INTRODUCTION

Weeds are the most serious threats among the farmers. Weeds compete with the crops for soil nutrients, sun- light and space, harbor insects, pests and microorgan- isms, some weeds release poisonous substances into the soil that are harmful to plants, human beings and

ARTICLE INFORMATION:

*Corresponding Author

Received 15th May, 2015

Accepted after revision 28th June, 2015 BBRC Print ISSN: 0974-6455

Online ISSN: 2321-4007 NAAS Journal Score : 3.48

©A Society of Science and Nature Publication, 2015. All rights reserved.

Online Contents Available at: http//www.bbrc.in/

animals. That means, they reduce farm and forest pro- ductivity, displace and degrade species. Herbicides, com- monly known as weed killers, have become an impor- tant part of landscape maintenance because chemical weed control often is the more economical way than hand or mechanical weeding. Some of these act by interfering with the growth of the weed and are often

43

US Deshmukh and PM Ramteke

synthetic “imitations” of plant hormones (Kellogg et al., 2000). Herbicides are also a primary method of vegeta- tion management in industrial areas, along highways and other non-cropland areas. There are many different types of herbicides available in the market. Some are of selective type, like Pendimethalin, 2, 4-D, Pursuit etc. selectively toxic to weeds. While Paraquat, Glyphosate are non-selective herbicides (Pfeiffer, 2011).

Herbicide use has increased significantly around the world over the past six decades. By 2001 approximately

1.14billion kilograms of herbicides were applied glo- bally for the control of unwanted vegetation in agricul- tural, lawn care, aquaculture and irrigation/recreational water management activities. In general, herbicides are low to moderate in toxicity towards humans and ani- mals, because most herbicides target chemical pathways that animals do not possess (e.g., photosynthesis), how- ever, there are exceptions; many can be dermal irritants since they are often strong acids, amines, esters, and phenols. Inhalation of spray mist may cause coughing and a burning sensation in the nasal passages and chest. Extended inhalation sometimes causes dizziness. Intake will usually cause vomiting, a burning sensation in the stomach, diarrhea and muscle twitching. Herbicides rep- resent 36% of global pesticide use, followed by insecti-

cides (25%), fungicides (10%) and other chemical classes (Joshi et al., 2012).

Pesticides have been useful to fight against pests of plants, animals and humans. Hepatotoxic, genotoxic

and neurotoxic effects of different pesticides have been evaluated in many in vivo and in vitro studies. The 2,

4-Dichlorophenoxyacetc acid (2, 4-D) is an herbicide widely used to control the growth of broadleaf plants. It is chemically derived from phenoxyacetic acid. The major uses of 2, 4-D is on cereal crops such as wheat, corn, oats, rye, barley and the cane crops (U.S. Envi- ronmental Protection Agency 2002) Tuschl and Schwab, 2003, Tayyab et al., 2010 and 2015.

According to Suresh Joshi (2012) the teratogenic, neurotoxic, immunosuppressive, cytotoxic and hepa- toxic effects of 2, 4-D have been well documented. 2,

4-D herbicide increases lipid peroxidation in animal and human cells in vitro. 2, 4-D has been shown to cause

cellular mutations which can lead to cancer. This muta- gen contains dioxins, a group of chemicals known to be hazardous to human health and to the environment. Exposure to 2, 4-D induces nephrotoxicity in rats during late pregnancy and early postnatal periods. Mikov et al., (2010) reported that 2, 4-D has a hypoglycemic effect in mice.

Farmers are facing the problem of labors, weeds are reducing the produce in the fields, therefore farm- ers have been shifted towards the synthetic herbicides, they are using tremendous amount of herbicides; with

the recurrence of weeds variety of herbicides are repeat- edly used. These herbicides may not affect the crop plant but may get accumulated in it, may percolate in nearby water body through soil. Possibility of exposure and chronic accumulation of such chemicals cannot be rejected. However considering the toxic effect of her- bicide, work was carried out to study the effect of sub- chronic exposure of 2, 4-D on albino male Wister rats with reference to variation in histological architecture of liver, kidney and testis, as very limited work is carried out rats or other mammals.

MATERIAL AND METHODS

Healthy male Wister albino rats were procured from Sud- hakarrao Naik Pharmacy College, Pusad, with an average weight of 170±10 gm. Rats were acclimatized to labora-

tory condition. During acclimatized rats were provided with food and water adlibitum. The animals were kept

in clean polypropylene cages (measuring 12”x10”x8”) with chrome plates grills. The rats were grouped in to two groups, six rats in each group; one group was kept as control, while other as experimental. Experimental rats were given 25% of LD50 (12mg/kg /body weight) oral dose of the 2, 4-D dissolved in tap water for 120 days. The control rats were sacrificed on 120th day. Whereas experimental rats were sacrificed on 121st day after giving 120 days oral dose according to norms of ethical committee (1060/ac/07/CPCSEA). The tissues like liver, testis & kidney of male rats were removed, fixed in Bouin’s fixative for 24 hrs, processed by paraffin wax impregnation method, cut using a rotary microtome at 5 μm thicknesses and stained with Hematoxylin and Eosin (H X E) for light microscopic examination.

RESULTS AND DISCUSSION

Administration of oral dose of 2, 4-D to rats for 120 days has shown the observable effect on the behavior of rats. Hyperactivity was seen just after the admin- istration of dose for 5-10 minutes. After substantial exposure to oral dose, rats showed symptoms of leth- argy, red nasal and ocular discharge, severe dehydra- tion, reduced feed and water intake, and pasty diarrhea with pungent smell. Hair fall, loss of weight also been observed but no any mortality was observed.

According to EXOTOX NET (1996), 2, 4-D is a respira- tory system irritant that can cause prolonged difficulty in breathing, coughing, burning, dizziness and tempo- rary loss of muscle coordination. When experimental animals were sacrificed for tissue, it was observed that the stomach was filled with fluid having pungent smell, also it has been observed that testis were shrunk (com-

parative reduction in weight), ulceration/edema on small intestine and swelled kidneys in all most all experimen- tal rats. According to EPA RED decision, the primary target organs of the chemical 2,4-D are eyes, thyroid gland, kidney, adrenal, ovaries and testis. According to Kobal and Budihna (1999), the macroscopic examina- tion of organs at the pathoanatomical dissection of the rats treated with 2, 4-D at doses of 100 mg. kg-1, and with MCPA at doses of 150 mg.kg-1 revealed slightly inflamed and swollen mucous membranes of the stom- ach and small intestines.

HISTOLOGICAL CHANGES

Effect of 2, 4-D herbicide on Liver architecture

Liver hepatocytes of control rats were polygonal in shape, mononucleate or binucleate (Fig1a). In 2, 4-D herbicide treated rats, the cell appear degranulated. Hepatic cord becomes disarrayed. Hydropic degeneration of some hepatocytes was also observed (Fig.1b). Gorzinski et al., (1987) reported necrobiotic changes in the form of hepa- tocellular cytoplasmic swelling and homogeneity in rats

US Deshmukh and PM Ramteke

received 15 mg/kg b.w. of 2, 4-D per day. Hallenbeck and Cunninghan- Burns (1985) observed focal necrosis of the hepatocytes in rats following treatment with 2, 4-D. Makinde et al., (2015) reported fish were exposed to 0.00, 1.40, 1.44, 1.48 and 1.52mg/l of 2, 4-D amine cause advancing phase of hepatic necrosis in the liver.

Effect of 2, 4-D herbicide on Kidney architecture

Kidney of a control rats revealed normal renal tubule and glomerulus (Fig. 2a). Examination of kidney sections of 2, 4-D treated rats showed inflammatory changes in the glomeruli with increased cellularity, capillary hyper- emia, exudation, hypertrophy, tubular degeneration. The glomerular membrane was slightly thickened. Interstitial nephritis and edema was visible in the section. Exten- sive vacuolization in renal tubules was also seen (Fig. 2b). Hard et al., (1999) reported that hyperplasia of renal tubu- lar epithelium implies an increase in the number of lining cells inside the tubules i.e. without proliferation beyond the basement membrane Tayeb et al., (2015) reported sub-acute exposure to different doses of 2, 4 dichloroph- enoxyacetic acid (2, 4-D) on rat kidney. Forty animals were divided into four equal groups and treated with dif- ferent doses of 2, 4-D: 0, 15, 75 and 15mg/kg body weight per day via oral gavage for 28 consecutive days. The his- topathological study revealed tubular damages, glomeru- lar alterations, vascular congestion and increased number of pyknotic nuclei in kidneys of all 2, 4-D treated groups.

Effect of 2, 4-D herbicide on Testis architecture

The testis of 2, 4-D treated rats exhibited shrinkage of seminiferous tubules and sperms appear to be aggluti- nated. (Fig.3b). However, normal cellular architecture is seen showing normal germinal epithelial cells, Primary and secondary spermatocytes in control rats (Fig. 3a). In this context, Sever et al., (1997) found that 2, 4-D reduced sperm counts and increased abnormalities in sperm humans.

CONCLUSION

The result obtained in the present investigation reveal that the sub chronic dose of 2, 4-D herbicide i.e., 25% of LD 50 induced considerable alteration in architec- ture of the liver, kidney & testis of rats. Biochemical and haematological alterations were also observed by various workers. Effects on endocrine regulation as well as influence on reproduction need to be studied thor- oughly. It can be concluded that widespread use of such herbicides in public places and agriculture fields are to be prohibited or restricted. There is a need to aware the people about the hazardous effects of herbicides.

REFERENCES

Gorzinski S. J., Kociba R. J., Campbell R. A., Smith F. A., Nolan R. J. and Eisenbrandt D. L. (1987). Acute pharmacokinetic and subchronic studies of 2, 4-dichloropheno- xyacetic acid. J. Fund. Appl. Toxicol, 9: 423-35.

Hallenbeck W. P. and K. M. Cunninghan-Burns (1985). Pesti- cide and Human Health. Springer- Verlag, New York, NY.

Hard G. C., Alden C. L., Bruner R. H., Frith C. H., Lewis R. M., Owen R. A., Krieg K. and Durchfeld-Mayer B. (1999). Prolifera- tive lesions of the kidney and lower urinary tract in rats. J. In: Guides for Toxicologic Pathology, vol URG-1, Society of Toxicologic Pathologists, Washington DC.

Kellogg R., Nehring R., Grube A., Goss D. W. and Plotkin S. (2000). United States Department of Agriculture Natural Resources Conservation Service.

Kobal S. and Budihna M. V. (1999). Toxicity of herbicides 2, 4-D and MCPR for Rat and Rabbits. J. Acta. Brno 68:281-290.

Makinde G.E.O., Olaifa, F. E., Banjo O. (2015). Acute Toxic- ity and Histopathological Changes in Gill and Liver of Catfish (Clarias Gariepinus) Juvenile Exposed to 2, 4-D Amine (Herbex D Sl®). J. of Biology, Agriculture and Healthcare 5(4): 145-149.

Pfeiffer M. (2011). Pesticides types of herbicides. Pesticides training resources. 2525 East Seneca Tascon, Arizona 85716- 3018. 520-323-3135.

Mikov I., Vasovic V., Mikov A., Golocorbin-Kon S., Stankov K., Mikov M. (2010). Hypoglycemic Effect of Herbicide 2, 4-Dichlorophenoxyacetic Acid (2,4-D). J. Pestic Phytomed (Belgrade); 25(4): 349-352.

Pesticide information profile (1996). EXOTOX NET EXTEN- SION toxicology Network publication.

Sever L. E., Arbuckle T. E. and Sweeney A. (1997). Reproductive and developmental effects of occupational pesticide exposure: The Epidemiological evidence. Occup. Med, 12 (2): 305-25.

Joshi S. C., Tibrewal P., Sharma A. and Sharma P. (2012). Evalu- ation of toxic effect of 2, 4-D (2, 4-Dichlorophenoxyaceticacid) on fertility and biochemical parameters of male reproductive system of albino rats. J. International Journal of Pharmacy and Pharmaceutical Sciences Vol 4.

Tuschl H. and Schwab C. (2003). Cytotoxic effect of the her- bicide 2, 4-Dichlorophenoxyacetic acid in hepg2 cell. J. Food and Chemical Toxicology 41: 385-393.

Tayeb W., Nakbi A., Trabelsi M., Attia N., Miled A. and Hamma- mia M. (2010). Hepatotoxicity induced by sub-acute exposure of rats to 2, 4-Dichlorophenoxyacetic acid based herbicide désor- mone lourd, Journal of Hazardous Materials, vol. 180: 225-233.

Tayeb W., Nakbi A., Trabelsi Miled A. and Hammamia M. (2015). Biochemical and histological evaluation of kidney damage after sub-acute exposure to 2, 4-dichlorophenoxyace- tic herbicide in rats: involvement of oxidative stress. J. Toxi- cology mechanisms and methods 22(9):696-704.

US Environmental Protection Agency (2002). Toxicity and Exposure Assessment for Children’s Health, Washington: J.U.S. Environmental Protection Agency.