Introduction

Fruits are an essential component of our diet because of their nutritional value, and have a fundamental function in the diet for the protection of physical condition and preclusion of diseases.¹ The majority of Indians are vegetarians, and their average diet constitutes about 150–250 g of fruits in the total meal per day, as was observed.² As several pests attack on the fruits, they are produced under very high input pressure. For better yield and quality, pesticides are frequently applied during the entire period of growth and sometimes even at the fruiting stage.³ In a country like India, the use of pesticides has become predictable to sustain and improve the current level of crop production by protecting the crop from pests.⁴ Approximately, the consumption of pesticides around the world, about two million tons per year; among them, 45% of pesticides are used in Europe and in USA about 25% are expended and the rest are consumed in the remaining world; about 3.75% is India’s share.⁵ Now days, India has turn into the 2^nd largest producer of pesticides in Asia after China and they are ranks 12^th worldwide.⁶ The consumption of chemical pesticides, According to Ministry of Agriculture and Farmers Welfare, in the duration of 2021-2022, chemical pesticides consumption in various states was 58720 metric tons. Out of 29 Indian states, pesticide consumption in India the top four states contributing to about 62.82% are Maharashtra, Uttar Pradesh, Haryana and Telangana. The rest of pesticide 20.83% consumption in other states and 16.35% in union territories. These pesticides are absorbed by the fruits and on consumption by human beings; it may be hazardous if safe waiting period is not adopted.⁷ The actual risk to human health is during the exposure of pesticide residues in major and resultant agricultural yields.⁸ Thus, there are various problems of human health’s which are related with pesticides, ranging from short-term impacts including nausea, headaches and skin itching, to long-term impacts has been reported,⁹ including various cancers,^10-11 birth defects, infertility,¹² blood disorders,¹³ nerve disorders ¹⁴ for example, amyotrophic lateral sclerosis ¹⁵ diabetes, Alzheimer,¹⁶ Parkinson,¹⁷ chronic lymphocytic leukemia (CLL), multiple myeloma and endocrine disruption.¹⁸ These undesirable impacts of pesticides on the  environment and human health have encouraged to researchers toward the monitoring of different kinds of pesticides in various types of food commodities. It would be economically unrealistic and practically impossible.¹⁹ Therefore, the objectives of our present research work were to monitor of organochlorine pesticides present in selected fruits of the summer season, namely Plum, Kiwi fruit and Pineapple, because of their toxic nature, persistence and tendency to bioaccumulate as well as they have been banned in india but these pesticides are still in use.²⁰ Therefore, it has been decided to monitor the pesticides. Due to increasing public awareness and legal issues involved with organochlorine pesticide residues in food commodities, there is a need to harmonize the monitoring of these pesticide residues in fruits.

Materials and Methods

Reagents and Equipments

All glassware and equipments should carefully wash with deionized water and rinsed by acetone and then must dry in oven at 150^oC temperature for overnight before use. For the extraction of pesticide residues from fruits following solvents like ethyl acetate, acetonitrile, acetone, n-hexane and cyclohexane should distill before used. Then, adsorbent neutral alumina, charcoal and florisil should activate before the use. Although, to eliminate probable phthalate impurities from anhydrous sodium sulphate was purify with acetone and also heated for 3h on 600^oC in muffle furnace. By using the technique GLC equipped with capillary columns with ⁶³Ni electron capture detector purified the samples of fruits were analyzed. The small equipments like mechanical shaker, warring blender and rotatory evaporator etc. used during the study. A Stock solution of Standard was prepared in n-hexane.

Samples Collection

The sample consists of 250g of each fruit i.e. Plum (Prunus domestica), Kiwi (Actinidia deliciosa) and Pineapple (Ananas comosus), purchase from local market then deposit in refrigerator at 4ºC and analyzed within 3 days of collection. Each collected sample of fruits has been wash with water and dried up. After drying, the extraction part of fruit samples will be chopped into small pieces and macerated with 25g anhydrous sodium sulphate in warring blender to make a fine paste.

Sample Extraction

50g homogenize fine paste of each sample (Plum, Kiwi Fruit and Pineapple) and subjected to shaken separately with 100 ml acetonitrile through mechanical shaker for 3 h. Acetonitrile solvent was used for extraction previously and later this layer was discarded because it act as a polar aprotic solvent due to its low chemical reactivity, high miscibility with water . Then, extract was filtered and the remaining filtrate were transfer into the separating funnel and shaken gently for 2 h. For the first sample i.e plum (50 ml cyclohexane), for second sample i.e kiwi fruit (50 ml n-hexane) and for third sample i.e pineapple (50 ml acetone and n-hexane {1:4}) was added and again shaken for 3h. Thereafter, to obtain the two dissimilar layers, separating funnel was permitted at decent position for about 3h. The upper layer (cyclohexane from plum; n-hexane from kiwi fruit and pineapple) was separated out from separating funnel. The sample extraction was recurring three times by using 50 ml cyclohexane in plum; 50 ml n-hexane in kiwi fruit and 50 ml acetone and n-hexane (1:4) in pineapple at each step. The collected extract was resolute at 40ºC temperature upto desiccation (5 ml) by using rotatory evaporator and then, dissolved in 5 ml n-hexane.

Purification

The composed extracts of samples will be subjected for clean up by column-packed chromatography during silica gel: activated charcoal (5:1 w/w): silica gel. Then, each extract of sample be primary passed through column as well as eluted with 40 ml n-hexane. Then, eluted extract will be more resolute upto desiccation and re-dissolved in 15 ml n-hexane for analysis of pesticides on GC-ECD.

Results and Discussion

First by running the stock solution of standards (Fig.1), we have determined retention time of peaks and peak area for the pesticides present in the standard corresponding to 0.2 µg/µl concentrations (Table.1). The peaks of different pesticides in the standard exhibited their peaks at different Rt values. The dissimilar isomers of BHC (benzene hexachloride) the peaks exhibited at Rt values 7.264 for α-BHC, 10.666 for γ-BHC, 12.108 for β-BHC and 13.553 for δ-BHC. The peak was found at Rt valve 15.230 for heptachlor and 18.093 for heptachlor epoxide; At Rt value the peak was found 16.828 for aldrin; The peak was found at Rt values as: 20.356 (endosulfan I), 23.731 (endosulfan II) and 26.861 (endosulfan sulfate); At Rt value the peak was found 22.460 for dieldrin; At Rt value the peak was found 18.655 for γ-chlordane and 19.761for α-chlordane; At Rt values the peak was found which correspond as: 23.041(endrin), 24.837 (endrin aldehyde) and 27.713 (methoxychlor). At Rt values the peaks were found as: 21.560 (4,4’-DDE), 23.459 (4,4’-DDD) and 25.122 (4,4’-DDT); The peak was found at Rt value 32.942 for endrin ketone.    

Table.1: Peak area and retention time of standard of organochlorine pesticides

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