Herbicide residues and environmental quality

At Karnal, rice and wheat were grown in Kharif and Rabi season in 2013-14 at Karnal in long-term herbicide trial. Butachlor and clodinafop were applied as rotational herbicides at 1.5 kg/ha. Sesbania crop was grown after harvest of wheat crop before transplanting rice for incorporation as green manuring.

The persistence of continuously used herbicides in rice-wheat system viz., butachlor (1.5 kg/ha) and anilophos (0.375 kg/ha) for 21 years, pretilachlor (0.75kg/ha) for 18 years to rice crop, pendimethalin (0.75 kg/ha) for 4 years and clodinafop-propargyl (0.06 kg/ha) for 13 years to wheat crop was studied. Soil (0-20 cm) and grain samples were collected at harvest for residue analysis. GC-MS/MS was used for the quantification of pretilachlor, butachlor, anilophos and clodinafop-propargyl while both HPLC and GC-MS/MS were used for the quantification of pendimethalin residues. Residues of pretilachlor, butachlor and anilophos in soil and rice grain at harvest were found below the maximum residue limits (MRL’s) (Table 5.1).

Table 5.1.1 Residue of herbicides used in rice-wheat cropping system.

Treatment dose (kg/ha)	Experimental Year	Crop	Herbicide residue at harvest (µg/g)
			Soil	Grain
Pretilachlor 0.75	18	Rice	0.007	0.006
Butachlor 1.5	21		0.003	<0.003
Anilophos 0.4	21		<0.003	<0.003
Pendimethalin 0.75	3	Wheat	<0.01	0.008
Clodinafop propargyl 0.060	13		<0.009	<0.009

Immediately after 5 hrs (0 day) of treatment, the average pendimethalin residues in the soil at 0-20 cm depth were 0.246 µg/g. The residues decreased successively as a function of time and more than 61% and 89% of the pendimethalin residues dissipated within 30 and 90 days of application, respectively. Residues declined below the detection limit (0.01 µg/g) in soil samples collected at harvest (Table 5.2).Disappearance of pendimethalin residues were according to equation: y= -0.0097x + 1.2957showing a half life of 31.30 days (Fig 5.1(b)).

Table 5.1.2 Residues of pendimethalin from wheat field soil at 0.75 kg/ha

Days after application	Residue (µg/g)	Dissipation %
0 (5 hrs)	0.246	0
3	0.168	31.7
15	0.130	47.2
30	0.095	61.4
60	0.051	79.3
90	0.027	89.0
120	0.014	94.3
Harvest	BDL	100

BDL (Below Detectable Limit) = 0.01µg/g

Fig 5.1.1: ( a) Residues (b) Dissipation kinetics of pendimethalin in soil.

Pendimethalin residue at harvest in soil and crop produce were below the limit of quantification < 0.01µg/g. Residues of pendimethalin in wheat grain were found 0.008 µg/g which were below the MRL’s. Clodinafop-propargyl residues were below the detectable limit (0.003 ug/g) in soil, grain and straw at harvest.

At AAU, Jorhat herbicide residues were evaluated at the permanent site during Kharif (Sali) and Rabi seasons. Soil samples at depths, 0 – 15 cm and 15 – 30 cm were collected after harvest of rice (18 ^th crop ) for analysis of macro and micro nutrients, bulk density ,organic carbon, carbon stock and pH The soil texture under rice – rice cropping system was sandy loam and CEC of soil is 6.18 mol(p⁺)/kg. The available N , P₂O_{5 ,}and K₂O (kg/ha) increased significantly with butachlor and pretilachlor treatments and decreases with depth over surface soil which may be due to the less weed competition and higher root uptake . Similarly, the available Zn, Cu, Mn, Fe and Ca increased significantly with butachlor and pretilachlor treated soil and higher concentration was associated with the surface s0il . Bulk density was significantly lower in surface soil (0 – 15 cm) than the subsurface (15 – 30 cm) soil due to deposition of organic matter content on surface soil for rice crop grown years together which finally reflects on the carbon stock of soil.

Table 5.1.3 Soil available organic carbon, bulk density and carbon stock after harvest of rice (18^thcrop)

Treatment	Organic C (%)		Bulk density (g/cc)		Carbon stock (mg/ha)
	0 – 15 cm	15 – 30 cm	0 – 15 cm	15 – 30 cm	0 – 15 cm	15 – 30 cm
Weedy	0.77c	0.52c	1.32a	1.62b	15.42c	18.38b
Mechanical weeding	1.01a	0.89a	1.34a	1.63b	20.46a	22.01a
Butachlor 1000 g/ha	0.96b	0.71b	1.35a	1.67a	19.55ab	17.99b
Pretilachlor 750 g/ha	0.80c	0.54c	1.49a	1.64a	18.05b	13.31c

Soil samples (0-15 cm) were collected at 0, 7, 14, 21, 30, 45 and 60 days of herbicide application to rice from respective treatments for total count for azotobcter , azospirillum and pseudomonas (Table 5.1.4. to 5.1.7).The collected soils were analyzed for microbial biomass carbon (microgram g¹soil), dehydrogenase activity (microgramTPFg^-1soil d ^-1), acid phosphatase activity (microgram pNP/g soil/h).

Table 5.1.4 Azotobacter population at various days of herbicide application (in 10⁶ cfu g soil^-1)

Treatment	0D	7D	14D	21D	30D	45D	60D
Weedy	28.67a	25.67a	26.67a	27.34a	29.67a	28.00a	28.33a
Mechanical weeding	25.00a	25.00a	26.00a	24.67a	25.00b	23.00b	25.00b
Butachlor 1000 g/ha	13.00b	8.33b	6.67c	12.00c	17.00c	16.67c	18.00c
Pretilachlor 750 g/ha	15.00b	10.33b	11.00b	15.00b	16.00c	18.00c	19.00c

Table 5.1.5 Azospirillum population at various days of herbicide application (in 10⁶ cfu g/soil)

Treatment	0D	7D	14D	21D	30D	45D	60D
Weedy	33.00a	32.67a	32.67a	35.33a	34.00a	35.00a	33.33a
Mechanical weeding	29.00b	27.33b	30.00b	29.00b	28.00b	25.33b	26.00b
Butachlor 1000 g/ha	17.00d	12.67d	9.33d	11.00d	15.00d	17.33d	19.00c
Pretilachlor 750 g/ha	21.00c	16.67c	15.33c	18.00c	19.33c	21.00c	20.33c

Table 5.1.6 PSB population at various days of herbicide application (in 10⁶ cfug/soil)

Treatment	0D	7D	14D	21D	30D	45D	60D
Weedy	23.00a	23.33a	22.00a	23.67a	20.33a	21.67a	24.00a
Mechanical weeding	21.33a	21.00b	20.33a	23.00a	21.67a	24.33a	23.00a
Butachlor 1000 g/ha	10.00c	7.00d	7.33b	9.00b	13.00b	15.00b	17.67c
Pretilachlor 750 g/ha	13.00b	11.00c	8.33b	9.67b	13.33b	17.33b	20.00b

Table 5.1.7 Deydrogenase activity at various days of herbicide application (g TPFg/ dry soil^/7days)

Treatment	0D	7D	14D	21D	30D	45D	60D
Weedy	184.16c	197.52c	199.74c	200.26c	198.18c	195.91c	196.99c
Mechanical weeding	185.56d	188.17d	182.46d	188.56d	184.00d	183.65d	185.33d
Butachlor 1000 g/ha	207.17a	220.26a	237.39a	229.72a	226.24a	214.03a	215.89a
Pretilachlor 750 g/ha	191.59b	205.22b	217.69b	222.07b	219.48b	208.75b	211.66a

Microbial population was relatively higher in soil under application of herbicides i.e. butachlor 1.5 kg/ha or pretilachlor 0.75 kg/ha. Microbial biomass carbon, dehydrogenase activity and acid phosphatase activity were relatively higher in soil under application of herbicides i.e. butachlor 1.5 kg/ha or pretilachlor 0.75 kg/ha method. Application of pretilachlor 0.75 kg/ha resulted relatively higher acid phosphatase activity and microbial biomass carbon over application of herbicides butachlor 1.5 kg/ha. (Table 5.1.3 to 5.1.7).

At CCSHAU, Hisar a long-term herbicide trial is under operation from 1999. In Rabi, clodinafop (60 g/ha) remained as permanent herbicide treatment where as sulfosulfuron was also applied as rotational herbicide this year at application rate of 25 g/ha. Butachlor applied at 1.5 Kg/ha remained as permanent herbicide treatment whereas pretilachlor was used as rotational herbicide at application rate of 1 kg/ha. Sesbania crop was grown after harvest of wheat crop and before sowing of paddy for incorporation as green manuring. Under green manuring and non green manuring, residues of clodinafop were found to dissipate 100 % after 30^th day under non-green manuring and under green manuring. Half life of clodinafop was found 6.4 days under non-green manuring; whereas, under green manuring, half life was 5.7 days. There were no residues of clodinafop in wheat grains and straw at harvest. Residues of sulfosulfuron dissipated slightly faster under green manuring than under non-green manuring conditions. The half life of sulfosulfuron was 3.9 and 5.6 days under green manuring and non-green manuring conditions, respectively.

Average initial deposit of pretilachlor (in non-green manuring) applied at 1 kg/ha was found 0.99 µg/g. On 7^th day, residues reached to 0.66 µg/g with dissipation of 33.3%. In green manuring, average initial deposits were 0.95 µg/g. Pretilachlor dissipated rapidly and about 88% of pretilachlor dissipated up to 15^th days. Half life of pretilachlor in non-green manuring was found 8.8 days, whereas in was .8 days under green-manuring (Table 5.1.8).

Table 5.1.8 Pretilachlor residues (µg/g) in non green-manuring and green-manuring conditions

Days after treatment	Pretilachlor (1.0 Kg/ha)
	Non-green manuring		Green manuring
	Average residues (µg/g)* ±SD	Dissipation (%)	Average residues (µg/g)*±SD	Dissipation (%)
0 (1h)	0.99±08	–	0.95±05	–
1	0.93±02	6.1	0.86±0.3	9.5
3	0.73±04	26.3	0.68±1.3	28.4
7	0.66±01	33.3	0.57±0.5	40
10	0.45±03	64.6	0.31±0.3	67.4
15	0.15±08	82.4	0.11±0.1	88.4
30	0.06±07	93.9	0.04±0.6	95.8
45	0.01±02	99.0	0.008±0.3	99.4
60	BDL	100	BDL	100
	Correlation Coefficient r = – 0.9831 Regression Equation y = 2.903 -0.0335 x t1/2 = 8.8 days		Correlation Coefficient r = – 0.9639 Regression Equation y = 1.074 -0.0395 x t1/2 = 7.8 days

*Average of three replicates.

Butachlor residues dissipated faster than non-green manuring conditions and dissipated to near 89 and 100 % on 30^th and 45 days under green manuring. Under non-green manuring, 75% degradation of butachlor was observed up to 30^th day. Half life of butachlor was found 13.3 and 9.2 days, under non-green manuring and green manuring, respectively. There were no residues of butachlor and pretilachlor in paddy grains and straw.

At Hyderabad a long-term field experiment is being conducted with Kharif rice and Rabi maize (zero tillage). Determination of initial representative composite soil samples before transplanting of rice and soil samples from different weed management treatments at the time of harvest revealed no significant changes in physico-chemical (pH, EC, OC) and fertility properties of the soil (Available N, P₂O₅ and K₂O) due to different treatments. Grain and straw /plant samples of rice collected at harvest showed 0.346 and 0.351 mg/kg pretilachlor residues in soil at 1 DAA in the two treatments. Pretilachlor residues persisted up to 30 DAA in the soil and reached below detectable level by 45 DAA. Residues of pretilachlor in rice grain and straw collected at harvest was below detectable level. Residues of bispyribac-sodium in the soil, rice grains and straw collected at the time of harvest were below the detectable limit of 0.02 ppm. In no-till maize soils, atrazine residues in soil were detected up to 60 DAA when applied as pre-emergence herbicide in combination with the paraquat. In post-emergence application residues could be detected up to 45 DAA. Atrazine residues were not detected in the maize grains or plant samples collected at harvest. Oxyfluorfen residues in soil were detected up to 45 DAA under zero tillage conditions. However, oxyfluorfen residues were not detected (<0.05 mg/kg) in soil, plant and grain at the time of harvest.

At TNAU, post harvest soil samples were collected in Rabi.  Soil samples were analyzed for the available nutrients (N, P, K), organic carbon pH and EC. In both the season, weed control treatments did not have significant effect on soil reaction, EC and other parameters. However the green manure application during Rabi season reduced the pH numerically during both the seasons. Soil, plant and grain samples collected during Rabi, 2014 were analyzed for butachlor, 2, 4-D and pretilachlor residue. A progressive decline in butachlor residue content was observed with advancement of crop growth. Nearly 80% pretilachlor got degraded within 30 days and it was found below detectable level at harvest. Pretilachlor degradation rate was higher under 75% inorganic N source + 25% organic source treatment. Pretilachlor residue was not found in grains and straw of 26^th crop under the treatment of alternate application of butachlor / pretilachlor. More than 90% butachlor degraded from the soil within 30 days in both the seasons. The continuous application of either butachlor + 2, 4-D or butachlor/ pretilachlor + 2, 4-D herbicide mixtures did not show build up of butachlor residue in the post harvest soil of 30^th crop.

In another experiment, nearly 80% of applied pyrazosulfuron-ethyl got degraded within 5 days of its application and it was found below detectable level at harvest. Grains and straw samples collected at the time of harvest were found to have below detectable level of pyrazosulfuron-ethyl residues. The rate of disappearance of pyrazosulfuron in soil followed first-order kinetics.Half-life of pyrazosulfuron calculated from the regression equation were found to be ranged from 2.60 to 3 days and 0.6 to 0.7 days across different treatments in soil and field water respectively.

At Bengaluru, butachlor residues were found below the limit of quantification in soil, grains and straw samples at harvest (113 days after herbicide application). At KAU, Thrissur residues in soil in the rice – rice system for 14 years (2001-2014) revealed that recommended levels of application of butachlor, pretilachlor and 2, 4-D in the rice field did not build up any residues in soil at the time of harvest. Their residues were below the detectable level in the grains and straw at harvest during the entire study period. Studies on the effect of herbicides on the population of bacteria and fungi showed that there was reduction in the population of soil microflora up to 30 days after spraying in all the herbicide applied plots. After 30 days, their population increased and tended to reach the original level by the time of harvest. This indicated that long term application of herbicides at the recommended rates under arable conditions is not likely to cause serious threat to the rice ecosystem. Residues of oxyfluorfen and pendimethalin were below the detectable level in the ginger rhizome after the harvest of the crop. While using diuron, residues were detected in fruits at higher levels of application; it is not advisable to recommend diuron at levels greater than 2.0 kg/ha. At Faizabad, butachlor residue in post harvested soil of rice under rice-wheat cropping system were determined by bioassay. Herbicide applied in rice under rice-wheat-summer green gram cropping system did not leave there toxic level of residue in soil of rice field.

Ground water samples were collected from tube-wells at Ludhiana at Moga and Kapurthala districts where farmers had applied pretilachlor, butachlor and anilophos in rice and clodinafop-propargyl and pendimethalin in wheat in rice-wheat cropping system. The residues of pretilachlor, butachlor, anilophos, clodinafop-propargyl and pendimethalin in water samples were found below detectable limit (<0.01 µg/mL). In another study, residues of paraquat and 2, 4-D in aquatic environment were evaluated in concrete ponds with one meter deep standing water at Ludhiana. Water hyacinth (Eichhornia crassipes) was grown in a tank of 2.2 sq.m. The tanks were sprayed with gramoxone (0.25 and 0.5%), and dimethyl amine salt and ethyl ester formulations of 2, 4-D (0.25 and 0.5%). It was observed that pH of water in different treatments declined up to 13^th day beyond which it increased (Table 5.2.1). The electrical conductance of water (EC) remained almost same throughout the experiment.

Table 5.2.1 Periodic pH of water samples collected at different intervals

Treatment (%)		Days after application
		0	3	6	10	13	19	24	36
Paraquat	Gramoxone 0.25	7.57	7.20	7.15	7.02	6.92	7.25	7.53	–
	Gramoxone 0.5	7.11	6.90	6.80	6.17	6.01	6.65	7.11	7.24
2’4-D	Amine salt 0.25	6.40	6.30	6.28	6.25	6.20	7.11	7.21	–
	Ester 0.25	6.89	6.55	6.25	6.19	6.12	6.40	6.99	–
	Amine salt 0.5	6.96	6.78	6.55	6.49	6.46	6.98	7.06	7.19
	Ester 0.5	6.95	6.6	6.40	6.25	6.18	6.37	7.11	–

Table 5.2.2 Persistence of herbicides (µg/mL) in water at different intervals

Treatment (%)	Days after spray
	0	3	6	10	13	19	24	36	45
Gramoxone 0.25	0.660	0.515	0.379	0.216	0.117	0.062	0.037	BDL	BDL
Gramoxone 0.5	0.959	0.914	0.669	0.570	0.452	0.280	0.108	0.035	BDL
Amine salt 0.25	0.769	0.433	0.201	0.146	0.100	0.069	0.039	BDL	BDL
Ethyl ester 0.25	0.666	0.330	0.169	0.115	0.069	0.036	BDL	BDL	BDL
Amine salt 0.5	1.002	0.640	0.330	0.161	0.115	0.097	0.069	0.025	BDL
Ethyl ester 0.5	0.925	0.537	0.268	0.144	0.095	0.060	0.034	BDL	BDL

BDL= 0.03 µg/mL

About 80% 2, 4-D and 40 to 67% paraquat dissipated within 10 days after application to water. 2,4 D dimethyl amine salt degraded to below the detectable limit (0.03 µg/mL) within 36 and 45 days at 0.25% and 0.5% applied doses. However, the dissipation of 2,4-D ethyl ester was comparatively faster and the residues were below the detection limit (0.03 µg/mL) within 24 and 36 days at dose rate of 0.25% and 0.5%, respectively. Paraquat residues were found to degrade below the detectable limit within 36 and 45 days at dose rate of 0.25% and 0.5%, respectively.

Field experiment was conducted to evaluate glyphosate residues in water at Ludhiana Wali drain, adjoining Village Sakkanwali in district Muktsar at Ludhiana. Roundup was applied at 1, 1.5 and 2% for control of sarkanda. Water samples were collected 3 hrs after the application of herbicide. For the quantification of glyphosate residues from water samples, the samples were derivatized with FMOC-Cl and analyzed by fluorescence spectrophotometer. Fluorescence spectrophotometer was found most sensitive for estimation of glyphosate. In fluorescence spectrophotometric samples were excited at 268 nm and the emission spectra was recorded (Fig 5.13b).

Fig5.2.1 (a) UV-Visible (b) Fluorescence spectra of glyphosate.

At Jorhat experiment was conducted in Rabi and Kharif by collecting water samples during January/February 2015 and August and September 2015 from water bodies at different locations adjacent to fields where herbicides were applied in winter rice crop. Butachlor residue in water declined up to 4 days slowly after which faster decline took up to 12 days for recommended levels. Butachlor residues were detectable up to 20 days of application. At HAU, Hisar water samples from the tube-wells at farmers’ fields were taken after 30-45 days of pretilachlor, butachlor, oxadiargyl, pendimethalin, metribuzin, sulfosulfuron and meso-iodosulfuron application. None out of 11 samples were having sulfosulfuron, meso+iodosulfuron, pendimethalin and metribuzin residues in ground water samples collected in Rabi 2014-15. In Kharif 2015, ten samples out of 35 were having pretilachlor residues having values ranging between 0.042 and 0.075 µg/ml. Residues of oxadiargyl and butachlor were not observed in any sample (Table 5.2.3).

Table 5.2.3 Pretilachlor, butachlor and oxadiargyl residues status in ground water at farmer’s field in paddy belt of Haryana (Kharif 2015)

Particulars	Water depth (ft)	Herbicide	Residues (µg/ml)
Ravi, Village Batta (Kaithal)	30	Pretilachlor	BDL
Sandeep, Village Batta, behind peer majaar (Kaithal)	25	Pretilachlor	BDL
Teja Patwari, Village Sain Majra (Ambala)	25	Pretilachlor	0.044
Nanak Ram, Village Chamal Majra (Ambala)	30	Butachlor	BDL
Baljeet Singh, Village Mangat Majra (Ambala)	30	Butachlor	BDL
Ravi, Village Sherpur (Y. Nagar)	25	Pretilachlor	BDL
Ramesh, Village Khanwala near Ledi (Y. Nagar)	20	Pretilachlor	0.056
Rajender village Khanwala near Ledi (Y. Nagar)	20	Pretilachlor	0.071
Near Railway crossing on road from Lapra to Kalanour (Y. Nagar)	20	Pretilachlor	BDL
Khilaram Narwal, Village Kalanour (Y. Nagar)	15	Pretilachlor	0.066
Razzaq, Village Kalanour (Y. Nagar)	20	Butachlor	BDL
Ashok Dhika, on Yamuna river bank Village Gumthala (Y. Nagar)	20	Pretilachlor	0.054
Kulvinder Village Naharpur (Y. Nagar)	20	Pretilachlor	0.042
Suresh, Village Jathlana (Y. Nagar)	15	Pretilachlor	0.075
Harpal, Village Jathlana (Y. Nagar)	20	Pretilachlor	0.053
Jaswinder, Village Labkari, Gadi Birbal (Karnal)	15	Pretilachlor	0.060
Newal Singh, Village Ghir (Karnal)	30	Butachlor	BDL
Gurchant Singh, Village Nasirpur (Ambala)	20	Butachlor	BDL
Ramchander, Village Kharak Pandwa, Kalayat (Kaithal)	25	Pretilachlor	0.048
Dalbir, Village Dhakal (Jind)	20	Pretilachlor	BDL
Devilal, Near Railway crossing Narwana (Jind)	25	Pretilachlor	BDL

At Pantnagar, water samples at 0-5, 15-20 and 30-35 days after application of herbicides in nearby fields were collected to determine persistence in water and change in water quality. 2,4-D residues in water collected from different places and location in both Rabi and Kharif season were below detectable limits of 0.009 g/ml. Study under laboratory condition did not show presence of herbicide residue (0.01 g/ml) after 30 days of application. 2,4-D persistence was less under field condition. At Hyderabad, persistence and contamination of the aquatic bodies by rice herbicides (pretilachlor and oxadiargyl) was determined by collecting water samples from the aquatic bodies in rice growing areas under Nagarjuna Sagar Project Left canal command area. In 2015, water samples were collected at two occasions from the Karimnagar district. At the time of first and second sampling drawn from the irrigation tanks, open wells or bore wells were found below the detection limit irrespective of the source. pH of the water samples was neutral to moderately alkaline.

At Combatore, water samples were collected from the cropped field bore well, pond (tail end of the area where the drained water from the rice ecosystems) during 3, 7 and 30 days after herbicide applications. Change in physico- chemical parameters influenced by different herbicides was also determine. A laboratory experiment was conducted in concrete tubs of height 90 cm with an internal diameter of 78 cm. To each pot 20 kg of soil was added to create natural environment and to study the partitioning of herbicides to the soil sediments. Then water hyacinth (Eichornia crassipes) was introduced into the tubs at 10 plants / tub and the plants were allowed for complete establishment to cover the entire tubs. After one week, 2, 4-D dimethylamine was sprayed at recommended and double the recommended dose. The two sets of herbicides treatments were followed as main treatments – W₁– With Eichornia crassipes; W₂ – Without Eichornia crassipes and Sub treatments: T₁– 2,4-D – 1.0 kg/ha; T₂ – 2,4-D – 2.0 kg/ha; T₃ – Control (no herbicide). Soil sediment and water samples were collected after spraying of herbicides at an interval of 0, 1, 3, 5, 10, 15, 20, 25 and 30 DAHA to study the residue of herbicides and the changes in pH, EC etc.

Under field conditions, water samples collected from garden land area did not contain residues of any herbicides in 1^st week of June to December 2015 months. Under submerged ecosystem, residues of herbicides analyzed were below detectable limit irrespective of source of water except for 2, 4 D and pretilachlor in the field water. In laboratory conditions, dissipation of 2,4-D from 2,4-D dimethylamine was relatively rapid and followed similar trends at both the concentrations in both water and sediments. Presence of water hyacinth reduced the quantity of 2, 4-D that reaches the water surface and also the soil sediment.

At Bengaluru, persistence of pyrazosulfuron-ethyl in water in transplanted rice was determined. Pyrazosulfuron-ethyl residues were found in the range of 0.0067 to 0.0022 mg/kg on 15^th to 30^th day. However on 45^th day it was below limit of quantification at recommended and double the recommended level of application. At KAU, persistence of glyphosate and POEA in aquatic system and their effect on fish were determined. Hgrophila polysperma bioassay was used to determine POEA in water. H. polysperma exposed to varying concentration of POEA (0.01, 0.02, 0.05, .075, 0.1, 0.5, 1.0, 2.0, 5.0, 10, 100, 1000µg/mL) and from dose response curve concentration of POEA was determined. The two sensitive parameters viz., percentage of leaves fallen and the percent leaves discoloured were compared. Half life of POEA in water was between 30 and 60 DAS in the glyphosate treatment with Salvinia. In the case of POEA without Salvinia treatment, the corresponding values were between 60 and 90 DAS. It could be seen that POEA persisted for longer period than glyphosate.

Fish samples were taken from each treatment under the experiment on ‘Persistence of glyphosate and POEA in aquatic system and their effect on fish. Visual observations on the dissected organs showed that neither glyphosate nor POEA (surfactant) caused changes in fish organs. However, the formulation Roundup® caused discoloration in the internal organs namely gills, intestine, heart, liver, ovary (with eggs). At 5 DAS, reddish tinge was observed in ovary (with eggs) and gills of the fish samples taken from the Roundup® treatment with Salvinia. However, at 45 DAS, the treatments viz., Roundup® with and without Salvinia did not show any difference from control except in the colour of gills (pale colour) which was common in all the chemical treatments.

Effect of Roundup at 5 days after application

Reddish tinge over gills and eyes Control- Normal

Soil and crop samples were collected at harvest from farmer’s fields from Ludhiana, Moga, and Kapurthala districts of Punjab in rice/wheat cropping system to determine extent of herbicides contamination in the soil and crop samples. Residue of butachlor, pretilachlor, anilophos, clodinafop-propargyl and pendimethalin were found below detectable limit (<0.01µg/g) in soil and crop produce. At Jorhat, soil samples at 0-15 cm and 15-30 cm were collected during June/July 2015 from farmers’ field growing winter rice after butachlor application. Butachlor residue at 1 kg/ha in soil, grains and straw after harvest of winter rice was found below detectable level (0.01 ppm). At HAU, Hisar, soil, wheat and rice grains and straw samples were from the sites where farmers are continuously using the pretilachlor and sulfosulfuron from many years. It was observed that 9 samples out of 20 locations were having sulfosulfuron and meso+iodosulfuron (RM) residues in soil which were ranging between 0.011 to 0.048 µg/g which was less than MRL of sulfosulfuron (0.05 µg/g) (Table 5.3.1).

Table 5.3.1 Residual of sulfosulfuron and meso+iodosulfuron (RM) in soil, wheat grain and straw at farmer’s fields in Haryana (Rabi 2014-15)

Sr. No	Name & address of farmer	Herbicide/s sprayed	Dose (g/ha)	Residues (µg/g)
				Soil	Grain	Straw
1	Virender, Village Danoda (Jind)	Leader (sulfosulfuron)	25	BDL	BDL	BDL
2	Raja, Village Samain (Tohana)	Leader (sulfosulfuron)	25	BDL	BDL	BDL
3	Dalbir Mann, Village Rasida (Jind)	Atlantis (meso+iodosulfuron RM)	14.4	0.03	BDL	BDL
4	Malkeet, Village Danoura (Ambala) (3)	Atlantis (meso+iodosulfuron RM)	14.4	0.026	BDL	BDL
5	Malkeet, Village Danoura (Ambala) (4)	Leader (sulfosulfuron)	25	0.018	BDL	BDL
6	Malkeet, Village Danoura (Ambala) (5)	Atlantis (meso+iodosulfuron RM)	14.4	0.032	BDL	BDL
7	Karam Singh, Village Rampur, (Ambala)	Leader (sulfosulfuron)	25	0.021	BDL	BDL
8	Malkeet, Village Garhi Birbal (Karnal) (1)	Leader (sulfosulfuron)	25	BDL	BDL	BDL
9	Sajjan, Village Samain (Tohana)	Leader (sulfosulfuron)	25	BDL	BDL	BDL
10	Tarsem, Village Nangla (Tohana)	Atlantis (meso+iodosulfuron RM)	14.4	0.033	BDL	BDL
11	Tarsem, Village Nangla (Tohana)	Leader (sulfosulfuron)	25	0.048	BDL	BDL

Nine out of 21 samples were having pretilachlor residues ranging between 0.004 – 0.024 µg/g in soil. Three out of 21 samples were having pretilachlor residues in rice grain ranging from 0.005 to 0.089 µg/g and 9 out of 21 samples were having pretilachlor residue between 0.014 to 0.089 µg/g in straw. Oxadiargyl residues were not observed in soil, grains and straw samples. At Pantnagar, soil and plant samples treated with 2,4-D at 0.50, clodinafop-propargyl at 0.06 kg/ha in wheat and butachlor 1.5, anilofos 0.75 kg/ha and 2,4-D at 0.50 kg/ha treated rice field were collected at harvest for its residue components. Residues of the herbicides were below MRL limits (0.01 g/g) in all components at the time of harvest of wheat and rice.

At Hyderabad, oxyfluorfen treated soil and onion samples were collected from the farmers fields. In all the soil samples collected at the time of harvest of the oxyfluorfen residues were below the detection limit. None of the onion bulb samples exhibited oxyfluorfen residues above the detection limit of 0.05 mg/kg. At Coimbatore, soil samples from the farmers fields of sattakkal pudur area was collected from four different locations during Kharif 2015 cropped with brinjal and banana treated with metribuzin and pendimethalin. Soil samples were also collected at the time of harvest from the sugarcane and maize grown fields during 2015 from Thondamuthur block. Coimbatore district that received atrazine and metribuzin. Concentrations of the studied herbicide residues in soil and crop produce were below BDL. At Thrissur Soil samples were collected from the farmer’s field at Chithali and Palakkad. Application of herbicides did not reduce the enzyme activity in the soil. All the herbicide treated plots registered higher enzyme activity than in the hand weeding treatment. Unweeded control treatment recorded higher enzyme activity than the other treatments.

At Faizabad, bioassay technique was used to determine adverse effect of herbicide on plants in soil samples taken at 0, 10, 30, 45, 60 days and at harvest. Visual phytotoxic effect on cucumber as influenced by herbicides applied in rice at different intervals was recorded (Table 50). Butachlor (1500 g /ha), and bispyribac sodium (25 g /ha) at pre-em. persisted up to 30 days in soil of rice while pretilachlor (780 g /ha) and anilofos (400 g /ha) at pre-em. persisted up to 45 days in soil of rice.

Table 5.3.2 Biometric observations regarding visual phytotoxic effect on cucumber as influenced by herbicides applied in rice in the farmers field at different intervals (in days)

Treatments	0	10	30	45	60	At harvest
Control	0	0	0	0	0	0
Butachlor 1500 g /ha pre. emergence	+++	++	+	0	0	0
Anilofos 400 g /hapre. emergence	+++	++	+	+	0	0
Pretilachlor 780 g /hapre-emergence	+++	+++	++	+	0	0
Bispyribac sodium 25 g /hapre-emergence	+++	++	+	0	0	0

Visual phytotoxic rating

No toxicity = 0; Slight toxicity = +

Medium toxicity = ++; Severe toxicity = +++

Maize crop was grown in Kharif season in 2015 at Karnal. Tembotrione was applied at 120 and 240 g/ha at 2-4 leaf stage of weeds. Residues of tembotrione metabolite (AE1417268) were determined in immature grains and mature maize grain, straw and soil collected at harvest. Limit of detection (LOD) and limit of quantification (LOQ) of tembotrione metabolite (AE1417268) was 0.003 and 0.005 ppm, respectively. Residues of tembotrione metabolite (AE1417268) were not detected in immature as well as mature maize grains, maize straw and soil samples taken at harvest. At Coimbatore determination of atrazine & its metabolites viz., hydroxy atrazine, desethyl atrazine, desethyl deisorpopyl hydroxy atrazine (DEDIPHA) by HPLC was optimized. Soil samples collected from maize field which received atrazine @ 0.5 kg / ha were subjected to atrazine and its metabolites detection. It was found that no residues were detectetd in soil at harvest.

At Ludhiana, persistence of pendimethalin and pinoxaden in soil (0-20 cm), rice grain and wheat samples at harvest under different tillage and residue management techniques was studied. Residue in soil and rice grains under different tillage and residue management techniques were found below detectable limits at the time of harvest. Pinoxaden residues ranging from 0.059 to 0.079 µg/g in the 0-20 cm soil depth 3 hr after the application and the concentration reduced to less than 0.01 µg/g in 30-45 days in all the treatments. Dissipation was comparatively faster under zero tillage treatment CT (T)-ZT-ZT, ZT (DS)-ZT+R-ZT and ZT (DS) +R-ZT+R-ZT. DT₅₀ ranged from 10.1 to 17.5 days in different treatments under recommended herbicide (W1) whereas, under integrated weed management, DT₅₀ was comparatively lower and ranged from 8.8 to 11.8 days under different treatments (Table 5.5.1).

Table 5.5.1 R², k, and DT₅₀ of pinoxaden in soil under different planting pattern

Treatment		W1			W2
		R2	k (/days)	DT50 (days)	R2	k (/days)	DT50 (days)
T1	CT(T)-CT	0.961	0.039	17.5	0.962	0.059	11.8
T2	CT(DS)-CT-ZT	0.973	0.041	17.0	0.997	0.063	11.1
T3	CT(T)-ZT-ZT	0.997	0.053	13.1	0.978	0.076	9.1
T4	ZT(DS)-ZT+R-ZT	0.993	0.054	12.7	0.992	0.078	9.0
T5	ZT(DS)+R-ZT+R-ZT	0.997	0.068	10.1	0.998	0.079	8.8

At Hisar, sampling of soil, grain and straw was done at harvest for analysis of pendimethalin. Pendimethalin residues (0.066 µg/g) in rice grains were observed in ZTDSR+(R)-ZTW+(R). In all other treatments, residues of pendimethalin were not detected in rice grains. In straw samples, pendimethalin residues varied from 0.007 to 0.059 µg/g in three DSR treatments. Pendimethalin residues (0.059 µg/g) in CTDSR-ZTW+(R) were above than MRL value of 0.05 µg/g. In soil, residues of pendimethalin were found to range from 0.040 to 0.070 µg/g in DSR treatments only. Residues of pretilachlor, bispyribac sodium and pyrazosulfuron were not detected in any of the soil, grain and straw samples.

Table 5.5.2 Pendimethalin residues (µg/g) in paddy grains, straw and soil in conservation agriculture under rice-wheat cropping system

Treatments	Pendimethalin residues* (µg/g)
	Soil	Paddy grains	Straw
ZTDSR+(R)-ZTW+(R)	0.070	0.066	0.032
ZTDSR-CTW	0.040	BDL	0.007
CTDSR-ZTW+(R)	0.045	BDL	0.059
CTR (PTR)-ZTW	BDL	BDL	BDL
CTR (PTR)-CTW	BDL	BDL	BDL

*Average of three replicates

At Pantnagar, residue of clodinafop-propargyl, metsulfuron-methyl in wheat and bispyribac -sodium in rice were below MRL (0.1 g/g) in both tillage and residue management techniques. At Hyderabad, significant changes in physico-chemical (pH, EC, OC) and fertility properties of the soil (Available N, P₂O₅ and K₂O) due to different treatments after harvest of the rice crop in different methods of establishment were not found. Pretilachlor residues persisted in the soil up to 30 DAA in the puddled rice soils where rice was established through transplanting and pretilachlor was applied at 600 g/ha dose. Pretilachlor residues in soil, rice grain and straw at harvest were below detectable level (BDL). In transplanted rice soils bispyribac- sodium persisted up to 8 days and reached to below the detection limit by 15 DAA. Residues of bispyribac-sodium in the soil samples, rice grain and rice straw samples collected at the time of harvest were below the detectable limit of 0.010 ppm in aerobic and transplanted rice treatments. Lower concentration of pendimethalin was observed in aerobic rice soils covered with residue of Daincha compared to the tilled soils. In all the treatments residues persisted up to 30 DAA. Initial concentration of atrazine in soil applied to maize as pre-emergence herbicide varied from 0.348 to 0.411 mg/kg. Atrazine and pendimethalin residues were below detection limit of 0.05 mg/kg in rice grain, plant and soil at the time of harvest.

At Gwalior, persistence of herbicides in soil at 0, 15, 30, 45, and 60 days applied to pearlmillet under pearlmillet-mustard and pearlmillet-mustard-green gram cropping system in conservation agriculture system was evaluated by cucumber and barley bioassay technique. A significant reduction in plant height and fresh weight of test plants was recorded up to 60 days after application of herbicides while dry weight was reduced up to 45 days only. The dry weight of cucumber plant was significantly reduced up to 30 and 45 DAS by isoproturon and pendimethalin respectively. No Significant reduction in growth of cucumber was recorded in soil after harvest of mustard. No significant reduction in growth of barley was recorded in soil after harvest of pearlmillet. Atrazine 750 g/ha PE + 2, 4-D 500 g/ha PoE and atrazine 750 g/ha PE + 1HW at 25 DAS applied to pearlmillet persisted in soil for 45 days. Different tillage practices in pearlmiillet-mustard-green gram cropping system could not affect the persistence of herbicides applied to pearlmillet. At TNAU, butachlor and pretilachlor residues were below detectable level (BDL, 0.005 ppm) in the post harvest soil, rice straw and grain irrespective of method of planting and type of tillage. During Kharif atrazine residues from different plots were below 0.01 mg/kg and no residues were detected in the control also.