Spectrophotometric investigation of Glutathione modulation by Thallium chloride in aqueous medium.

Thallium has been shown to significantly influence various tissues of living organisms; Exposure to Thallium can disturb mitochondrial function, degenerate neurons, and interfere with the function of critical metabolic enzymes and co-enzymes. Glutathione (GSH) an essential biomarker is considered a key factor in harnessing the thallium toxicity. In the present study the interaction of Thallium (Thallium Chloride) and glutathione was investigated spectro-photo-metrically in aqueous media. The renowned Elman's experimental protocol was followed at a wavelength of 412nm for Glutathione quantification in each sample. The pH of each sample was maintained at 7.6 using Phosphate buffer during the entire course of the experiment. A concentration as well as time dependent depletion of glutathione after exposure to various concentration of Thallium metal was observed, revealing chemical interaction between the metal and glutathione. The exact mechanism of interaction of Thallium and glutathione is still to be investigated. However, this piece of research suggests that a decrease in the concentration of Glutathione may be due to Thallium-GSH abduct or oxidize glutathione (GSSG) formation. This study was performed in-vitro as a model of in vivo.

Interaction of palladium inorganic salt and organic complex with glutathione content of liver homogenate.

Glutathione is an essential antioxidant of living organism that provides a primary protection against metals toxicity. A significant amount of glutathione is present in blood erythrocytes, plasma and liver hepatocytes to protect them from oxidative damage from both external and internal oxidants. Metalo-element palladium has numerous pharmacological, clinical and toxicological compensations, like palladium is used as anti-viral, anti-bacterial, neuro-protective and anti-tumor agent. However studies have also indicated some mild to serious toxic effects of palladium metallo-elements. In the presence study the interaction of palladium inorganic salt and organic complex with glutathione (GSH) content of liver homogenate was examined spectro-photometrically. 20% (w/v) liver homogenate was prepared of the collected liver of rabbit in 5% TCA (tri-chloro-acetic acid) solution and 1mm EDTA, using a potter-eveljhem homogenizer with motor driven Teflon pestle. The GSH content quantification was carried out by Elman's method. Our finding showed that there was a depletion of GSH content by both palladium inorganic salts and organic complexes, concentrations wise as well as with time elapse as level of GSH content decrease from (43.6% to 72.62%) with Palladium Nitrate and from (24.09 to 59.5%) with Bis-benzonitrile Palladium II Chloride as compared to control, and further dropped with time incubation from 0-90 minutes from (49.7 to 87.1%), with Palladium Nitrate and from (29.3% to 67.6%) respectively. The result showed that the effect of both inorganic salt of palladium was more enhanced as compare to its organic complex. It was suggested from our finding that the depletion in the glutathione content of liver homogenate may be due to oxidation of glutathione or due to glutathione metal abduct formation by both inorganic salt and organic complex of palladium. This study in situ is a model of in vivo.

Report: Palladium glutathione, N-acetylcysteine, D-penicillamine conjugation chemistry.

The metalloelement Palladium has a number of potential Pharmaco-clinical advantages. Palladium compounds have antiviral, antibacterial, neuroprotective and antitumor properties. However studies have also indicated some mild to serious toxic effects of Palladium metalloelements. Biothiols are important antioxidants that provide protection against metals toxicity. The interaction of metalloelements with biothiols can provide valuable information about the level of toxicity of the metalloelements and about the protective role of biothiols thereof. In this piece of work the effect of salt and complexes of Palladium on the status of different thiols (GSH, NAC, and D-Pen) in aqueous medium, were examined, The thiol quantification was carried out using Elman's method through UV-visible spectrophotometry and 1H- NMR. Results of the study performed in aqueous medium showed that level of different thiols depleted after the addition of the inorganic salts and organic complexes of Palladium. The mechanism of interaction of Palladium with thiols was examined using H-NMR. The results indicate that the depletion in the level of thiols may be due to 1:1 or 1:2 conjugation of Palladium with thiols. These conjugation reactions further suggest that the Palladium have xenobiotic nature causing oxidative stress and thiols play their role in detoxification and biotransformation of these metalloelements.

The role of Glutathione, Cysteine and D-Penicillamine in exchanging Palladium and Vanadium metals from albumin metal complex.

Thiol groups are extensively present across biological systems being found in range of small molecules (e.g. Glutathione, Homo-cysteine) and proteins (e.g. albumin, haemo-globin). Albumin is considered to be a major thiol containing protein present in circulating Plasma. Albumin contains a single thiolate group located at cysteine-34(cys-34) at its active site. Albumin also binds a wide variety of metals and metals complexes at various sites around the protein. Usually heavy metals are preferentially attached with the thiol group of albumin. The binding of heavy metals at cys-34 provides a mechanism by which the residence time of potentially toxic species in the body can be increased. In this research we have assessed the oxidative modification of and metal binding capacity of cys-34 with heavy metals Palladium and Vanadium to investigate the ease with which it is possible to effect disulfide-thiol exchange at this sites/or remove a metal bound at this position. Both the metals were treated with albumin and then the albumin metals (Pd and V) complexes were treated with small thoil molecules like Glutathione, Cysteine and D-Penicillamine. Our finding showed that the albumin thiol group retained the metals with itself by forming some strong bonding with the Thiols group, it is concluded from this finding that if by chance both the metals enter the living system; strongly disturb the chemistry and physiological function of this bio-molecule.

Effect of organo and inorganic lithium salt on human blood plasma glutathione- A comparative study.

Investigation of toxicological effect of various metals is the field of interest for toxicological scientists since four to five decades and especially the toxicological effect of those drugs containing metals and there use is common because there is no other choice except to use these metal containing drugs. Inorganic as well as organic salts of lithium are commonly used in prophylaxis and treatments of many psychiatric disorders. The aim of the present study was to see the difference between the effect of organic and inorganic salt of lithium commonly used in psychiatric disorders on the GSH of human blood plasma. It is the scientific fact that ionic dissociation of organic and inorganic salts of any metal is always quite different hence to prove this fact, the effect of lithium citrate (organic salt of lithium) and lithium carbonate (inorganic salt of lithium) was investigated on human blood plasma GSH to find the difference between the effect of two. Ellman's method was used for the quantification of glutathione contents in plasma. It was found that lithium citrate decrease plasma GSH contents less than lithium carbonate indicating that organic salts of lithium are safe than inorganic salts of lithium when are used in psychiatric disorders. Further to analyze the effect of organic and inorganic salt of lithium on blood plasma GSH with the increase in incubation time was also evaluated and was found that both concentration and time dependent effect of organic salt of lithium shows that this salt has decreased plasma GSH contents of human blood less than inorganic salt of lithium either by promoting oxidation of GSH into GSSG or by lithium glutathione complex formation. These results suggest the physicians that the use of organic lithium salts is much safer than inorganic salts of lithium in terms of depletion of blood plasma GSH contents.

Depletion of GSH in human blood plasma and cytosolic fraction during cadmium toxicity is temperature and pH dependent.

Toxicities of heavy metals is a burning issue and a topic of interest among the toxicologists throughout the world. Metals are always in use of man since long but in recent years the use of cadmium has increased in the form of various cadmium compounds such as cadmium compounds as stabilizers in plastic pipe industries and in the preparations of different alloys etc. Cadmium is even used in phosphate fertilizers and thus comes directly or indirectly in contact with human eatables like crops, vegetables and fruits. Once it is absorbed it affects almost all the organs and systems of human body especially blood components and kidneys. Always the chemical reactions of different chemicals are dependent on some influential factors, among these factors the effect of pH and temperature of the media in which these chemicals interact with each other are very much important. Keeping in view this fact we have evaluated the effect of cadmium nitrate tetra hydrate on GSH of human plasma and cytosolic fraction. Estimation of thiol was done by Ellman's modified method and was found that the interaction of cadmium nitrate tetra hydrate and GSH of these blood components was more at a pH and temperature, which were near to physiological pH and temperature of human body. This fact was proved as the estimated thiol concentration left after the interaction of cadmium nitrate tetra hydrate and thiol of these blood components was minimum at pH and temperature near to human blood pH and temperature. We concluded that the possible reason for depletion of GSH of these blood components was conversion of GSH into Cd(SG) (2) and/or GSSG formation.

Cadmium-glutathione complex formation in human t-cell and b-cell lymphocytes after their incubation with organo-cadmium diacetate.

Cadmium intake is associated with oxidative stress that causes depletion of intracellular as well as extra cellular reduced glutathione. There is strong evidence indicating that reactive oxygen species and reactive nitrogen species generated in the presence of cadmium could be responsible for its toxic effects in many cells and tissues. Depletion of reduced glutathione in various cells, especially in T and B-lymphocytes, causes extreme damage to the antioxidant defense system of body. The aim of this research work was to investigate the metabolic changes that occur in T and B lymphocytes after their incubation with organ cadmium diacetate by using Ellman's spectrophotometric method of thiol quantification. The results of the present study indicate that cadmium depleted T and B lymphocytes GSH to a harmful extent. It is proposed that this depletion is due to the bivalent cadmium glutathione complex formation, oxidation of reduced glutathione (GSH) to its oxidized form, or both.

Study of the effect of inorganic and organic complexes of arsenic metal on the status of GSH in T. cells and B. cells of blood.

Arsenic is a major threat to large part of the population due to its carcinogenic nature. The toxicity of Arsenic varies with its chemical form and oxidation states. Glutathione (GSH), a major intra-cellular tripeptide plays a major role in arsenic detoxification. The present study was designed to provide insight into the extent of changes in GSH level by inorganic arsenic in the form of Arsenic trioxide (ATO) and organic arsenic in the form of nitro benzene arsenic acid (NBA). Lymphocytes (T.cells and B.cells) were investigated for determination of change in GSH metabolic status caused by arsenic. The depletion of GSH level positively correlated with increasing arsenic concentration and time of incubation. The decline in GSH level was consistent with increasing pH and physiological temperature. Our findings show that changes in GSH status produced by Arsenic could be due to adduct (As-(SG)3) formation. This change in GSH metabolic status provides information regarding mechanism of toxicity of inorganic and organic arsenicals. These findings are important for the rational design of antidote for the prevention of arsenic induced toxicity.

Change in metabolic status of glutathione by palladium nitrate in blood components.

This piece of research work present the toxicological impact of varied concentrations of palladium nitrate [Pd (NO3)2] by changing the chemical status of glutathione and the way how glutathione plays its role in detoxification and conjugation processes of [Pd (NO(3))(2))] in whole blood components (plasma and cytosolic fraction). The impact of different concentration of [Pd (NO3)2] on reduced glutathione level in whole blood component (plasma and cytosolic fraction) were measured spectrophotometrically following Standard Ellman's method. Compared with control sample, significant decrease in the GSH content in whole blood components (plasma and cytosolic fraction) was obtained with various concentrations (100µM-1000µM) of palladium nitrate. Depleted GSH level was more pronounced with time incubation period (0-90) minutes. These finding shows that changes in the GSH status produced by palladium nitrate could either be due to palladium nitrate and glutathione( Pd-SG) complex formation or by conversion of reduce glutathione (2GSH + Pd(+2) - GSSG). This change in the GSH metabolic status provides information regarding the mechanism of palladium, in blood components.

Study of the effects of BaCl(2) on the level of glutathione in plasma and cytosolic fraction in whole blood.

Barium has important role in the field of medical sciences, but it has been found in various studies that barium can cause numerous toxic effects. Studies have proven the strong affinity of the metalloelements for the sulfhydryl group (SH), present in reduced glutathione (GSH) and other biological molecules. In this context, the study about the possible interaction of BaCl(2) with glutathione in whole blood components was of interest, as an indication about the extent of barium toxicity and the role of glutathione in the conjugation and detoxification of the metalloelement barium. The concentration dependent and time dependent effect of BaCl(2) on the level of GSH in plasma and cytosolic fraction in whole blood was investigated, following Ellman's method. It was found that BaCl(2) causes a decrease in the GSH level, which is more pronounced with increasing concentration of BaCl(2) and with time incubation as well. The observed effect GSH concentration may be presumably due to production of oxidized glutathione (GSSG) or then due to barium-glutathione (GS-Ba-SG) conjugate formation.

Evaluation of the interaction of vanadium with glutathione in human blood components.

Metallo-elements including Vanadium (V) have strong affinity for sulfhydryl (-SH) groups in biological molecules including Glutathione (GSH) in tissues. Because of this fact it was of interest to further investigate the interaction of Ammonium Vanadate [NH(4)VO(3)] with Glutathione as a biomarker of toxicity and the role of Glutathione in the detoxification and conjugation pr(o)Cesses in whole blood components including plasma and cytosolic fraction. Effects of different concentrations of Ammonium Vanadate [NH(4)VO(3)] on the level of reduced Glutathione in whole blood components (Plasma and Cytosolic fraction) were examined. GSH depletion in plasma and cytosolic fraction was Ammonium Vanadate's concentration-dependent. Depleted GSH level was more pronounced with more incubation time period. These findings show that changes in the GSH status produced by Ammonium Vanadate could be due to either by adduct formation of Vanadium and glutathione i.e. (V-SG) or by increased production of oxidized Glutathione (2GSH +V(+5) → GSSG). This change in GSH metabolic status provides some information regarding the mechanism of toxicity by Ammonium Vanadate and the protective role of glutathione.

Role of Glutathione in protection against mercury induced poisoning.

Mercury is harmless in an insoluble form, such as mercuric sulfide, but it is poisonous in soluble forms such as mercuric chloride or methylmercury. Mercury is a neurotoxin. Outbreaks of mercuric chloride poisonings have made it clear that adults, children, and developing fetuses are at risk from ingestion exposure to mercury. It is very important and interesting to study the reaction of mercuric chloride and Glutathione as biomarker of Glutathione role in detoxification and conjugation in components (Plasma and Cytosolic Fraction). The effect of mercuric chloride's different concentrations was examined on GSH present in plasma and cytosolic fraction. Decrease in GSH level was dependant on mercuric chloride concentration. The decrease in GSH level of blood components was more prominent with the time of incubation of mercuric chloride. Decrease in the concentration of reduced state Glutathione may be due the interaction of reduced state Glutathione (GSH) and mercuric chloride to form oxidized Glutathione (GSSG) or mercuric-glutathione complex. This change in GSH metabolic status provides information regarding the role of GSH in detoxification of mercuric chloride. The effect of mercury metal on Glutathione in blood components has been discussed in this paper in vitro condition as a model for in Vivo condition.

Comparison of high-pressure liquid chromatography and microbiological assay for determination of ciprofloxacin tablets in human plasma employed in bioequivalence and pharmacokinetics study.

Ciprofloxacin was given orally to 28 healthy male volunteers for single oral dose of 500mg; Plasma samples were collected at different time's interval between 0 and 12h and analyzed both by high pressure liquid chromatography and by a microbiological assay. The detection limits (LOD) were 0.02µg/ml and 0.1µg/ml, for both methods respectively. For each method, coefficients of variation (R(2)) were 0.9995 and 0.9918 in plasma and limit of quantitation (LOQ).02 and 0.5µg/ml. The Comparison of means maximum concentration 2.68 µg/ml at 1.5 hr for test and 2.43 µg/ml are attain in HPLC method of Reference at 2hrs respectively. The plasma concentrations measured by microbiological assay of reference tablet are 3.95µg/ml (mean ± SE) at 1 hour and 3.80µg/ml (mean ± SE) at 1 hour. The concentrations in plasma measured by microbiological method were markedly higher than the high-pressure liquid chromatography values which indicates the presence of antimicrobially active metabolites. The mean ± SE values of pharmacokinetic parameters calculated by HPLC method, for total area under the curve (AUC 0-oo) were 13.11, and 11.91 h.mg/l for both test and reference tablets respectively. The mean ± SE values of clearance measured in l/h were 44.91 and 48.42 respectively. The elimination rate constant Kel [l/h] showed 0.17 l/h for test and 0.15 l/h reference tablets and likewise, absorption half-life expressed in hours shown 0.67 h for test and 1.04 h for reference respectively. The Mean Residence Time for test is 5.48 h and 5.49 h for reference. The mean ± SE values of pharmacokinetic parameters (Microbiological assay) for total area under the curve (AUC 0-oo) were 22.11 and 19.33 h.mg/l for both test and reference tablets respectively. The mean ± SE values of clearance measured in l/h were 29.02 and 31.63 respectively. The elimination rate constant Kel [l/h] showed 0.21 l/h for test and 0.20 l/h reference tablets and likewise, absorption half-life expressed in hours shown 0.86h for test and 0.56 h for reference respectively. The Mean Residence Time for test is 5.27 h and 4.67 h for reference. Significant difference observed between two methods.

The protective role of glutathione in silver induced toxicity in blood components.

Silver nitrate (AgNO3) effect on the status of reduced glutathione (GSH), the major non-protein thiol in blood components were examined using freshly isolated plasma and cytosolic fraction of whole blood of healthy human volunteer. AgNO3 had significant effects on GSH contents reduction in plasma and cytosolic fraction of blood. Effect or reduction in GSH content was more pronounced or significant at AgNO3 concentration with time intervals (minutes to hours). Depletion of GSH in plasma and cytosolic fraction of blood by the addition of AgNO3 indicates that AgNO3 penetrated the cell membrane and reacted with GSH, depleted its contents mostly due to oxidation of reduced Glutathione (GSH) to glutathione disulphide (GSSG) 2 GSH-GSSG+2H+ or likely that AgNO3 after penetration formed a Silver-Glutathione (Ag-SG) complex. Time dependent effect of AgNO3 on the chemical status of glutathione was discussed in vitro in this paper which in principle presents the model of in vivo reaction, which will be the precise indirect impression of AgNO3 detoxification resulting by the interaction with GSH.

Bioavailability of ciprofloxacin tablets in humans and its correlation with the dissolution rates.

Bioavailabilities of seven tablets of ciprofloxacin were determined. The correlation between in-vivo bioavailability parameters and in-vitro dissolution rates were studied. Ciprofloxacin concentration from the blood was determined by microbiological assay technique. The release pattern of ciprofloxacin from tablets, which were determined by dissolution USP paddle method and spectrophotometric method was used to determine the concentration. All the parameters of ciprofloxacin bioavailability i.e., Tmax, Cmax, AUC and absorption rate constant (Ka) showed no significant correlation with dissolution rates at T30%, T50%, T90% and dissolution at 30 mins with aims as in-vivo bioequvalence waiver. The value of dissolution test used as quality tool for predicting in-vivo performance of drug product is significantly enhanced, if the in-vivo-in-vitro relationship (correlation or association) is established.

Comparative bioequivalence and pharmacokinetics of ciprofloxacin in healthy male subjects.

Pharmacokinetics and bioequivalence studies of two ciprofloxacin tablet brands (trial batch of ciprofloxacin (Test) & Reference) were conducted in 14 healthy male volunteers after oral administration. Each brand (test and reference) consisted of 500 mg of ciprofloxacin. The drug was analyzed in plasma samples with a microbiological assay using Streptococcus faecalis as test organism. The elimination half-life of 3.00 +/- 0.21 and 3.28 +/- 0.11 h was calculated for both brands. The peak plasma concentrations of (3.59 +/-0.26 ug/mL) and (3.34 +/- 1.20 ug/mL) was attained in about 1.48 +/- 0.11 hour and 1.47+/-0.05 for both Test and Reference ciprofloxacin respectively. The mean +/- SE values for total area under the curve (AUC O-alpha) were 26.15 +/- 1.35, and 24.95 +/- 0.93 hmg/1 for both test and reference tablets respectively. The mean +/- SE values of clearance were 24.83+/-1.63 and 24.73+/-1.11 1/h for both formulations respectively. The ratio of elimination rate constant Kel [l/h] was 1.14 percent difference between the test and reference tablets and likewise, half-life (t1/2beta) expressed in hours showed the ratio of 0.91 percent. This study indicated that all the pharmacokinetic and bioequivalence parameters for both ciprofloxacin formulations are statistically non-significant, hence both formulations are bioequivalent.

Assessment of bioequivalence of ciprofloxacin in healthy male subjects using HPLC.

Pharmacokinetics and bioequivalence studies of two brands of ciprofloxacin 500 mg (Ciprofloxacin & Ciproxin) were evaluated in 14 healthy male volunteers after oral administration. The drug was analyzed in plasma samples by using HPLC. The peak plasma concentrations of (2.28+/-0.04 mg/L) and (1.9+/-0.02 mg/L) was attained in about 1.71 hours and 2.00 hours for both Test and Reference ciprofloxacin, respectively. The mean +/- SE values for total area under the curve (AUC) were 11.91+/-0.26 and 13.11+/-0.27 h.mg/L for both test and reference tablets respectively. This study indicated that all the differences in bioequivalence parameters for both ciprofloxacin formulations are statistically non-significant, hence both formulations are bioequivalent.