The Study On The Toxic Characteristics Of Badu Shengji San And The Effects Of Calamine On The Toxicity Of Mercury

Badu Shengji San (BDSJS) as one of external preparations containing mercury,which has been used for thousands of years in TCM (tradational Chinese medicine) has a unique curative effect on the treatment of chronic refractory skin ulcer. However,mercury is an extremly toxic metal and can accumulate in human bodies,which has potential danger in clinical application. It is significant to research the accumulation and alleviation of the toxicity of mercury of BDSJS as a typical external preparations containing mercury for clinical safe utilization.Our previous studies had proved that mercury was the main toxic component of BDSJS and kidney was its toxic target organ,however,compatible herbs could reduce the toxicity of mercury. Calamine which is another major component of BDSJS is made of zinc oxide and zinc can reduce blood mercury content and protect renal injury induced by mercury in recent study. Therefore, we hypothesized that calamine may be the most important mineral drug that may attenuate the toxicity of mercury. So the aim of this article was to clarify the accumulation characteristics of mercury of BDSJS and to expound the effects and mechanisms of calamine on the toxicity of mercury.To clarify the accumulation characteristics of mercury in BDSJS, damaged-skin rats were treated with BDSJS0,25,50,100mg/kg/d for4weeks. Rats were sacrificed after the last dose, recovery4weeks and recovery8weeks respectively. The consentration of mercury in blood,urine and renal cortex, coefficient of kidney and pathological changes,urinary NAG and β2-MG, biochemical indexes of hepatic and renal function were detected.Our results showed that administration of BDSJS for4weeks could cause the accumulation of mercury in blood and mainly in kidney; blood and urinary mercury recovered after suspended for4weeks and8weeks respectively; more than70%mercury were removed in renal after8weeks; BDSJS100mg/kg could damage renal tubule, but it was reversible and could recover after suspended for4weeks.We had found that BDSJS50mg/kg was a safe dose for rats.According to the result above,we calculated the dose range of human beings and evaluated the accumulation and toxicity of mercury of patients suffered from chronic skin ulcer who were treated with BDSJS for4weeks. The concentration of mercury in blood,urine,urinary NAG and β2-MG, biochemical of hepatic and renal function were detected after the last dose, recovery2weeks and recovery6weeks respectively. Our results showed that treatment of BDSJS for4weeks, blood mercury concentration revealed a rising trend, urine mercury concentration increased obviously;serum ALT,AST, BUN,Cr,urinary β2-MG had no obvious changes; the concentration of NAG in urine increased obviously; after suspended for6weeks, the concentration of mercury and NAG recovered to normal in urine.We also compared the difference of nephrotoxicity induced by BDSJS between ulcerous and damaged-skin rats which were two different skin conditon models. We took the pathological kidney coefficient, urine RBP and NAG and renal pathological changes as indexes. Our results showed that,with the same dose, renal toxicity of BDSJS on ulcer-skin rats were worse than that on the damaged-skin rats,which suggested that doctors should pay attention to the difference of absorption and accumulation of mercury according to different wounds depth.To expound the effects and mechanisms of calamine on the toxicity of mercury, several experiments in vivo and in vitro were carried out.In vivo experiments, skin-damaged rats were respectively treated with hydrargyri oxydum rubrum,which is a herb of BDSJS containg mercury and hydrargyri oxydum rubrum added calamine.After the last dose, rats were sacrificed, the concentration of mercruy in kidney, blood and urine, urinary NAG and RBP and renal MDA、T-SOD、CuZn-SOD and GSH were measured.We also observed the activities of ATPase. The result showed that after administration of hydrargyri oxydum rubrum20mg/kg/d for1week, equivalent calamine had a partial protective effect on renal toxicity and could reduce the blood mercury concentration and increase urinary excretion of mercury and decrease MDA level significantly,compared with the rats in hydrargyri oxydum rubrum group. We also observed the effect of equivalent calamine on mercury concentration of blood at different time in damaged-skin rat in single does adminstration of hydrargyri oxydum rubrum. The results showed that blood mercury concentration in rats of hydrargyri oxydum rubrum and hydrargyri oxydum rubrum added calamine groups increased gradually after the treatment,and after withdrawal drug8hous and4hours reached the hightest respectively.In order to clarify the mechanism that calamine decresed the mercury concentration in bood, plasma protein binding rates by ultrafiltration were carried out. The results showed that compared with25ng/L mercury group, protein binding rate in25ng/L Hg+Zn group had a decreased trend in plasma of rat and human plasmas; but the protein binding rate in mercury(25ng/L)+zinc group decreased obviously in human serum albumin. So, the plasma protein binding rate of mercury would decrease as a rusult of the competition of zinc.In vitro experiments, we explored the effect of zinc ion on the toxicity of mercury by cultivating the human renal proximal tubular epithelial cells (HK-2), with the changes of mitochondrial membrane potential, DNA content, nucleus area, cell membrane permeability and cell number by Cellomics High Content Imaging Analysis (HCS) method.Our results showed that, comprared with the same dose of mercury, the combination of zinc and mercury could significantly inhibit the increase of DNA content,and increase the level of mitochondrial membrane potential and cell membrane permeability.So,zinc may have no protective effect on the toxicity of mercury to HK-2cells.In conclusion, calamine can alleviate the toxicity of mercury to some extent, the mechanism may be related to decreasing the concentration of blood by competing with mercury in binding to serum albumin, accelerating the excretion of mercury in urine and zinc can also protect renal damage induced by oxidative stress of mercury by reducing the content of MDA and increasing the level of SOD in renal tissue.