Hematobiochemical changes of lead Poisoning and amelioration with Ocimum sanctum in wistar albino rats

An experiment was carried out to study the hematobiochemical changes of chronic lead poisoning in adult male wistar albino rats for a period of twelve weeks. Adult 216 healthy rats were randomly divided into six groups viz. control (groupI), higher dose of lead acetate @60mgs/kg bwt (group II), Lower dose of lead acetate @ 30 mgs / kg bwt (group III), Higher dose of lead + Ocimum @ 400 mgs/ kg bwt (group IV), lower dose of lead + Ocimum @ 400 mgs/ kg bwt (group V), Ocimum control (group VI). All lead treated and ameliorated groups given Lead acetate/ lead + Ocimum orally for three days in a week for a period of twelve weeks. The mean PCV, Hb, values were reduced significantly (P<0.05) in lead treated rats as dose dependent manner. Where as significant improvement was noticed in Ocimum treated groups Increased TLC and PLC values as dose dependent manner. A significant reduction in PNC was noticed in ocimum treated groups. Significant (P<0.05) decrease in serum total protein values, serum glucose and increased creatinine values were observed in lead treated groups as dose dependent . Increased protein & glucose and decreased creatinine values obtained in Ocimum treated groups. The alterations in hematological and biochemical parameters in the present study indicates decreased lifespan & fragility of RBC and damage to liver, kidney and Pancreas in lead poisoned wistar albino rats.


Introduction
agents like Thiamine, vitamin E, selenium, zinc etc.There was a meager information was available Lead is an abundant, ubiquitous, dangerous and regarding herbal products as ameliorating agent.important toxic environmental contaminant of global Keeping in view, Ocimum sanctum was used as an concern due to its significant role in modern industry ameliorating agent in present research.Ocimum and it is still being used recklessly (Valverde et al., sanctum (OS) commonly known as 'Tulasi' in Hindi is 2002).Lead occurs in a variety of organic and a medicinal plant commonly grown in India.The use inorganic compounds with a multitude of additional of this herb has been reported in Indian traditional uses in the manufacture of protective paints for iron systems of Medicine and its modern applications are and steel, explosives, rodenticide, batteries etc.The receiving wide spread attention day by day.Different manipulation of lead for these uses has caused lead parts of this plant have been claimed to be valuable in a contamination of air, dust and soil.It can be present in wide spectrum of diseases.It has been observed that air in the vicinity of factories and on the highways.tulasi has antioxidant, antibiotic, antiatherogenic, Lead levels in air, water and soil have been increasing immunomodulatory, anti-inflammatory, analgesic, during the last 10 years and it is considered to be one of antiulcer, chemopreventive and antipyretic properties the most important environmental pollutants of both (Surender Singh et al., 2007).urban and semi urban areas.Vegetables are polluted by

Material and Methods
lead from the air and a considerable amount of lead contamination is found in cereals and broad leafed Procurement of experimental animals, lead vegetables.Because of its persistence in the acetate and Ocimum sanctum (OS) environment, exposure to lead has become a major Male Wistar albino rats with body weight around 150g were utilized for the present experiment.Rats public health concern (Vaglenov et al., 2001).
were acclimatized to the experimental conditions for Several authors tried various ameliorating one week, after acclimatization, animals were grouped reported that 100, 200,300,400 mg /kg bwt of OS were administered to Wistar rats followed by intra and housed in standard poly propylene rat cages (three rats 1 cage) during the experiment.They were peritoneal injections of DMBA (7, 12, Dimethyl Benz 0 0 (a) Anthracene) (30mg /kg bwt) 90 minutes after the maintained at 25 ±1 c and a 12:12 hour interval light / dark cycle through out the experimental period for 12 final dose of the extract.At this dose of DMBA weeks(3months) by taking necessary precautions, induced the genotoxicity by formation of micronuclei.standard laboratory hygienic conditions by providing The animals pretreated with OS @ a concentration of laboratory animal feed and water adlibitum.The 300mg and 400mg /kg bwt was significantly reduced approval of the institutional animal ethical committee micronuclei formation i.e. reduced genotoxic effect of was obtained prior to commencement of the DMBA.Hence the higher dose of OS i.e. 400mg /kg experiment.
bwt was selected as dose.The Lead acetate ((CCH COO ) Pb 3H O, M.w Hematology Blood collected in 10% EDTA solution from all = 379.33)with a laboratory reagent grade was the groups at each sacrifice was used for the estimation procured from the Qualigens Fine chemicals, Bombay, India and Ocimum sanctum (OS) from the of total erythrocyte count (TEC), Total Leucocyte Natural Remedies Pvt. Ltd.Bangalore, India.

Experimental trial
Microhematocrit method (Jain 1986) and Hemoglobin Total of 216 healthy young male rats were (Hb) by sahli's method (Coles, 1986).The blood randomly assigned to the control and treatment smears were directly prepared and stained by groups.Six groups of rats consisting of 36 rats in each leishman's stain for Differential Leucocyte count group were used for the study.To know the LD value 50 (DLC) by Battlement method (Jain, 1986). of lead acetate a separate study was conducted with 36 Biochemical assay Blood was collected from all groups at each rats before starting of the experiment and according to sacrifice directly in to the sterile test tube and allowed that dose was calculated.Lead acetate and OS gavaged 0 per orally by using distilled water as vehicle.The to clot.The serum was collected and stored at 4 C i.e. groups and the doses employed for the present study in a refrigerator until use and was used for the were shown in Table -

Selection of Dose Results
The  1991).Decreased serum creatinine levels were decrease in TLC and no significant change in noticed in Ocimum ameliorated groups than lead neutrophil count in wistar rats fed with lead acetate @ treated groups as dose dependant manner.This might 1ppm, 100ppm and 1000ppm orally for 28 days.The be because of nephroprotective effect of Ocimum as increased values of TLC, PNC might be due to lead reported by Mukesh Kumar Sharma et al. (2005).inflammation in tissues (Yagminas et al. 1990).

Lymphocytopenia might be due to pathological
The results in the present study suggest that the changes that were taken place in spleen and lymphnode.
hematologically TEC,PCV,Hb and PLC values were Significant improvement in TEC, Hb, PCV, PLC significantly decreased and increased TLC & PNC and values and significant reduction in the TLC, PNC 1. estimation of Total serum proteins (Transasia Six rats from each group were randomly biochemical Ltd), glucose (M/S Excel Diagnostics sacrificed at every fortnight intervals after starting the Ltd) and creatinine (kamineni life sciences Pvt.Ltd)

Table - 1: Different groups with no. of animals and treaments.
lead acetate dose was selected based on the The mean TEC, PCV, Hb and PLC values pilot study and the OS dose was selected based on the

Table - 2: Hematological parameters (mean and SE) in animals of different experimental groups
PNC were observed in lead fed rats values in Ocimum ameliorated groups were noticed (group II & III) when compared to control.Significant (Group IV & V) when compared to lead fed groups.This might be due to hemoprotective immune stimulatory improvement was noticed in TEC, Hb, PCV, PLC and anti inflammatory effect and antioxidant property values and significant reduction in the TLC, PNC of Ocimum (Surendar singh, 2007).values in Ocimum ameliorated groups (Group IV & V) Biochemically TSP values were decreased when compared to corresponding lead fed groups.The significantly in lead treated rats as dose dependent overall mean Total erythrocyte count (TEC), packed manner and the results are in accordance with Stowe et cell volume (PCV), Hemoglobin (Hb), Total leucocyte al.(1973) and Hameed et al.(2008 Decreased levels in count (TLC), Percent neutrophil count (PNC) and TSP might be due to binding of lead to albumin (Stone Percent lymphocyte count (PLC) were given in Table-2.and Soares, 1976), disturbance in the protein Total serum proteins and glucose levels metabolism in the liver consequent to accumulation of decreased significantly (P<0.05) and significant lead leads to liver injury(Sharaf et.al.2008).Reduced increase in serum creatinine values were observed glucose levels were observed in all lead treated groups throughout the experimental period in lead treated rats (II & III) throughout the experimental period as dose (group II & III) as dose dependent manner when dependant manner.The earlier authors reported compared to control.In Ocimum ameliorated groups hypoglycemia i.e.Ashour (2002) in rabbits and (IV & V) the TSP and glucose values were improved Ashour et al. (2007) in male albino rats.In the present and decreased serum creatinine levels were noticed in study, the hypoglycemia might be due to dysfunction Ocimum ameliorated groups than lead treated groups of liver, thyroid (Goud et al.1985) and pancreatic as dose dependant manner ( Table -3).damage that was evident microscopically.A Discussion significant improvement was observed in TSP and The present study revealed that the mean TEC, glucose levels of Ocimum ameliorated groups in dose PCV, Hb and PLC values were reduced significantly dependant manner and this might be due to (P<0.05) and significant increase in TLC, PNC were hepatoprotective effect of Ocimum (Razvi Syad Ubiad observed in lead fed rats (group II & III) when et al. 2000).Significant increase in serum creatinine values in all lead treated groups in a dose dependant compared to control and suggesting that decreased manner when compared to control.Similar changes values of TEC, Hb and PCV might be due to decreased reported by Ashour et al. (2007) and Suradkar et al.Stone and Soares 1976).Similar microscopic examination of kidneys that might have observations were made by MuGahi et al. (2003) in lead to impaired glomerular infiltration (Dev et adult rats.In contrary Suradkar et al. (2009) reported al.