Bacterium isolated from coffee waste pulp biosorps lead: Investigation of EPS mediated mechanism.

Kleibsiella pneumoniae Kpn555, isolated from coffee waste pulp showed high level of tolerance to lead with a minimum inhibitory concentration of 900 mg/L. On its growth in nutrient broth supplemented with lead, brown clumps were visualised at the bottom of the flask. On scanning and transmission electron microscopic studies the brown clumps were corroborated to be bacterial cells with lead biosorbed on the cell surface and accumulated inside the cytoplasm. Biochemical and FT-IR analysis of the extracellular polymeric substance produced on exposure to lead revealed its chemical nature as glycolipid with protein moieties. Purified EPS (100 mg/L) could remove 50% of lead from aqueous solution (200 mg/L). Isolation of plasmid from Klebsiella pneumoniae Kpn555 revealed the presence of a plasmid of size 30-40 kb. This capability of the bacteria was proven to be plasmid mediated as the Escherichia coli DH5α cells transformed with the plasmid of Klebsiella pneumoniae Kpn555 also could tolerate 900 mg/L of lead and form brown clumps. This study shows that these bacteria, aided by EPS could serve as an effective agent for the removal of lead from contaminated water environmental samples.

Characterization of Herbal Antifungal Agent, Origanum vulgare against Oral Candida spp. Isolated from Patients with Candida-Associated Denture Stomatitis: An In vitro Study.

BACKGROUND: Candida Associated Denture Stomatitis is the prevalent fungal pathosis in denture wearers, especially in immunocompromized patients. Existing antifungal agents are ineffective since the Candida species become resistant and also, they become toxic. Origanum vulgare is a herbal plant with high anti-fungal activity against Candida of blood and urine origin. However, it has never been explored against Candida from oral cavity. MATERIALS & METHODOLOGY: Dry leaves of the plant were purchased and authenticated. Oil extraction was done using Hydro-distillation method. Clinical isolates of Candida from denture wearers was speciated using CHROMagar. Well Diffusion test was used to confirm the antifungal activity. Hydro-distillation & Maceration methods of extraction were compared. MIC/MFC was determined using CSLI guidelines. Infra-Red Spectroscopy was used to identify the active functional group. RESULTS: O.vulgare showed 30±3mm of zone of inhibition as against 19mm for fluconazole. The suitable extraction method was Hydro-distillation. MIC & MFC were found to be 0.024% and 0.097% respectively which was much lesser than for fluconazole (0.25%). The active functional group had chemically similar structure as Carvacrol, usually found in antifungal herbs. CONCLUSION: within the limitations of the study, it was concluded that (a)O.vulgare is anticandidal for clinical isolates of oral Candida, (b) Hydro-distillation is an effective method as compared to Maceration (c) MIC & MFC are much lower than that of fluconazole (d) the major functional group was structurally similar to Carvacrol.

Chronic toxicity of diethyl phthalate-A three generation lactational and gestational exposure study on male Wistar rats.

Diethyl phthalate (DEP) is widely used in the perfume industry as a vehicle for fragrances and in personal care products making human exposure of DEP significant to adults as well as neonatals, as confirmed by levels recorded in blood as well as breast milk samples of human populations in some parts of the world. Therefore, a study was undertaken to understand the toxic effect of DEP over three generations in male Wistar rats. Healthy male and female albino rats of Wistar strain weighing 75-100g (6-7 weeks old) were randomly assigned to two groups of six each. Group I (Control) male and female rats were fed on normal diet and water ad libitum. Group II (DEP) male and female rats were given DEP dissolved in corn oil mixed with the diet at 50mg/kg of the diet/day. Hundred days after the treatment, females were mated with males for 10 days. Exposure to DEP was continued throughout mating, gestation until termination at weaning, which was 150 days of total treatment period of the parental generation. The F1 and F2 generation pups were then segregated on the basis of their sex and six male and female pups of both generations were allowed to grow till they were 75-100g in weight. The treatment was then carried out similar to the parental generation but with reduced dose of 25mg/kg of the diet/day for F1 generation and 10mg/kg of the diet/day for F2 generation. Hundred days after the treatment, females were mated with males for 10 days. Exposure to DEP was continued throughout mating, gestation (21 days) until termination at weaning (21 days), which was 150 days of total treatment period of the F1 and F2 generation. Liver and serum ALT, AST and triglycerides were significantly increased over the three generations, which was much more significant in the F2 generation DEP treated group. The serum cholesterol and liver glutathione and glutathione reductase showed a significant decrease over the three generations, which was much more significant in the F2 generation DEP treated group as compared to the parental and F1 generation DEP treated rats. Histology of the liver showed remarkably enhanced fatty degeneration in the F2 generation DEP treated rats as compared to parental and F1 generation DEP treated rats. Vacuolations were much more significant in the F1 generation DEP treated rats as compared to the controls and F2 generation DEP treated rats. It can be concluded from this study, that continuous exposure through food, gestation and lactation over three generation's inspite of dose reduction of DEP leads to an enhanced toxic effect in the latter generations.

Combined and individual administration of diethyl phthalate and polychlorinated biphenyls and its toxicity in female Wistar rats.

Polychlorinated biphenyls (PCBs) are persistent environmental pollutants and known to act as xenoestrogens. PCBs and diethyl phthalate (DEP) are ubiquitous environmental pollutants because both are used as plasticizers and in various other industrial applications. Therefore, a study was undertaken to evaluate the interactive toxicity of DEP and PCBs in young female Wistar rats. Healthy young female albino rats of Wistar strain weighing 100g (7-8 weeks old) were randomly assigned to five groups of six each. Group I female rats were fed on normal diet and water ad libitum. Group II female rats were maintained on normal diet mixed with corn oil at 16.5mg/kg diet/day and 0.94mg/kg body weight/day as oil control. Groups III and IV female rats were given Clophen A60 and DEP dissolved in corn oil mixed with the diet at 50mg/(kgdietday), which is approximately equal to 2.85mg/(kgbodyweightday), individually to each group. Group V female rats received a mixture of DEP and Clophen A60, each dissolved in corn oil mixed with the diet at 50mg/(kgdietday), which is approximately equal to 2.85mg/(kgbodyweightday). Treatment was carried out for 150 days and after the completion of treatment, serum and liver enzymes and other biochemical parameters in the serum and liver were assessed. Liver weight to body weight ratio showed significant increase in Clophen A60 and Clophen A60+DEP treated rats. In the three treated groups, there was significant decrease in liver glutathione (GSH) and glutathione reductase (GR). Alanine amino transferase (ALT) was significantly increased in the liver of the three treated groups and in the serum of Clophen A60 and DEP alone treated groups and significant decrease only in the serum of Clophen A60+DEP treated rats. Significant increase in liver and serum lactate dehydrogenase (LDH) and acid phosphatase (ACP) activity was observed in the three treated groups. Alkaline phosphatase (ALP) activity was significantly increased only in the serum of the Clophen A60 and Clophen A60+DEP treated rats, whereas significant decrease in the serum and liver of DEP alone treated rats was observed. Aspartate aminotransferase (AST) activity and cholesterol levels were highly significant in the liver and serum of DEP treated rats. In addition, cholesterol level was significantly increased in the liver and serum of Clophen A60 treated rats and only in the liver of Clophen A60+DEP treated rats. Succinate dehydrogenase (SDH) activity was significantly increased in the liver of Clophen A60 and Clophen A60+DEP treated rats and highly significant increase in the serum of Clophen A60+DEP treated rats. There was significant increase in triglyceride levels in the liver and serum of Clophen A60 and Clophen A60+DEP treated rats, whereas significant increase in triglyceride levels in the serum of DEP alone treated rats was observed. Glycogen levels were significantly increased in the liver of Clophen A60+DEP treated rats, whereas serum glucose levels showed significant decrease, but in Clophen A60 alone treated rats showed significant increase in liver glycogen and serum glucose, whereas DEP alone treated rats showed significant increase in only serum glucose levels. Lipid peroxidation was increased in the liver of DEP treated rats, which was highly significant, compared to significant increase in Clophen A60 and Clophen A60+DEP treated rats. Histology of liver showed severe vacuolation, loss of hepatic architecture and granular deposits in the hepatocytes of DEP and Clophen A60+DEP treated rats, whereas in Clophen A60 alone treated rats, hepatocytes showed hyper pigmentation mild loss of hepatic architecture in centrilobular and periportal area.

Toxicity study of maternal transfer of polychlorinated biphenyls and diethyl phthalate to 21-day-old male and female weanling pups of Wistar rats.

Polychlorinated biphenyls (PCBs) are environmental pollutants known to act as xenoestrogens. PCBs and diethylphthalate (DEP) are ubiquitous environmental pollutants because both are used as plasticizers and in various other industrial applications. Therefore, a study was undertaken to evaluate the interactive toxicity of DEP and PCB in 21-day-old male and female pups of Wistar rats. Healthy young male and female albino rats of Wistar strain weighing 75-100g (6-7 weeks old) were randomly assigned to four groups of six each. Group I male and female rats were fed a normal diet and water ad libitum. Group II and III male and female rats were given PCB (Clophen A60) and DEP dissolved in corn oil mixed with the diet at 50 mg/kg of the diet (2.85 mg/kg body wt) individually to each group. Group IV male and female rats received a mixture of DEP and PCB (Clophen A60), each dissolved in corn oil mixed with the diet at 50 mg/kg of the diet (2.85 mg/kg body wt). Hundred days after the treatment, females were mated with males for 10 days. Exposure to DEP and PCB was continued throughout mating, gestation until termination at weaning, which was 150 days of total treatment period of adults. The pups from each group were then segregated on the basis of their sex. Six male and female pups each (approx. 21 days old) from each group were chosen randomly and were killed for toxicity study. Liver-to-body weight ratio showed significant increase in the male and female pups of PCB- and PCB+DEP-treated rats, whereas male pups of DEP alone treated rats showed significant increase and female pups showed significant decrease as compared to controls and other treated groups. Significant increase in liver and serum lactate dehydrogenase (LDH) and acid phosphatase (ACP) activity in the male and female pups of the three treated groups was observed. Alkaline phosphatase (ALP) activity was significantly increased only in the serum of male and female pups of the three treated groups, whereas significant decrease in the liver of male pups of the three treated groups. In the female pups, significant decrease in liver ALP was observed only PCB- and PCB+DEP-treated groups. Histology of liver showed severe vacuolation and steatosis in the hepatocytes of PCB-treated male and female pups and in PCB+DEP-treated group, vacuolation, and steatosis was much more predominant as compared to the PCB and DEP alone treated groups. DEP alone treated groups, both male and female pups showed mild vacuolations in the liver. A synergistic interactive toxic effect of PCB and DEP was evident in both male and female pups in the following study.

Chronic toxicity of diethyl phthalate in male Wistar rats--a dose-response study.

Diethyl phthalate (DEP) is widely used in personal care products, plastics and medical devices at various concentrations, but its information is limited on its toxicity associated with exposure at high as well as low doses for a prolonged period. Therefore, a study was undertaken to understand the dose-response toxic effect of DEP in male Wistar rats. Control rats were fed on normal diet and water ad libitum. Rats were given DEP dissolved individually in corn oil mixed with the diet at 10, 25 and 50 mg/kg of the diet/day, which is equal to 0.57, 1.425 and 2.85 mg/kg body wt/day. After 5 months of treatment animals were sacrificed, enzymes and other biochemical parameters in the serum and liver were assessed. Liver weight to body weight ratio showed a significant increase only in 10 ppm DEP treated rats. A significant increase was observed in the serum ACP, LDH, ALT enzyme levels of 10 mg/kg treated rats as compared to control, 25 and 50 mg/kg treated rats. Other biochemical parameters like glycogen, total cholesterol, total triglycerides and lipid peroxidation were also increased in the liver of all the three treated groups. In the 10 and 50 mg/kg diet/day treated rats, there was a significant decrease in liver total GSH as compared to controls and 25 mg/kg treated rats. Histology of liver showed severe vacuolations, fatty degeneration and loss of hepatic architecture in the 10mg/kg treated rats, whereas in the 25 and 50 mg/kg treated rats only loss of hepatic architecture and granular deposits in the hepatocytes was predominant. Histology of liver by electron micrographs showed a significant dose-dependent proliferation of mitochondria in the hepatocytes, while the 10mg/kg treated rats showed increased number of peroxisomes in the hepatocytes. It is evident from this study that treatment with higher concentrations of DEP results in mitochondrial proliferation as well as accumulation of glycogen, cholesterol and triglycerides within the liver, but exposure to lower concentrations for longer periods results in increase in peroxisome numbers leading to severe hepatocellular changes which can be confirmed by significantly increased liver weights, elevated enzyme levels in the serum and liver and impaired metabolism of glycogen, cholesterol and triglyceride as well as altered liver histology.

Toxicity study of diethyl phthalate on freshwater fish Cirrhina mrigala.

Diethyl phthalate (DEP) is used as a plasticizer, a detergent base, in aerosol sprays, as a perfume binder in incense sticks and after-shave lotions. It is known to be a contaminant of freshwater and marine ecosystems. Therefore, a study was designed to determine the toxic effects of DEP on a freshwater fish, Cirrhina mrigala. The fish was treated with 25, 50, 75, and 100 ppm (w/v) DEP dissolved in acetone to determine the LC50. Positive controls were treated with acetone only. There was 100% mortality observed within 24 h in 75 and 100 ppm, and 50% mortality in 50 ppm treated fish in 72 h. Those treated at 25 ppm showed only 10% mortality within 72 h and remaining fish continued to survive. The surviving fish were treated with 25 ppm DEP once daily for 3 days with every change of water (Group III). One group was maintained as negative control in dechlorinated water (Group I) and the other group received acetone once daily for 3 days with every change of water and was used as positive control (Group II). Fish were killed by cold narcosis on an ice block and dissected to obtain liver, muscle, and brain samples; 10% homogenates in ice-cold saline were prepared. Brain and muscle acetylcholinesterase (AchE) activity was measured. Liver aspartate (AST) and alanine aminotransferase (ALT), and liver and muscle succinate dehydrogenase (SDH) alkaline and acid phosphate (ALP and ACP) were measured. There was a significant increase in liver and muscle ACP and ALP in DEP-treated fish compared with positive and negative controls. There was a significant increase in muscle SDH and liver ALT (ALT) in DEP-treated fish compared with positive and negative controls. Brain AchE level was significantly decreased in DEP-treated fish compared to positive and negative controls. These results indicate that DEP brings about significant changes in the activity of certain liver and muscle enzymes. These alterations in enzyme activity may have long-term effects on that are continuously exposed to low doses of DEP in the aquatic environment.