MSU LIBRARIES .22— \v RETURNING MATERIALS: P1ace in book drop to remove this checkout from your record. Elfl§§ w111 be charged if book is returned after the date stamped be]ow. INCREASING THE EFFECTIVENESS OF l-TRIACONTANOL APPLIED TO PLANTS By Alan William McKeown A DISSERTATION Submitted to Michigan State University in partial fulfilment of the requirements for the degree of DOCTOR OF PHILOSOPHY Department of Horticulture 1982 ABSTRACT INCREASING THE EFFECTIVENESS 0F l-TRIACONTANOL APPLIED T0 PLANTS BY Alan William McKeown Triacontanol (TRIA) is a 30 carbon primary alcohol which stimulates the growth of plants and may increase crop yield. The increase in growth and yield has been variable in greenhouse and field studies. Possible sources of this variability were investigated, in an attempt to develop practical recommendations for the use of TRIA on food crops. The factors studied included formulation, dose, pH of spray solutions, night and day temperatures, and nitrogen and phosphorus nutrition. In growth chamber and greenhouse studies, a colloidal dispersion of TRIA applied at concentrations of 0.] to ID ug/L stimulated growth of corn (E22.EEXE.L-) seedlings. These doses are l00 to 1,000 times less than the relatively low doses used with the original chloroform TWeen 20 formulation in prior research. The pH of the spray affected the response of plants to TRIA. Water suspensions with a pH of 8.0 or more were found most effective with the original formulation when applied to corn, rice (0ryza sativa L.) and soybeans (Glycine max L.) Night temperatures influenced the response of corn seedlings to TRIA. Night temperatures of l0°C and I5°C prior to TRIA application increased the response of corn to TRIA. The temperatures during the day after treatment did not alter the magnitude of the response of corn to TRIA. Field research in Michigan in I979 and I980 and in Ontario during l98l showed inconsistent yield responses in several crops. TRIA increased the yield of winter wheat (Triticum aestivum L.), sweet corn, tomatoes (Lycopersicon esculentum Mill.) and cucumbers (Cucumis sativus L.) in some tests during this period. Approximately “0% of the tests in the field with the chloroform or colloidal formulations resulted in a positive response, with lOZ for the acetone emulsion. Tomato yields varied with dose of TRIA applied to the foliage. The highest yields in the field were obtained with applications of about l0 ug/L of the colloidal dispersion. Treatment of tomato seeds with TRIA or application to roots of tomato transplants increased the yield. The same colloidal dispersion of TRIA increased the dry weight of corn at 0.I ug/L in controlled environment studies. There was a differential trend of response of corn and tomatoes to TRIA when grown at different levels of phosphorus fertility in the field. It is postulated that TRIA may be more effective on plants growing under mild stress conditions, such as low night temperatures. ACKNOWLEDGEMENTS I would like to thank Dr. S. K. Ries for his help and guidance and the members of the guidance committee, Drs. Cress, Meggitt, Price and Putnam. Additionally I wish to thank Dr. E. Everson of the Crop Science Department for his assistance with the wheat experiments in Michigan. I appreciate the technical assistance provided by V. Wert and L. Reynolds. I would also like to thank the Buckeye Cellulose Company of Memphis, TE. for the supply of TRIA used in the 1981 experiments and the Ontario Ministry of Agriculture and Food for their support during the 1981 research at Simcoe, Ontario. ii TABLE OF CONTENTS Page INTRODUCTION........ ..... . ................. . ........................... I LITERATURE REVIEW ..... ..... ... ........... . ....................... ..... 3 MATERIALS AND METHODS .... ....... ......... ............................. II Formulations ................ ..... . ................................. II Procedures for controlled environments ...................... . ...... l2 Seed treatments ...... ....... . ................. ..... ................ I3 .Field experiments .. ....... . ......... ... ............................ IA Experimental designs and statistical analysis . ..................... I7 RESULTS .................. ......... . ..... . ............................. 18 Controlled environments .............. . ............. ..... ........... I8 pH of spray .. ........... ..... ....................... . .............. I8 Night temperatures ... .............................................. 22 Dose of TRIA ..... . ....... ................... ........... . ..... . ..... 25 Seed soaks ........ ..... ..... ....... ... ............................. 25 FIELD EXPERIMENTS ..... ........................... . .................... 3i l979 ........ ............... . ........... ..... ..... ... ............... 3] i980 ............... ............... . ............. . ..... . ............ 32 I98] .... ..... . ................... ..... ....... . ..................... 35 DISCUSSION ...... . ..... ... ........... . ........... . ..................... A2 SUMMARY ....... ........................................................ 5] LITERATURE CITED ................ ......... ....... ...................... 53 iii Table 10. LIST OF TABLES Page Growth of field corn seedlings as affected by TRIA-A applied to run-off in sprays of various pH's obtained with 1 mM phosphate buffer. .......... . ................... 20 Differential growth responses of corn seedlings to TRIA-A applied to run-off in sprays of various doses and pH's with or without 10 mM phosphate buffer. ------------------ 20 Growth of corn grown in pots outdoors as affected by the pH of the spray solution and TRIA-A ------------------- 21 The response of corn seedlings to TRIA-A applied at various spray pH's in 10 mM phosphate buffer. ------------ 21 The response of soybeans to sprays to TRIA-A at different pH's. ..... ....... . ......... . ..... ..... .................. 22 The response of rice seedlings to TRIA-A in a 10 mM phosphate buffered emulsion. ...... ..... ....... .......... 22 Dry weight accumulation of corn seedlings grown at different night temperatures in response to TRIA-A at 100 ug/L in pH 8.0 1 mM phosphate buffer. ............... 24 Response of corn to pre and post treatment night temper- atures with 30°C day temperatures to TRIA-B. ... ......... 24 Growth of corn seedlings at different night temperatures and 30°C days before and after treatment with TRIA-C. .... 25 The response of 8-day old corn seedlings to dose of TRIA-A sprayed to run-off. . ......... ....... ..... . ........ 25 Page Table II. Growth of cucumbers in response to doses of TRIA-A with I mM pH 8.0 phosphate buffer sprayed to run-off. ------- - --------- 27 I2. Increased dry weight of corn seedlings treated with various volumes of sprays of I00 ug TRIA-A/L. ------------------------- 27 I3. The effect of different spray concentrations of TRIA-C ‘ applied at 350 L/ha on the growth of corn seedlings. ----- '°-- 28 IA. Germination of tomatoes after a 30 min seed treatment with TRIA-C. Lots of 50 seeds were counted after germination for 72 hr. ................. ............. ................. ........ 23 I5. Growth of tomato seedlings after treatment of the seeds with TRIA-C at different concentrations for 30 min. Seeds were sown with or without pre-germination for 72 hr. -------------- 30 I6. Differential effect of pre-germination and TRIA-C on the height of tomato seedlings 25 days after planting. ------------ 30 I7. Differential effect of treatment water temperature on the response of tomatoes to TRIA-C. ......... ..... ..... ............ 3| l8. Dry weight of tomato seedlings from seeds treated with different pH'5 of TRIA-C. ............... ....... .............. 3| l9. Increased yield of soft winter wheat treated with various volumes and concentrations of unbuffered TRIA-A. ............. 3h 20. Yield of sweet corn when sprayed to drip with TRIA-A in a pH 8.0 obtained with a I mM phosphate buffer. ................. 3h ZI. Dollar return of cucumbers treated with TRIA-A applied in pH 8.0 I mM phosphate buffer to run-off. ......... . ............. .. 35 22. Yield of soft winter wheat treated with TRIA-A and TRIA-B applied to run-off in two locations. .......................... 35 Table 23. 2h. 25. 26. 27. 28. Increase in unmarketable sweet corn in response to TRIA-C applied at the 3-Ieaf stage in different volumes of spray. There was no effect of spray volume nor an effect of TRIA-C on marketable yield. ................... Yield of tomatoes treated with various concentrations of TRIA-C. There was no difference between 3A0, 670 and l,0l0 L/ha of spray at the different concentrations. .... The weight of 25 randomly selected ripe tomato fruit as affected by dose of TRIA-C and phosphorus. .............. Marketable yield of sweet corn as affected by TRIA-C applied at 3h0 L/ha in l mM phosphate buffer and pH of spray at pH 5.0 and 9.0. ........ . .................... Yield of tomatoes in response to TRIA-C applied to the roots for 5 min. .. ..... . ....................... . ........ Tomato yield from plants grown from TRIA-C treated seeds that had been direct-sown or grown for transplants. ..... vi Page ... 37 ... 38 ..._38 '° #0 .. AI INTRODUCTION The world is facing an ever-increasing population, consequently an ever- increasing demand for food. A major problem is to supply sufficient food for the increasing population. It is important to investigate the potential usefulness of any compound or technique which may improve the yield and/or quality of crops. Triacontanol (TRIA) is one such compound. Applications of a few mg/ha have increased crop yield. TRIA is a 30 carbon primary alcohol first isolated as a component of alfalfa leaf wax by Chibnall et al (7). Its ability to stimulate plant growth was first reported by Ries (22). Since the original discovery of plant growth regulating activity TRIA has been shown to stimulate the yield of several annual crop species (2I, 2“). Single foliar applications of TRIA at concentrations of 0.0l to ID mg/L to seedlings increased the yields of sweet corn (Zea_mgy§_L.) cucumbers (Cucumis sativa L), soybeans (Glycine max L.) and wheat (Triticum aestivum L.) an average of IZZ. Little information is available on the effect of TRIA on yield components or quality, but TRIA may stimulate plant growth. The implications of consistent increased crop yields resulting from very small amounts of TRIA are enormous. However, plants have not consistently responded with increased yield or bio-mass production to applications of TRIA under field or greenhouse conditions. The reasons for this failure to consistently stimulate yield or bio-mass production are not clear. Furthermore, the exact environmental conditions, 2 TRIA concentrations, timing and method of TRIA application and other factors required to obtain increased yields have not been delineated. However, if techniques are developed so that consistent yield increases could be obtained, then TRIA would be an excellent management tool for increasing crop productivity. The objectives of this research were to: (l) Investigate the effect of the pH of different formulations on the response of plants to TRIA. (2) Investigate types of formulation and doses of TRIA. (3) Evaluate the effect of pre and post treatment temperatures on the response of plants to TRIA. .The major problem delaying the commercial use of TRIA in crop production both in the field or greenhouse is this lack of a consistent response. This was also the central problem in this research. The major difficulty in this study was to find a protocol that would consistently reproduce the response of plants to TRIA, so that desired environmental and physical variables could be investigated in detail. Much emphasis was placed on environmental factors in order to develop such a protocol. LITERATURE REVIEW Triacontanol (TRIA, CH3 (CH2)28 CHZOH) is a 30 carbon primary alcohol which was identified as a component of alfalfa (Medicago sativa L)t leaf wax by Chibnall et al in I933 (7). Coarsely chopped alfalfa hay banded at ll7 kg/ha was reported by Ries et al (22) to increase the yield of tomatoes (Lycopersicon esculentum Mill.) by l0 tonne/ha. The yields of cucumbers (Cucumis sativus L.) and lettuce (Lactuca sativa L.) were also Increased by this treatment. The increase in yield was greater than that expected from the added nitrogen from the alfalfa. It was subsequently demonstrated that a chloroform extract of alfalfa hay as well as ground alfalfa applied pre-plant to the soil increased the dry weight of corn (Eeg_mgy§_L.) seedlings grown in a greenhouse (22). The chloroform extract added at the equivalent of 0.I to l0 mg/L of emulsion also increased the growth of rice (Oryza sativa L.) grown in solution culture. TRIA was isolated and identified as the active component of the extract (22). Synthetic TRIA, both when applied in the nutrient solution to rice plants and to the foliage of corn seedlings grown in soil increased the dry weight compared to untreated plants (22). The response of plants to TRIA under laboratogy conditions. The overall response of plants to TRIA is characterized by a very rapid increase in growth rate as measured by dry weight accumulation and increases in leaf area (23). Ries and Hert (23) demonstrated increased dry weight and leaf area of rice plants in solution culture within 3 hr of treatment with ID ug/L of TRIA in the nutrient solution. Kjeldahl-detectable nitrogen I. also increased due to the treatment. They showed that the net assimilation rate (NAR) of rice was highest the first 8 hr after treatment; however, there was no effect on the NAR after 2h hr (23). This suggests a very short-term direct effect of TRIA on plant metabolism. Ries and Wert (23) showed that the increase in plant dry weight would occur when rice was treated for 6 hr in the dark. This evidence suggests that plant response to TRIA is not wholly dependent directly on the photosynthetic pathway, and that metabolism of a stored product may be involved. If the growth response to TRIA depends on the metabolism of a stored product, then the manipulation of night temperatures should induce differences in the response. Lower night temperatures would lead to greater food reserves, hence a larger pool available to respond to TRIA application. Effects of night temperatures on this possible TRIA effect have not been investigated. Bittenbender et al (2) further investigated the dark response of rice to TRIA. Carbon dioxide was shown to have an apparent regulatory role on dry weight gain in the dark in response to TRIA (2). Rice plants did not respond to TRIA in the dark when the C02 level around the plant was less than 50 or greater than 360 uL/L. The maximum dark response occurred when C0 levels were l00 to 200 uL/L, i.e. 2 less than ambient atmospheric levels. The effect of C02 on the daytime response of plants to TRIA is not known. The role of C02 in the response of plants to TRIA remains unclear. Eriksen et al (8) demonstrated a 30% increase in dry weight of tomatoes grown in solution culture after 2% days when l00 ug/L TRIA was added to the solution before flower bud formation. No effect of TRIA was observed with a single application; TRIA needed to be supplied with every change of the nutrient solution. No effect on corn was observed in solution culture 5 with similar treatments (8). In these experiments, Eriksen et al also observed an apparent effect on the photosynthetic system of tomatoes (C3 plant) treated with TRIA under several concentrations of oxygen. Photosynthetic and 2% 0 . 2 2 Net photosynthesis was reduced by 39% in control plants and 27% in TRIA- efficiency of treated and control plants was compared at 2l% 0 treated plants when compared at 2l% vs 2% (8). This would indicate more available photosynthate in TRIA-treated plants. There was no effect of TRIA on photosynthesis of corn (8). The results agree with those of Ries and Nert (23) who showed no apparent effect of TRIA on rice (Ch plant) photosynthesis. It may be possible that TRIA affects C and Ch plants 3 differentially. Short-Term biochemical responses of plants to TRIA. In laboratory experiments rapid changes in metabolism induced by TRIA have been demonstrated. For example, changes in Kjeldahl-detactable nitrogen have been reported within 80 min (II, l3, IA, 25). Ries et all (25) showed increased in KjeldahI-detectable soluble protein, free amino acids and reducing sugars within A min of treatment. Dry weight was shown to increase in ID min and leaf area within 20 min (25). As many of these experiments were 80 min in duration, the long-term biochemical effects of treatment with, if any, TRIA are not known. The relationship of these short-term biochemical events with long-term effects of TRIA on plants remains to be demonstrated. Houtz (II) has shown that phosphate uptake from the nutrient solution increased as a consequence of treating rice with TRIA. Ramani and Kannan (20) demonstrated a TRIA-induced increase in uptake and transport of phosphate and rubidium ions by sorghum (Sorghum vulgare L.) grown in solution culture. Drought resistant cultivars of sorghum absorbed more phosphate and rubidium ions than did drought susceptible cultivars (20). 6 It is evident that TRIA influences the uptake of some mineral ions, and that cultivar differences exist. Bittenbender et al (2) showed that the TRIA response of rice was not influenced by the source of nitrogen (NH: vs N03). Singletary (I8) demonstrated with soybeans (Glycine max L.) that there was no effect on nutrient uptake grown in solution culture. He found however, that TRIA at I0 ug/L increased the growth of corn seedlings grown in solution culture with half of the normal N and P levels compared to those grown in full strength solutions. Clearly, there is an effect of TRIA on the short-term nutrient status of the plant. However, the effect of the plants' initial nutritional status on its ability to respond to TRIA needs clarification. 'Formulations of TRIA. Originally, crystalline TRIA was dissolved in chloroform to make a stock solution. Aliquots of this stock were added to water containing 0.l% v/v Tween 20 to form a stable suspension (23). An acetone-CaCl -water emulsion developed by Heliber (26) was used for a 2 period of time In I980. In l98l, a stable colloidal dispersion of TRIA was developed by Laughlin et al (IA). This formulation of TRIA has several advantages: it is an aqueous dispersion, virtually eliminating problems of safety as compared with using organic solvents. It is prepared under controlled conditions, as opposed to previous formulations of chloroform or acetone. Furthermore, the colloidal dispersion of TRIA has been shown to be as much as l,000 times as effective as the original TRIA-A formul- ation in short-term studies with rice and corn (IA). This formulation of TRIA has been shown to stimulate the growth of rice in solution culture when applied at concentrations as low as l ng/L (2.3 x IO-IZM) (IA). These minute quantities are about the physiologically active endogenous levels of known naturally-occurring plant hormones. This suggests a profound physiological role for TRIA in the plant. 7 In fact, the concentrations of applied TRIA that have elicited responses in plants are much lower than those used for other exogenously applied growth regulators, often applied at concentrations up to several hundred ppm. This suggests that TRIA has a very potent effect on plant metabolism. Jones et al (I2) showed that other long-chain hydrocarbons such as octacosanol (CH3 (CMZ)26 CHZOH) were powerful inhibitors of the activity -IZM was shown to inhibit TRIA applied to of TRIA. Octacosanol at 2.A x IO rice at 2.3 x l0-8M (l00 ug/L) (l2), thus small amounts of impurities in the formulation may drastically effect the activity of the TRIA. However, the nature of the type of inhibition is unknown. Response of plants to TRIA under field and gleenhouse conditions. Single foliar applications of TRIA ranging from 20 mg/ha to 2.2 g/ha increased marketable yields an average of l3% for sweet corn, IA% ($/ha) for cucumbers (more fruit/plant), l0% soybeans, l7% for tomatoes (more early fruit) and 8% for wheat (2l, 2A). Howeveer, in these studies an effect of TRIA could not be consistently obtained in every experiment. Also the optimum concentration of TRIA was not determined. Bouwkamp and McArdle (A) showed that TRIA applied at a concentration of l00 ug/L to sweet potatoes (Ipomoea batatas L.) in the field had no effect on yield of roots at harvest. Leaves were sampled at IA hr, 5 and 57 days after treatment; weighed and analyzed for nitrogen. The dry matter of leaves was increased by TRIA at IA hr and 5 days, while % nitrogen increased after 5 days. There were no differences at harvest. It is evident that a short-term effect of TRIA was obtained but this effect was not reflected in an increased yield of sweet potato roots. 8 Bosland et al (3) did not obtain an increase in yield or change in soluble solids of 'PMR-AS' muskmelons (Cucumis melo L.) when plants were treated in the 8 to l0-Ieaf stage with 0.0l to l0.0 ppm TRIA as a foliar spray. This stage of growth is later than that used by Ries et al (2A) when TRIA increased the yield of cucumbers, indicating that age may be a factor in the lack of response. In greenhouse experiments TRIA increased the dry weight of cucumber, field corn, soybean and carrot (Daucus carota L.) seedlings 3A%, 3A%, I9% and 65% respectively of TRIA at concentrations of 0.0l to ID mg/L (2A). However, consistent responses to TRIA were not obtained and optimum doses and conditions were not defined. Pocock (l7) reported variable responses in potted sugar beets (Beta vulgaris L.) with plant age and concentration of TRIA. TRIA applied at concentrations of 0.00I to l mg/ha when the first tree leaves are expanding decreased the growth of beets by l0%. However, the dry weights increased when TRIA was applied after the first two leaves were fully expanded. Three weeks after treatment these plants had increased dry weight by 30% as compared to the controls (I7). There was no effect of TRIA on sugar concentration at harvest (l7). Application of TRIA to the seeds of several species by soaking in TRIA dissolved dichloromethane mixtures has stimulated dry weight production in greenhouse studies an average of 50% (2A). No effect of TRIA could be found with direct-seeded crops in the field (2A). However, marketable yield of tomatoes was increased 27% when grown from transplants produced from treated seeds (2A). Reasons for this lack of consistency are not known. Ries et al (2A) investigated the response of wheat, corn and tomato seedlings from treated seeds grown at night and day temperatures of l0/ISOC, lS/ZOOC, 20/250C and 25/300C. Percent dry 9 weight increases over control and temperature were linearly related, indicating that temperature influenced the response. Increases due to the temperatures studied varied from 20 to 80%, indicating that temperature influenced the magnitude of the response to TRIA. Henry and Primo (9) reported increased bio-mass of Il-day-old 'Great Lakes' lettuce, but not 'Grand Rapids' grown in solution culture in a growth chamber indicating a cultivar difference in response to TRIA under these conditions. Charlton et al (6) reported no effect of TRIA at 0.I to l00 ug/L on several analogues applied to the seeds of durum wheat in greenhouse experiments. Hoagland (l0) demonstrated no effect of TRIA at l0-SM on seed germination of IS crop and weed species. As with the effect of TRIA on crops grown in the field, there are incon- sistent reports of its effect on growth in shorter germ greenhouse studies. The reasons for this lack of consistency are not clear. General comments on the action of TRIA on plants. Since the original discovery of the growth stimulating activity of TRIA, various reports (l7, 2i, 2A) show increased long-term growth, and A show no effect (3, A, 6, 8). Increased yield was not obtained in every field experiment (21). No successful responses on crops grown as perennials have been reported to date. Little is known about the effect of TRIA on various yield and quality parameters. Reasons for the inconsistent responses of plants to TRIA are not clear. Climatic, soil conditions, species, cultivar, concentration and formulation and formulation quality may all influence the ability of plants to respond positibely to TRIA. The optimum dose of TRIA, time of application, stage of growth and the mode of application required to obtain the maximum response of plants all remain undefined for any l0 species. In addition, another unknown factor is the effect of spray pH on the plants' response to TRIA. A low pH has been shown to increase the uptake of NAA (l6). However, the effect of pH on the physiological activity of long-chain alcohols is unknown. The action of TRIA on plants has been most consistent in short-term lab oratory studies (2, ll, l2, l3, IA, 22, 23, 25). Consistent increases in dry weight, Kjeldahl-nitrogen, free amino acids and reducing sugars have been obtained in rice and corn seedlings in short-term biochemical studies (ll, I3, 25). Short-term biochemical responses to TRIA in these studies have been shown to be rapid, in some cases a matter of minutes. Reasons for the inconsistency between field, greenhouse and laboratory studies are not known, and the relationship between observed biochemical responses and total plant behaviour remain unclear. Since yield is a function of the total integrated plant-environmental interaction during its life, it is difficult to evaluate the importance of short- term, transitory biochemical effects on long-term growth and yield. TRIA can be a plant growth stimulant. However, little is known about the conditions required to induce positive effects on the yield of crops. If a means is found to consistently increase yields of crops when treated with TRIA, then it may become an important management tool in crop production. MATERIALS AND METHODS Formulations. A solution of TRIA was prepared with the TRIA dissolved in chloroform (l)mg/l0 ml). Aliquots were added to distilled water containing Tween 20 (polyoxyethlene sorbitan monolaurate) 0.l% v/v as a surfactant (23). This emulsion was heated to drive off the chloroform, and is referred to as TRIA-A. TRIA was also prepared by dissolving in acetone, aliquots of one which were added to a 2% acetone/ water emulsion (26). This will be referred to as TRIA-B. The colloidally-dispersed formulation (IA) was added directly to distilled water and will be referred to as TRIA-C. All control treatments were made as their individual formulations, but without TRIA. A l mM or ID mM phosphate buffer was used to obtain various Spray pH's with the TRIA-A formulation. Sodium hydroxide and sulphuric were used to adjust the pH of the TRIA-B and TRIA-C formulations. Typically, TRIA was applied to the run-off point, using a glass 250 ml chromatography sprayer (A.H. Thomas Co., Philadelphia, PA) with air as the pr0pellant. Concentrations of TRIA tested ranged from 0.I to l0,000 ug/L. ll l2 Procedures in controlled environments. Greenhouse experiments were conducted under the following environmental conditions, supplemental lighting, both fluorescent and High Intensity Discharge lamps (67 W/m2 at 1.2 m), the photoperiod was l6 hr, the night temperature was approximately ZIOC. 'Heinz I350' tomatoes and 'Greenstar' cucumbers were grown in ID cm clay pots and 'Pioneer 3780' field corn in I8 cm clay pots. A l:I:l soil mix of sand, peat and loam was used for the period from March 1979 to September I980 and a I:l soilzpeat mix was used after this time. Nutrients were applied twice per week at 250 ml per pot (IOO ml for ID cm pots) with l.6 g/L solution of I2:2I.l:6:6 (NszK) or I g/L of 20:8.8:I6.6 soluble fertilizer. All water was applied in measured amounts once the experiment was started. Plants were thinned to A uniform seedlings per pot and carefully blocked for size in all experiments. Typically corn was treated at the early three-leaf stage, 6 to 8 days after planting, depending on time of year and harvested 7 days later. 'Corsoy' soybeans were sprayed when the first true leaves appeared. The procedures for the rice experiments were previously described (23). Plants were dried to a constant weight at 70°C prior to weighing. In the growth chamber studies night temperatures of IOOC, ISOC, 20°C and 25°C nights were used and the day temperatures were maintained at 30°C. For several of the night temperature experiments, corn heat units (CHU) were calculated using the formula of Brown (5). Plants were treated at the number of CHU accumulated to the third-leaf stage of corn grown with 22°C nights. Plants grown at 22°C nights were harvested after 5 days (A30 CHU); plants from the other treatments were harvested after the same heat units had accumulated. l3 Plants were watered by hand with 250 mL 2 times per week. Fertilizer was applied as in the greenhouse eXperiments. Light levels were measured widuaLambda Li I85 with Quantum Sensor (Lincoln, Ne.) and a Weston F56 sunlight illumination meter (Newark, N.J.) and were l0,000 lux at 0.75 m at East Lansing and Simcoe respectively. Sixteen-hour day and 8-hour nights were used for most experiments. After the end of March l98l, both controlled environment and field research was conducted at the Horticultural Experiment Station, Simcoe, Ontario. In l98l all of the TRIA used was the colloidalIy-dispersed TRIA-C. Corn experiments were similar to those at M.S.U. with the exception that IS cm plastic pots and peat soil mix (Metro 200) were used and the sprays were applied at 350 L/ha with a trigger-type atomizer. Seed treatments. Seeds of 'Earlibright' tomatoes were treated for 30 minutes in surfactant and concentrations of TRIA-C, ranging from 0.00I to l.0 mg TRIA/L. The surfactant control was 0.I mg/L. Seeds were removed at the end of treatment and dried overnight at room temperature. For pregermination, seeds were placed in I.5 kg plastic freezer bags with 250 mL of high resistance deionized water. At the end of 2 hr the water was drained off and approximately l00 mL of fresh, deionized water was added. The bags were filled with air and sealed with a twist tie and the seeds germinated at room temperature (ZIOC) for 3 days. In order to investigate the possibility that with the colloidal dispersion, the wax on the seed coat was preventing TRIA from reaching the embryo, the seeds were soaked 30 min In DCM with constant stirring. After soaking, the seeds were dried overnight at room temperature. l4 Field experiments. Normal cultural practices for Michigan were used in Michigan in I979, I980 and those recommended for Ontario were used in l98l. Cultural practices in both areas are similar. Wheat experiments in I979 were conducted at both East Lansing and Saranac, Michigan. The East Lansing eXperiment used several TRIA-A treatments applied at 2 different stages of growth on A cultivar combinations and doses of TRIA-A. The following cultivars of soft winter wheat were used in 1979: 'Augusta', 'Tecumseh', 'Ionia' and 'Genesee'. 'Ionia' and 'Genesee' were lost due to severe lodging. Treatments were applied to drip in the test at Saranac in order to wet the plants thoroughly. It was found that CO as a prepellant lowered the spray solution pH, 2 therefore all further experiments were sprayed with a pump-up compressed air Sprayed to avoid this complication. Further experiments in I979 and I980 were sprayed to the run-off point to completely wet the plants, as with the greenhouse experiments. Experiments were conducted to elucidate the Optimum dose of TRIA-A for 'Gold Cup' sweet corn and 'Greenstar' cucumbers. Ears of sweet corn of marketable size were picked by hand, weights were recorded before and after husking. Pickling cucumbers were harvested once-over and graded based on Pickle Improvement Committee of the International Pickle Packers Association standard prices. Yield was converted to $/ha as follows: fruit up to 2.7 cm in diameter was given a value of $l3.25/l00 kg; 2.7 to 3.8 cm $6.23; 3.8 to 5.l cm $A.A2; 5.l to 5.7 cm $2.2; 5.7 to 6.A $l.ll; greater than 6.A cm $0.55. 'Meinz I350' tomatoes were harvested by hand and the weight of ripe marketable fruit was recorded. IS 'Gold Cup' sweet corn and 'Greenstar' cucumber experiments were conducted at the East Lansing and at the Carksville Horticultural Research Stations in I980 to investigate the effect of low nitrogen and high phosphorus nutrition on the response of creps to TRIA. The Dryden sandy loam soil at Clarksville has a higher natural fertility than the Spinks sandy loam soil at East Lansing. Similar rates of nitrogen (80 kg/ha) and phosphorus (I60 kg/ha) were applied to the corn and cucumbers at both locations as a sidedressing. 'Pikred' tomatoes were hand-weeded as it was felt that the stress induced by cool, drying winds may have weakened the plants, and possibly predisposed them to injury from the herbicide in combination with the TRIA treatments. Harvest methods in I980 were similar to those used In I979. In I980, eXperiments were conducted at East Lansing, Saranac and Tuscola County with 'Augusta' and 'Frankenmuth' soft-white winter wheats. Different rates of TRIA-A and TRIA-B were compared. Plots were sprayed at the early jointing stage and prior to anthesis. The East Lansing plot was lost due to winter Injury. An experiment was conducted to evaluate the effect of phosphorus (0.5% P) applied to the leaves and TRIA at the early filling state of wheat. Several cool season crops were screened for response to TRIA. It was hypothesized that the cooler night temperatures in the spring may favor the response. 'Improved Progress' peas (Pisum sativum L.) were direct-seeded, cole craps, 'Market Prize' cabbage (Brassica oleracea L. var capitata), 'Waltham' broccoli (E, oleracea L. botrytls group), 'Imperial I0-6' cauliflower (E, oleracea L. botrytls group), 'Early hybrid 6' chinese cabbage (E, raga L. chinesis group) were started in l6 the greenhouse and transplanted to the field. In l98l field research was conducted at the Horticultural Experiment Station, Simcoe, Ontario. The l98l experiments were designed to find the optimum dose of TRIA-C to use on 'Snow Crown' cauliflower, 'Earlivee' sweet corn and direct-seeded 'Veepro' tomatoes. Experiments were conducted to evaluate effects of nitrogen and phosphorus fertilizer and the dose of TRIA on sweet corn and tomatoes. Nitrogen at A5 kg/ha was applied pre-plant and 20 kg/ha of phosphorus was banded after planting. Plots were split for an additional A5 kg/ha N at the recommended time of application for both craps to evaluate the effect of 'low' and 'high' nitrogen. In addition, an experiment with or without starter fertilizer (l0:23:8) at l kg/l00 L to the roots of bare root 'Earlibright' tomato transplants. Tomato seeds were treated with TRIA-C as in greenhouse eXperiments and grown for transplants or direct-seeded. The two direct-seeded tomato eXperiments were harvested using 3 Hart- Carter harvester. For multipick hand-harvest experiments, tomatoes were picked at or beyond the breaker stage and graded according to size. The large were over 5.0 cm and the small 2.5 to 5.0 cm in diameter. At the final harvest, vines were stripped of all fruit over 2.5 cm diameter to obtain an estimate of total fruit yield. Cauliflower was harvested 3 times per week. At harvest, the trimmed head was weighed and the florets removed and weighed. Sweet corn of marketable size was harvested by hand. The number of harvested plants were recorded and number of ears/plant were calculated. After weighing, the husks were removed, the cobs graded as marketable or unmarketable and reweighed. A cob was considered unmarketable if it was short (under IS cm), unfilled, diseased or damaged. 17 Experimental desiggs and statistical analysis. Experiments were usually arranged in randomized complete block designs. Split plot designs were used in some field experiments. Analysis of variance was conducted on all data using apprOpriate statistical methods (l9). Trend analyses or orthogonal/non-orthogonal comparisons were made where relevant. RESULTS CONTROLLED ENVIRONMENTS Spray pH. After observing a decrease in spray pH after spraying with the C02 propelled sprayer, an experiment was conducted to evaluate the effect of pH on the response of corn to TRIA-A. There was a differential growth response of corn treated with TRIA-A applied in various pH buffered sprays (Table l). TRIA-A did not stimulate growth of corn seedlings when applied at a pH of 6.8, reduced growth when applied at pH 7.7 and stimulated growth at pH 8.0. In a further experiment TRIA-A differentially affected the growth of corn seedlings when applied in buffered or unbuffered sprays usingcompressed air or C02 as the propellant (Table 2). The source of propellant had no effect other than that induced by changing the pH of the spray. The increased growth of corn seedlings in reSponse to concentration of TRIA-A in pH 8.0 sprays was best described as a quadratic trend. The decreased growth of corn to TRIA-A treated with low pH sprays was best described as a linear trend. Furthermore, the growth of corn was reduced by the high pH sprays. No effect of spray pH on the response of corn to TRIA was found when the plants were grown outdoors (Table 3). In another experiment there was no effect of pH on the growth of corn treated with TRIA-A (Table A). In one experiment TRIA-C, applied in a spray at pH 9.0 (356 mg/plant) stimulated the growth of corn seedlings, as compared to controls (326 mg/plant), but did not stimulate growth when applied at pH 5.0 (3A6 mg/plant) or 7.0 (336 mg/plant). Controls sprays at a pH of 8.0 18 l9 Table l. Growth of field corn seedlings as affected by TRIA-A applied to run-off in sprays of various pH's obtained with phosphate buffer. Dry Weight (mg/shoot)z pH TRIA-A (IOO ug/L) 6.8 7.7 8.0 - A28 A52 379 + AA2 » 359*"x A68** z The F value of the interaction for pH x TRIA was significant at the I% level. x**The F values for the comparison of the control with TRIA pH 7.7 and 8.0 were significant at the I% level. Table 2. Differential growth responses of corn seedlings to TRIA-A applied to run-off in sprays of various doses and pH's with I0 mM phosphate bufferz. Dry Weight Treatments (mg/shoot)Y TRIA-A (ug/L) Average V Buffer Post spray pH 0 l00 l000 - 6.I A58 A37 A2A** + 8.0 A00 AA3 A32“w 2 Carbon dioxide or air was used as the spray prepellant. Y The F value for the interaction of pH x TRIA-A was significant at the I% level. x* The F value for the linear trend of TRIA-A with dose was significant at the 5% level. ‘ w**The F value for the quadratic trend of TRIA-A with dose was significant at the I% level. 20 Table 3. Growth of corn in pots outdoors as affected by the pH of the spray and TRIA-A. Phosphate buffer TRIA-A Dry weight?"2 pH (l mM) (l00 ug/L) (mg/shoot) 6.3 O 0 38] 6.3 O IOO A09 8.0 + 0 392 8.0 + '00 AAI 2*The F value for TRIA was significant at the 5% level. There was no significant effect of pH on the growth of plants. Table A. The response of corn seedlings to TRIA-A applied at various spray pH's in ID mM phosphate bufferz. Dry weight*Y Treatments (mg/shoot) pH 3.0 30A 7.0 30l 8.0 33] TRIA (ug/L) 0 297 ICC 328 2 There was no effect of pH on the response of plants to TRIA. Y*The F value for TRIA was significant at the I% level. 21 Table 5. The response of soybeans to sprays of TRIA-A at different pH's. Phosphate buffer TRIA Dry weight pH (I mM) (ug/L) (g/shoot)z 6.3 0 0 I.l6**Y 6.3 0 l00 l.0A 8.0 + O l.06 8.0 + l00 l.l9** z The F value for pH x TRIA was significant at the 0.05% level. Y*,**The F values for the comparison of pH 6.3 control with pH 8.0 control was significant at the I% level. The F value for the comparison of control with TRIA at pH 8.0 was significant at the 5% level. Table 6. The response of rice seedlings to TRIA-A in a l0 mM phosphate buffered emulsion. TRIA Dry weight?"2 pH (l00 ug/L) (mg/plant) 5.6 0 76 5.6 + 79 8.0 0 72 8.0 + 87 zIltThe P value for TRIA is significant at the 5% level. 22 decreased the growth of soybeans (Table 5). TRIA-A applied in sprays of pH 8.0 overcame this inhibitory effect. Growth of rice seedlings was stimulated by TRIA-A, the greatest increase occurred when treated in pH 8.0 emulsions (Table 6). Phosphate buffers were not used with TRIA-B as precipitates formed when the emulsion was prepared. Night temperatures. Growth of field corn seedlings increased in response to treatment with TRIA-A at 10°C and 15°C nights (Table 7). Additionally, plant dry weight was increased by growing in the lower night temperatures. Plants grown with l0°C nights often had injury on the leaves. This may have been due to guttation fluids or chilling. Several other tests were conducted with similar temperature regimes; however, little effect of TRIA was obtained in these experiments. When night temperatures were changed after spraying with TRIA-B, the greatest response occurred with plants grown at l5°C night temperatures before treatment (Table 8). Plants grown at ISOC nights continuously were the largest but had a lower percent increase than those receiving 25°C nights post- treatment. In a similar experiment conducted with TRIA-C, night temperatures of ISOC (30°C days) favoured the response, while pre-treatment night temper- atures of 25°C inhibited the response of TRIA (Table 9). In further experiments day temperatures were investigated as well as spray dispersion temperature. Plants grown at 30°C or 35°C day (ISOC nights) were larger than those grown in the greenhouse. There was no effect of day temperature on the response of corn seedlings to TRIA-C. TRIA-C was applied in sprays at 20°C, 50°C to test the hypothesis that solution temperature would influence the response of TRIA. At a 30°C solution temperature, 0.I ug TRIA-C/L inhibited the growth of corn. This effect, however, could have been due to the warm dispersion rather than TRIA. 23 Table 7. Dry weight accumulation of corn seedlings grown at different night temperatures inzresponse to TRIA-A at I00 ug/L in pH 8.0 l mM phosphate buffer . Dry weighty (mg/shoot) Day/NightoTegperatures Control TRIA ( C) 30/10 510 6A8**x 30/l5 383 AAO* 30/22 AlO AlO 2Plants were harvested after approximately A30 corn heat units had accumulated. yThe F value for temperature linear and quadratic were significant at the 5% level. X *,**The F values for the comparison of control with treatment at the respective temperatures were significant at the 5% levels respectively. Table 8. Response of corn to pre and post treatment night temperatures with 30 C day temperatures to TRIA-B. Dry weight2 (mg/shoot) Night temperature 0C TRIA-B (Pre/post treatment) (I00 ug/L) ' I5/l5 l5/25 25/l5 25/25 - A88 353 A73 358 + 508 393 A85 388 2The F value for the interaction of pre x post night temperatures and TRIA-B was significant at the 5% level. 24 Table 9. Growth of corn seedlings at different night temperatures and 30 C days before and after treatment with TRIA-C. Dry weight2 (mg/shoot) Night temperaturecC (Pre/post treatment) TRIA-C (1.0 ug/L, 350 L/ha) I5/I5 I5/25 25/15 25/25 - 323 360 370 378 + 3514*y 395* 352 38k 2The F value for pre treatment x TRIA-C was significant at the 5% level. Y*The comparison of control vs TRIA-C with l5°C nights prior to treatment was significant at the 5% level. Table ID. The response of 8-gay-old corn seedlings to dose of TRIA-A sprayed to run-off . TRIA-A Dry weight *Y (pg/L) (mg/shoot) 0 682 I00 848 I000 7A6 zUnbuffered spray. y*The F value for quadratic trend of dose of TRIA significant at the 5% level. 25 Dose of TRIA. Over the course of this work many experiments were conduc- ted to find the optimum concentration of TRIA to use with all 3 formulations both when sprayed to drip or at set volume per unit area. The response of corn seedlings to applications of increasing concen- trations of TRIA was best described as a quadratic curve with IOO ug/L of unbuffered TRIA-A being maximal (Table ID). The response of cucumbers exhibited a cubic trend, with the largest increase from I00 ug/L (Table II). Decreased growth resulted with ID ug/L TRIA-A. The response was also found in the stem, cotyledon and the first true leaf. No effect was found in the second or third (both still small and expanded) leaves. In one of the pH experiments IOO ug/L of TRIA-A applied at pH 8.0 proved superior to l,000 ug/L (Table 2). The growth response of corn to unbuffered spray of TRIA-A at l00 ug/L applied in various volumes induced a growth response best described as a quadratic trend (Table l2). The largest increase in dry weight of corn occurred between AIO and 820 L/ha (A0 to 80 mg/ha) of spray applied. In a test with TRIA-C applied in concentrations of 0 to I00 ug/L at 350 L/ha, 0.I ug/L TRIA-C stimulated growth (Table I3). It appears that the concentration of TRIA-C required to elicit the response is less than the optimum for TRIA-A (l00 ug/L). Other evidence for this increased effectiveness of TRIA-C was apparent in the studies with night temperatures when TRIA-C stimulated growth at l ug/L (Table 9). Seed soaks. TRIA increased the rate of germination of tomato seed as measured by emergence of the radicle after 72 hr (Table IA). However, after planting in the greenhouse, this effect was lost. In another similar experiment there was no effect on germination. However, TRIA-C at 0.I mg/L increased growth measured by height and stem diameter after l7 days and dry weight and height at harvest (25 days) (Table IS). The effect on stem 26 Table II. Growth of cucumbers in response to doses of TRIA-A with l mM pH 8.0 phosphate buffer Sprayed to run-off. TRIA-A Dry weight*2 (ug/L) (mg/shoot) 0 527 '0 AAA IOO 560 1,000 539 2*The F value for cubic trend to dose of TRIA is significant at the I% level. Table l2. Increased dry weight of corn seedlings treated with various unbuffered volumes of spray of l00 ug TRIA-AIL. Spray volume TRIA-A Dry weight”2 (L/ha) (l00 ug/L) _ (mg/shoot) l,000 0 356 200 . + 387 A00 + hlo 600 + AOI 800 + hog 1,000 + 353 2“The F value for quadratic trend to dose of TRIA is significant at the I% level. 27 Table l3. The effect of different spray concentrations of TRIA-C applied at 350 L/ha on the growth of corn seedlings. TRIA-C Dry weight (Hg/L) (mg/shoot) O 380 0.I Al2*z l.0 397 I0.0 382 I00.0 388 z'IrThe F value for the comparison of control with 0.I ug/L TRIA-C was significant at the 5% level. Table IA. Germination of tomatoes after a 30-min seed treatment with TRIA-C. Lots of 50 seeds were counted after germination for 72 hr. TRIA-C Germinationz (mg/L) 2 Dry (untreated) A l2 Surfactant 35 0.0] 52 0.I 7i l.0 A6 2The F value for the following comparisons were significant (% level): untreated with surfactant (l%); surfactant with 0.0I mg/L TRIA-C (5%); surfactant with 0.I mg/L TRIA-C (l%); TRIA 0.01 mg with 0.1 mg/L (5%). 28 diameter was lost by harvest, Larger plants were obtained from pre- germinated seeds. Furthermore, there was a differential effect of pre- germination and TRIA-C on the height at harvest (Table I6). Plants from pre-germinated seeds treated with 0.I mg/L were shorter than those at 0.0I mg/L. However, this effect did not exist after l7 days, indicating that the effect was lost in the 6 days prior to harvest. In the first 2 experiments, excess seed was treated and the remainder saved and planted at a later date. There was no effect of TRIA-C on seeds held IT”‘ 6 weeks at room temperature. Seeds from experiment 2 were planted in 2 locations, to investigate the ability to 'hold' the response to TRIA and any differential response in the greenhouse and growth room. There was no effect of TRIA at either location. The soaking time for seed treatment of tomatoes with TRIA-C dispersions was investigated; however, there were no effects of TRIA-C due to time of seed treatment. The lack of consistent effects of TRIA in tomato seed treatments indicated that seed size and indirectly, vigour might influence the seeds' ability to respond to TRIA. Visually, in most cases, plants would show responses; however, there was a lot of interplant variability which may have masked the response. In 2 experiments, seed was sized and showed that there was no interaction of seed size with TRIA on the growth of the seedlings in one test. In this test, tomato seed about 3.I mm in diameter treated with TRIA-C resulted in larger plants after ll days. However, the effect was not detectable l7 days after treatment. In another experiment, there was an increase in seedling dry weight for the larger seed sizes. In some of the treatment combinations the TRIA-C reduced the height of the tomato seedlings, producing stockier plants which would be better for transplants than the control plants. 29 Table 15. Growth of tomato seedlings after treatment of the seeds with TRIA-C at different concentrations for 30 min. Seeds were sown with or without pre-germination for 72 hr. TRIA Dry weight Stem diameter Seed Treatment (mg/L) (mg/shoot) (mm) 0 o 379 A.5 Water 0 323 A.6 Surfactant 0 350 A.8 Surfactant 0.0I 387 A.9* Surfactant 0.I AA8=Iiz 5.l* Surfactant l.0 39l 5.0* 2*The F value for the comparison of TRIA-C with controls was significant at the 5% level. Table I6. Differential effect of pre-germination and TRIA-C on the height of tomato seedlings 25 days after planting. Height (cm) TRIA (mg/L) Pre- 0 0 0 0.0I 0.I l.0 germination (Dry) (H20) (Surfactant) 0 l0.l 9.8 l0.5 ll.2*z l2.A** l0.8 + I0.9 l0.6 l0.A l2.0* Il.0 ll.6 Z*,**The F value of the comparison of treatment with mean of control is significant at the 5% and l% levels respectively. 30 Table I7. Differential effect of treatment water temperature on the response of tomatoes to TRIA-C. Dry weight (mg/shoot)z TRIA (mg/L) Temperature 0C Surfactant 0.0l 0.I l.0 20 375.A 377.9 3I6.7*y 3l8.3* 50 360.A 363.3 370.0 362.9 z The F value for treatment temperature x TRIA-C was significant at the 5% level. y*Comparisons of TRIA-C vs control was significant at the 5% level. Table l8. Dry weight of tomato seedlings from seeds treated with different pH's of TRIA-C. Dry weight (mg/shoot)z TRIA (mg/LI pH 0 0.01 0.1 1.0 (Surfactant) 5 A39 A15 A71 528** 7 528 5l6 A75 A52* 9 A35 A7A A96 A57 z The F value of the interaction for pH x TRIA-C was significant at the 5% level. Y*,**The F values for the linear trend of TRIA-C were significant at the 5 and l% levels respectively for that pH of treatment dispersion. 3l Soaking seed in hot water is recommended for disease control for many crops. It was hypothesized that increasing the treatment solution temperature might affect the seedlings' response to TRIA, resulting in both disease control and plant stimulation. TRIA-C decreased tomato plant dry weight with 200C treatment water (Table 17). Work with foliarly-applied sprays indicated that a higher pH of TRIA-A favoured the plants' response to TRIA, hence the pH of the water for seed treatment was investigated. There was a differential effect of pH and TRIA-C on the growth of tomato seedlings (Table I8). At pH 5.0 increasing TRIA increased plant dry weight (20%) at pH 7, TRIA decreased plant dry weight, while at pH 9.0 TRIA-C had no effect on plant dry weight. The largest tomato plants occurred in treatments at a pH 5.0, containing l.0 mg/L and In the surfactant control at a pH of 7.0, with 528 mg/shoot each (Table I8). There is no reasonable explanation for the increased growth of the surfactant control at pH 7.0. FIELD EXPERIMENTS. 1922, The yield of 'Augusta' winter wheat was increased in I979 by foliar applications of I,000 ug/L TRIA-A applied in I,A00 L/ha of spray (Table I9). The amount of spray and dose applied to the plants appeared to be a factor, since l,000 ug/L applied to wheat at l,A00 L/ha provided higher yields than the same concentration applied at A70 L/ha. The upright leaves of the wheat were difficult to wet with spray. TRIA-A applied to wheat at anthesis at l00 ug/L induced a slight reduction in yield. The yield of ”Goldcup' sweet corn increased linearly with dose of l00 or l,000 ug/L TRIA-A at a pH of 8.0 (Table 20). The weight of husked ears was l8 and 25% larger than the control after treatment of plants with l00 or l,000 ug/L TRIA respectively. In a second experiment with sweet corn, there was no response to TRIA-A that could be measured. 32 In I979, TRIA-A had no effect on tomatoes sprayed in the greenhouse several days prior to transplanting, or after establishment. TRIA-A at IOO ug/L increased the yield by |8% as measured in dollars/ha (based on Pickling Cucumber Improvement Committee Standard prices) of 'Greenstar' cucumbers (Table 2i). 1289, TRIA-B applied to tomatoes grown on unmulched soil and on black or silver mulches did not increase the marketable yield of tomatoes. There was an increased yield of the tomatoes planted on both kinds of mulch. Likewise in a study with different formulations of TRIA-B there was no difference in tomato yield. The addition of CaCl2 and CaCl2 + Tween 20 to the spray solution had no effect on the response of plants to TRIA. It has been reported that TRIA increased the rate of phosphate uptake in rice (II). It was postulated that TRIA applied with a starter solution might also enhance plant growth. However, research to test this hypothesis showed no significant response from plants treated with TRIA in the transplant water. Yields in this trial were low, due to inclement weather in the spring. The transplants were also damaged by cold, dry winds several days after transplanting. Plants without starter fertilizer suffered severely, while those with the fertilizer were damaged less. This may have been a factor in all three tomato experiments (as well as muskmelon), as all suffered chilling damage. An experiment to determine the effect of TRIA on soluble solids of muskmelon fruit showed no significant difference in yield or soluble solids. These results agreed with the research of Bosland (3). 33 Table I9. Increased yield of soft winter wheat treated with various volumes and concentrations of unbuffered TRIA-A. Spray applied L/ha TRIA ug/L Tonne/haz 1,A00 o 5.A2 A70 I00 5.00 A70 1,000 5.38 1,A00 100 5.23 I,AOO I,000 5.80 2*The F value for l,000 ug/L TRIA applied in l,A00 L spray/ha was significantly greater than all treatments at the 5% level. Table 20. Yield of sweet corn when sprayed to drip with TRIA-A at a pH 8.0, obtained with a l mM phosphate buffer. TRIA-A Weight of husked cobs*z (Hg/L) (tonnes/ha) 0 18.5 100 21.9 1,000 23.2 2*The F value for the linear trend to dose of TRIA is significant at the I% level. 34 Table 2i. Dollar return of cucumbers treated with TRIA-A applied in pH 8.0 l mM phosphate buffer to run-offz. TRIA-A YIeld*Y (ug/L ($lha) 0 AAA 50 A59 IOO 525 200 AOI 2 Dollar returns based on pickling cucumber improvement committee prices. Y*The F value for cubic trend to dose of TRIA is significant at the I% level. Table 22. Yield of soft winter wheat treated with TRIA-A and TRIA-B applied to run-off in two locations. Yield (tonne/ha)z Location Saranac TUscoTa I I y I I TRIA ( Frankenmuth ) ( Augusta ) (ug/L) TRIA-A TRIA-B TRIA-A 1111 A-B 0 A.A3 A.35 5.3A 5.57 l00 A.29 A.53 5.A5*x 5.32* z The F value for formulation x TRIA was significant at the 5% level in both locations. Y The F value for the comparison of TRIA-A with TRIA-B was significant at the 5% level. "*The F values for the comparison of TRIA with control were significant at the 5% level. 35 Experiments were conducted to assess the effect of supplemental nitrogen and phosphorus on the response of sweet corn and pickling cucumbers treated with TRIA-B. There was no effect of nutrients on the response of plants to TRIA. Cucumbers and corn responded to supplemental applications of fertilizer at East Lansing but not at Clarksville, perhaps because the soil fertility as measured by soil tests was higher at Clarksville than at East Lansing. Small trials with several cool season crops failed to indicate any yield increase in response to TRIA-B applied in I980. Winter wheat trials conducted in I980 with both TRIA-A and TRIA-B formulations, two cultivars, 2 times of application and 2 locations showed different results (Table 22). At Saranac there was no response with the cultivar 'Augusta'. TRIA-B increased the yield of 'Frankenmuth', whereas TRIA-A was ineffective. The opposite held for 'Augusta' at Saranac. TRIA-A and TRIA-B decreased the yield of 'Frankenmuth' winter wheat at Tuscola (5.A9 vs 5.l8 t/ha for TRIA). In East Lansing there was no significant difference among any of the treatments where TRIA was applied during the grain filling period. The addition of phosphate had no effect on the TRIA response nor did addition of CaCl to the spray emulsion. 2 1981, TRIA-C did not increase the yield of 'Snow Crown' cauliflower in l98l. The surfactant and TRIA-C at l,000 ug/L reduced the weight per plant of 'Earlivee' sweet corn (Table 23). TRIA at 0.I ug/L increased the weight of unmarketable cobs. Most of the unmarketable ears were short and not completely filled to the tip. Furthermore the percentage of harvested plants varied with different combinations of TRIA and amount of spray applied (Table 23). There was no effect of TRIA-C on ears per plant. An increase in number harvested increased with TRIA-C at 670 and l,0l0 L/ha. 36 The harvest of more plants in TRIA treatments may explain the increase in number of unmarketable ears due to the smaller cobs from smaller or later plants. During a longer season, these cobs would have likely been accep- table resulting in higher total yields from TRIA-treated plants. Yield of ripe tomato fruit was increased 3A% and 50% by 0.I and I0 ug/L of TRIA-C respectively, while there was no response at I.0 ug/L (Table 2A). There was no effect of volume of spray applied. The highest yields of ripe fruit occurred when the seedlings were treated with ID ug/L of TRIA-C. Total yield followed that of ripe fruit. The percentage of green fruit was least from plants treated with 0.I and l0.0 pg TRIA-C/L. Soil tests at the Simcoe Experiment Station indicated the presence of high levels of phosphorus and potassium,hence little response in yield from supplemental phosphorus was expected. There was also no response of sweet corn to supplemental applications of nitrogen. Phosphorus increased the marketable ears per plant and total number of ears/plant regardless of TRIA treatment. There was a differential response in number and weight of unmarketable ears to phosphorus and applications of TRIA. As with the previous corn experiment, most unmarketable ears were short and not filled to the tip. With no additional phosphorus, TRIA-C at 0, 0.1, 100 ug/L decreased the weight of unmarketable ears (7l7, 678, A53 kg/ha); when 20 kg/ha phosphorus was added TRIA-C increased the weight of unmarketable ears (AAS, A53. 750 kglha). Application of nitrogen to direct-seeded tomatoes increased the yield of ripe fruit. There was no effect of TRIA-C on either total tomato yield or number of ripe fruit. However, random samples of 25 red fruit showed differential responses of weight/fruit to TRIA and phosphorus (Table 25). One ug/L TRIA-C resulted in lower fruit weight with no phosphorus and slightly higher with added phosphorus. The effect of TRIA on size and numbers 37 Table 23a. Increase in unmarketable sweet corn in response to TRIA-C applied at the 3-leaf stage in different volumes of spray. There was no effect of spray volume nor an effect of TRIA-C on marketable yield. Treatment Marketable Unmarketable ears (husked) ears TRIA (ug/L) (g/plant) (kg/ha) Water 3I9 828 Surfactant 288*2 865 0.I 32l I305* l.0 323 733 l000 288* 923* ziiThe F value for the comparison of treatment with water control was significant at the 5% level. Table 23b. Percentage of sweet corn plants harvestedz. TRIA (ug/L)*Y L Spray Controls 0.l l.0 l000 3A0 93.A 7A.8** 78.A 85.9 670 8l.2 92.5 92.2 97.l* I0l0 86.2 82.l 9l.2 93.7 2 The F value of Liters spray applied x TRIA-C was significant at the 5% level. Y*,**The F value for the comparison of TRIA-C with controls were significant at the 5 and l% levels respectively. 38 Table 2A. Yield of tomatoes treated with various concentrations of TRIA-C. There was no difference between 3A0, 670 and lOIO L/ha of spray at the different concentrations. TRIA-C tonnes/ha Percent ug/L Ripe Total Green 0 38.1A**z 60.27* 3A.6* 0.1 50.93 67.A8 19.A 1.0 36.80 51.7A 30.1 10.0 57.3A 79.A8 21.5 zw’w * The F value for cubic trend to dose of TRIA was significant at the 5% and l% levels respectively. Table 25. The weight of 25 randomly selectgd ripe tomato fruit as affected by dose of TRIA-C and Phosphorus . Weight/fruit (g) TRIA-C (ug/L) Phosphorus (kg/ha) 0 l.0 l0.0 O 80.0 72.8 79.2 20 80.0 85.65y 76.8 2The F value for Phosphorus x TRIA-C was significant at the 5% level. Y*The comparison of TRIA l.0 ug/L at 0 with 20 kg/ha P was significant at the 5% level. 39 as the yield of ripe fruit/clump was the same. Foliar applications with a pH of 9.0 did not affect the weight of marketable corn. One ug/L of TRIA-C at 3A0 L/ha reduced the marketable ears/plant and number of marketable cobs (Table 26). Similar to the other corn experiment (Table 23), TRIA-C increased the number of un- marketable ears. TRIA-C stimulated early yield of tomatoes when applied to the roots before transplanting (Table 26). Addition of the fertilizer increased the amount of small fruits in the 'early'harvests. TRIA at l0 ug/L increased the yield of early red fruit; however, the weight/fruit was less, indicat- ing that the increase was due to number of fruit rather than size of fruit. There was a differential response to starter fertilizer and TRIA for small fruit in the early harvest. More small fruits were harvested from plants treated with TRIA-C and no starter (5.l t/ha) and fewer small fruits when the starter was added with TRIA-C (A.5 t/ha). No differences in total yield were obtained. Direct-seeded tomatoes yielded less than transplants. Transplants for the trial were from a test in which TRIA-C increased the rate of germin- ation (Table IA). The surfactant treatment and l.0 mg/L increased the amount of early small fruit as compared to the 'dry' control (Table 28). In the later yields, 0.0I mg/L decreased the number of breakers, while in early harvests TRIA-C at 0.I mg/L decreased the weight of fruit. There was no difference in total yield, indicating that TRIA may affect only early yields. It would appear that the surfactant (tallow alkyl sulphate) or soaking the seeds in water may affect growth under certain conditions. 40 Table 26. Marketable yield of sweet corn as affected by TRIA-C applied at 3A0 L/ha in phosphate buffers at pH 5.0 and 9.0. TREATMENT Marketable TRIA (pg/L) ears/plant 0.0 .968 l.0 ,ahonz l00.0 .913 z"F value for the comparison of control with L pg TRIA-C was significant at the 5% level. Table 27. Yield of tomatoes in response to TRIA-C applied to the roots for 5 minutes. Treatment Ripe fruit wt./fruit Early harvest early TRIA (Hg/L) tonnes/ha (9) 0 9.l l0l Surfactant 3.3 96 1.0 ‘9.7 9A** 10.0 10.7=‘