Developmental toxicity, antioxidant, and marker enzyme assessment of swertiamarin in zebrafish (Danio rerio).

A secoiridoid glycoside called swertiamarin has been widely used as a herbal medicine for many decades. In particular, swertiamarin from the Enicostema axillare herb has been used as a multipurpose drug to treat innumerable health problems. As this medicine is consumed orally, its toxicity level should be determined. To examine the safety of this compound, toxicology work was done in zebrafish, and this is the first report to describe swertiamarin toxicity in zebrafish. Zebrafish embryos were used in this swertiamarin toxicity study, and morphological changes were observed. Further, the compound was also studied in adult zebrafish to determine the impact of the compound on the fish liver. Enzyme profiling with superoxide dismutase, glutathione peroxidase, catalase, reduced glutathione levels, glutathione S-transferase, lactate dehydrogenase, glutamic oxaloacetic transaminases, lipid peroxidation, Na+ /K+ -ATPase, and glutamic pyruvic transaminases) was evaluated (p ≤ 0.05). Results suggest that swertiamarin is a safe drug only at a low concentration (40 µM). This study also shows that even herbal medicinal compounds may be toxic to humans at higher dosages. Hence, irrespective of whether a drug is synthetic or natural, it needs to be tested for its toxicity before use in humans.

Subchronic effects of 2,3,7,8-TCDD or PCBs on thyroid hormone metabolism: use in risk assessment.

Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), 3,3',4,4',5-pentachlorobiphenyl (PCB 126), or 2,3,3',4,4',5-hexachlorobiphenyl (PCB 156) on thyroid hormone metabolism were studied in 13-week feeding studies in female Sprague-Dawley rats. The diets were supplemented with the compounds tested at concentrations ranging from 0.2 to 20 micrograms/kg diet for TCDD, 7 to 180 micrograms/kg diet for PCB 126, or 1.2 to 12 mg/kg diet for PCB 156, respectively. Significant correlations were found for all three compounds between reductions in plasma total thyroxine (TT4) levels and inductions of the microsomal phase II enzyme UDP-glucuronosyltransferase by using T4 as a substrate (T4UGT). Furthermore, the coinduction of certain phase I and II isozymes, i.c., cytochrome P450 1A1 (CYP1A1) and UGT1A1, by these compounds, clearly suggests the involvement of an Ah receptor-mediated mechanism in the disturbance of thyroid hormone metabolism by these polyhalogenated aromatic compounds. These results provide a mechanistic base for the use of certain effects on thyroid hormone metabolism by polyhalogenated aromatic compounds in risk assessment. By using these effects, potencies of PCB 126 and PCB 156 relative to TCDD ranged from 0.008 to 0.1 for PCB 126, and from 0.00007 to 0.004 for PCB 156, respectively. These values correspond very well with relative potencies of PCB 126 and PCB 156 by using some other well-known Ah receptor-mediated toxic and biochemical parameters.

Assessment of biological activities of mixtures of polychlorinated dibenzo-p-dioxins (PCDDs) and their constituents in human HepG2 cells.

Dose-response curves of the induction of P4501A1-dependent 7-ethoxyresorufin O-deethylase (EROD) were analyzed in human hepatoma HepG2 cells treated with defined mixtures of polychlorinated dibenzo-p-dioxins (PCDDs) and their 2,3,7,8-substituted constituents, similar to previous studies with rat hepatocytes and H4IIE cells (Schrenk et al. 1991). PCDDs appear to act less potent in human HepG2 cells in comparison with rat cells. For example, EC50 values of 2,3,7,8-Cl4DD were 8-fold and 19-fold higher than in rat H4IIE cells and hepatocytes, respectively. EC50 values of PCDDs were compared with that of 2,3,7,8-Cl4DD and expressed as 2,3,7,8-Cl4DD equivalents (TEs). Although the rank order of PCDD potencies was similar, TEs for some PCDDs (1,2,3,7,8-Cl5DD; TE = 0.75 and 1,2,3,7,8-Cl6DD; TE = 0.61) were found to be higher than in the rat system. In contrast to rat cells no significant induction of EROD could be detected with Cl8DD in HepG2 cells up to its limit of solubility. Experimentally determined TEs of PCDD mixtures containing 49 constituents were found to be largely due to additive effects of their 2,3,7,8-substituted constituents.

Assessment of biological activities of mixtures of polychlorinated dibenzo-p-dioxins: comparison between defined mixtures and their constituents.

As a first step to assess biological activities of complex mixtures of polychlorinated dibenzo-p-dioxins (PCDDs), induction of 7-ethoxyresorufin O-deethylase (EROD) by defined mixtures and their constituents has been analysed in vitro. Two cell systems have been compared: primary hepatocyte cultures and hepatoma H4IIE cells. EC50 values of PCDDs were compared with that of the most potent compound, 2,3,7,8-Cl4DD (2,3,7,8-tetrachlorodibenzo-p-dioxin) and expressed as 2,3,7,8-Cl4DD equivalents (TEs). TEs for three defined mixtures containing up to 49 PCDDs could be predicted from the sum of TEs for the 2,3,7,8-substituted congeners. Efficacies (maximal enzyme induction) of less potent PCDDs (1,2,3,4-Cl4DD, Cl8DD and of a mixture containing 86% Cl8DD and of benz(a)anthracene were lower in hepatocytes (by 33%) and in H4IIE cells (by 50%). The results suggest that biological activities of complex PCDD mixtures are largely due to additive effects of their 2,3,7,8-substituted constituents.

Quantitative assessment of enzyme induction by peroxisome proliferators and application to determination of effects on triglyceride biosynthesis in primary cultures of rat hepatocytes.

Potencies for the induction of peroxisomal fatty acyl-CoA oxidase (FACO) and microsomal laurate hydroxylase (LH) were determined for clofibric acid (CPIB), ciprofibrate (Cipro) and gemfibrozil (Gem) in primary cultures of rat hepatocytes based on complete concentration-response analysis and determination of theoretical maximum inductive responses for Cipro. CPIB and Cipro each induced FACO and LH in a concentration-dependent manner. Scatchard analysis of the data allowed calculation of EC50 values (mM) of 0.82 and 0.028 (for FACO) and 0.22 and 0.0081 (for LH) for CPIB and Cipro respectively. The EC50 ratios (CPIB/Cipro) were identical (29-fold) for induction of FACO and LH, supporting the concept that these enzymes are induced by CPIB and Cipro through a common mechanism. By comparison, Gem was relatively ineffective as an inducer of FACO and LH. Furthermore, Gem did not antagonize Cipro-mediated enzyme inductions, suggesting that Gem is a peroxisome proliferator of low potency rather than a partial agonist. Based on the potency and time-course profiles observed for induction of FACO and LH, the effects of CPIB, Cipro and Gem on triglyceride (TG) biosynthesis were determined in the cultured rat hepatocytes. Conditions of maximal FACO and LH induction by the drugs did not result in inhibition of TG biosynthesis in the cells. These results support the in vivo evidence which indicates that FACO and LH induction are not causally linked to the hypotriglyceridemic actions of peroxisome proliferating drugs.