. Percent of possible sunshine data for the Lansing, Michigan area was obtained from the National Weather Service office at Capitol City Airport in Lansing. This data was used instead of actual solar energy measurements. Weekly averages of percent of possible sunshine were cal- culated and are shown in Figure 2. The daytime temperature in the greenhouse was very dependent on the amount of sun- shine. Cloudy weather caused cooler temperatures (Figures 1 and 2, weeks 2 and 4). Sunny days resulted in higher greenhouse temperatures. 0n cloudy days the cold outside air kept the greenhouse temperatures close to the thermo- stat setting of 13°C. WATER BALANCE Water inputs to the soil columns were in the form of irrigation with simulated effluent or distilled water applications. The volumes of all irrigations and distilled water applications for each treatment combination are com- piled in Table 2. The simulated effluent treatments were 5, 10, and 15 cm/wk as described in "Materials and Methods". The maximum irrigation rate at the Muskegon County Wastewater Facility is about 10 cm/wk. The 10 cm/wk irrigation rate was used in this experiment to imitate the maximum rate at Muskegon. Irrigation rates of 5 and 15 cm/wk were included to see how well the crops could strip nitrogen at a rate lower or higher than the rate of water and nutrients applied 26 .ommF .ew Ame< 0F mFmF .w >m<=mmum 20mm z< u N mmeFm xumz P b om om ow om tom 12 new row 3NIHSNDS ETBISSOd dO lNHOHBd 27 mewF ommF mmeF mmeF meF mmNF mme mmmF mmmF mmNF cmNF mMNF mE=Fo> FmFOF mmoF ooF CON CON mNF om omF ooF omF o o 0 mem3 vaFFmeo ommmF QMNF oMNF mmNF omNF mMNF mmNF mmNF mmNF mmNF omNF omNF pcszFFm uwmezeFm "a mFFoF qu «NoF eNoF mqm «Fm vum wNm «mm qu er va mE:Fo> FmFOF mmoF ooF CON CON mNF om omF ooF omF o o o Lopez umFFFmeo doom va «Nw er wNw va va ¢Nw ¢Nw «Nw «Nm va pcszme UmuwFseFm "adj mmNF «Na «NoF eNoF mam Noe Nmm NFm Nam NFv NFc NFv wE:Fo> FmFOF moFN NFm NFm NFo .me om omF ooF omF o o o Lmumz vwFFFmeo Nmmq NF¢ NFq NF¢ NF¢ NFe NFv NFc NFe NF¢ NFq NF¢ ucmsFmFm umeerFm z. 9.33 quoF FF OF 0 m N. o m w m N F mF u N m¢mm wNNm mmm Foe mFm aoF FFN qmF Fm FFN mom mmN mom m vam me «om NmoF mFF «mm va qmo mFFF FmF FoFF mFoF mF mwom va omm mmm mmm FNw mmq 0mm «FF vom FNF Foo oF m>muzmoo mFFm mFm omm mam NmF ONN NmF mm mFN FNm FmN mow m vaFF mom mmm mwm mmm FFoF mFFF omm cmmF me omFF omoF mF mmmo Fov ¢mm mFm wmo moo FwF wmm «Fm woo NNF MFo oF zmou ONwe mFm mNF va ovv mmm me NFN mmm me vom wow m x3FEo Fancy FF oF m N F o m a m N F zoFF mw< u m m4mm-zmoo tom c. . 415% F8 5;. M m I r I I too 4. A. . nAu . I 4 , /_ l mm zmou / OF M I I « I V z... lOw w n u m. . . loo rooF 33 for the corn to recover. The drainage of these columns re— mained higher than rye for the rest of the experiment. As expected, increasing the water input increased the drainage of a given treatment combination. Figure 4 shows the average volume of water drained for each irriga- tion treatment. Each irrigation treatment has been averaged across the three crop systems. Those weeks which had rel- atively high temperature and more minutes of sunshine exhib- it the lowest drainage rates and weeks with low temperatures and less sunshine have higher drainage rates. The 5 cm/wk simulated effluent treatment shows a sudden increase in drainage from week 8 to 9. This is due to an increase in the distilled water applications for this simulated effluent treatment. Beginning week 8, an extra 5 cm/wk of distilled water was applied to all columns which had been receiving 5 cm/wk of the simulated effluent. This was in addition to the distilled water applied as an estimate of evaporation from the soil surface (see Table 3). This extra 5 cm/wk of distilled water was added be- cause no measurable amount of drainage was occurring. Water was a limiting factor in the growth of plants in these soil columns. The approximate volume of water evapotranspired was calculated by subtracting average drainage volumes for each treatment combination (Table 3) from the total volume of liquid applied (Table 2). This data is presented in Table 4. 34 .meomu mmomo< nmwm oszmuzmou mow wow me FmF NqN oFm mme mON Fm va o m Fmv Foo mmv mow mFN woN mFm NF mam ow oFF mF mNm OFv mom FFm moN mmN OFN om oNN NoF FmF oF zmou mam mmN FFm mom FoF FNN 00m MFF Fm woF o m FE x3FEo FF oF m N F m m a m N F zoFFm mNF<3 do mzaho> NFm ommmezm mmF<3 no m2340> omh<42u4m m>m 4 ooF CON com oo¢ com com OOF com com (031v1n31v3) 1w NI GBUIdSNVHIOdVAE aaivm 38 PLANT GROWTH Corn height was measured during the fifth and ninth weeks and at the termination of the experiment. The data is presented in Table 5. Within corn treatments corn height increased with time and simulated effluent applica- tion rate. This growth response to increasing simulated effluent rate was due to greater amounts of water, nitrogen, and phosphorus supplied to the plant. Corn plants in the corn-rye treatments showed no significant response to in- creasing simulated effluent rate. At the time of harvest, corn plants in corn treatments were about twice as tall as the stunted corn plants in corn-rye treatments. Rye plants were not measured TABLE 5 - HEIGHT OF CORN PLANTS. WEEK CROP IRRIGATION 5 9 TERMINATION cm/wk cm 5 26 51 57 CORN 10 28 59 68 15 28 62 74 5 21 27 31 CORN-RYE 10 20 27 30 15 23 29 31 At the termination of the experiment all plant mat- elr‘l'als above the soil were harvested. Plant materials re- moved when the rye was cut back in the corn-rye treatments 39 were included. Table 6 shows the average dry weight of plant materials harvested from each treatment combination. Individual weights for corn and rye from the corn-rye treatments are shown. Corn treatments produced the most dry plant materials at all levels of irrigation. This would be expected under field conditions also. Corn—rye treat- ments produced less dry matter than rye treatments. This is due to severely stunted corn plants and the early cutting of rye which reduced its growth. If corn and rye were intercropped in the field we would expect the dry matter yield to be nearly equal to the sum of the yields of the crops if grown seperately. TABLE 6 - AVERAGE DRY WEIGHT OF PLANT MATERIALS. SIMULATED EFFLUENT TREATMENT CROP TREATMENT 5 cm/wk 10 cm/wk 15 cm/wk 9 CORN 2.60 4.16 5.54 CORN-RYE CORN PLANTS 0.48 0.42 0.50 RYE PLANTS 1.64 2.55 3.32 TOTAL 2 12 2 97 3 82 RYE 2.16 3.06 4.32 Because of the difficulty of getting good seper- ation of soil from roots, no attempt was made to quantify the root differences between treatments. Only visual ob- servations were made. All three crop treatments had con- tinuous root systems to the base of the columns at all three irrigation levels. There were very obvious rooting differences between irrigation treatments on rye. 40 Approximately twice as many roots were in the upper 20 cm of the colums containing rye treated with 15 cm/wk as were in the columns containing rye treated with 5 cm/wk. Corn root systems were much less dense than rye. There were no visual differences in corn rooting due to increased irriga- tion rate. NUTRIENT BALANCE The main purpose of this experiment was to study the nitrogen losses in the drainage water Of each treatment combination. The nutrients applied were nitrate, ammonium, and phosphate. Nitrate concentrations of drainage water samples were determined each time samples were collected. Ammonium, phosphate, and nitrite concentrations were only measured occasionally. Under the conditions of this experi- ment it is unlikely that these nutrients would be lost in the drainage water. Phosphorus movement in soils is very limited. The phosphorus applied to the soil columns would probably be absorbed in the first few cms of soil. The concentration of phosphorus was less than 0.05 ppm in all drainage samples analyzed. Some ammonium could have been fixed by the soil or used by the plants. Most of the ammo- nium would have been oxidized to nitrate before it could reach the base of the columns. All drainage samples analy- zed for ammonium contained less than 0.05 ppm ammonium nit- rogen. Nitrite is of concern in wastewater treatment systems because it is highly toxic to higher plants and animals. Nitrite is not stable in aerobic soils. In this experiment 41 it should have been quickly converted to nitrate. Nitrite concentrations in drainage samples were always less than 0.05 ppm nitrite nitrogen. The rate of simulated effluent applied to soil columns determined the amount of nitrogen each received. The 5, 10 and 15 cm/wk treatments received 2.7, 5.4 and 8.2 mg of nitrogen per week respectively. The average nitrogen content of the simulated effluent during the eleven week experiment was 6.6 ppm. Nitrogen additions to the soil columns are explained in Table 7. TABLE 7 - NITROGEN APPLIED IN SIMULATED EFFLUENT. WEEKLY] NITROGEN TOTAL NITROGEN EFFLUENT EFFLUENT APPLIED APPLIED FOR RATE VOLUME EACH WEEK* EXPERIMENT cm/wk m1/wk mg/wk mg 5 412 2.72 29.9 10 824 5.44 59.8 15 1236 8.16 89.7 * AVERAGE NITROGEN CONCENTRATION OF SIMULATED EFFLUENT WAS 6.6 ppm. The concentrations of nitrate nitrogen in drainage samples from each treatment combination have been averaged for each week, and are presented in Table 8. Initially the average nitrate nitrogen concentra- tions in the drainage samples were very low (0.1 to 1.2 ppm nitrogen). There was a lag time before the nitrate applied in the simulated effluent leached through the soil column. 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