A Study On The Bioaccessibilitv Of Cu, Zn And Pb In Soils And Dusts

To assess the health risk of heavy metals in soils/dusts quickly and reliably, in-vitro digestion test, which has progressed rapidly in the past two decades, is widely used to determinate the bioaccessibility of heavy metals in soils/dusts. The bioaccessibility of heavy metals in soils/dusts refers to the solubility of heavy metals in simulated human digestive juice. Although various in-vitro digestion tests have been developed, no universally accepted method is established. Differently designed in-vitro digestion tests have apparently differences in compositions and procedures, which has caused the lack of comparability between results of different in-vitro digestion tests and the subjectivity in selection and application of in-vitro digestion tests. The aim of in-vitro digestion test research is to establish a unified, standardized, well accepted and widely used in-vitro digestion test. Before achieve this goal, several issues must be confirmed. They are:1) the effects of intestinal digestion and digestive enzymes addition on the results of in-vitro digestion test;2) the differences and connections of in-vitro digestion test and other methods for assessing soil heavy metal pollution;3) the application meaning of in-vitro digestion test in evaluating the health risk of soil heavy metals.Three differently designed in-vitro digestion tests:SBET (Simple Bioaccessibility Extraction Test), PBET (Physiologically Based Extraction Test) and SGET (Simple Gastrointestinal Extraction Test), were chosen in this study. SBET is a method only with gastric digestion but without pepsin addition. PBET is a method including gastric and intestinal digestion with corresponding digestive enzymes. SGET is composed of gastric digestion with pepsin and intestinal digestion without bile salts and pancreatin. In this study, the differences of extraction ability between these three methods were compared, the suitable types of in-vitro digestion tests for the assessment of soil Cu, Zn and Pb was revealed, the role of digestive enzymes (pepsin, bile salts and pancreatin) in simulated digestive juices of SBET, PBET and SGET was discussed, the extraction efficiency of in-vitro digestion test (PBET) and six commonly used single-extraction methods (0.1M HNO3,0.4M HO Ac,0.1M NaNO3,0.01M CaCl2,0.05M EDTA and0.5M DTPA) was contrasted, a possibility for the integration of soil heavy metal bioaccessibility and phytoavailability was analyzed, the function of constituents in PBET (citrate, malate, acetic acid and lactic acid) was discussed, the compositions of PBET was modified and simplified, and at last the bioaccessibility of Cu, Zn and Pb in urban soils and street dusts from Hangzhou city was evaluated. Then, several conclusions were obtained:(1) The extraction ability of different in-vitro digestion tests varied significantly. SBET, which only included gastric digestion, had the highest efficiency to extract Zn and Pb from soils. In PBET and SGET, compared to the results of gastric phase, the solubility of soil Zn and Pb in the intestinal phase was obviously low. The observations indicated that the bioaccessibility of soil Zn and Pb is mainly pH-dependent. When soil entered the neutral intestinal phase, the dissolved soil Zn and Pb in acid gastric phase would be deposited as a result of pH rise. Therefore, the bioaccessibility of soil Zn and Pb in gastric digestion represented the maximum solubility of soil Zn and Pb in digestive tract. However, in PBET and SGET, when soil entered the intestinal phase, the solubility of soil Cu did not necessarily decrease or sometimes even increased due to the digestive enzymes. It was suggested that the bioaccessibility of soil Cu in gastric digestion did not represented the maximum solubility of soil Cu in digestive tract. Therefore, it is concluded that the in-vitro digestion tests only including gastric digestion, such as SBET, can be used to assess the bioaccessibility of soil Zn and Pb for the purpose of protection and precaution, but the in-vitro digestion tests comprised of gastric and intestinal digestion with digestive enzymes, such as PBET, should be selected to assess the bioaccessibility of soil Cu.(2) To uncover the function of digestive enzymes (pepsin, bile salts and pancreatin) on the release of soil heavy metals in in-vitro digestion test, three in-vitro digestion tests (SBET, PBET and SGET) were selected. The bioaccessibility of soil Cu, Zn and Pb in each method were respectively evaluated with and without digestive enzymes and the differences compared. The results showed that the bioaccessibility of soil Cu, Zn and Pb increased, decreased or hardly changed after the addition of digestive enzymes. This phenomenon can be explained by the reaction between digestive enzymes and metal ions. The combination of digestive enzymes and metal ions can help soil heavy metals stay soluble, but on the other hand, the combination may cause the precipitation of digestive enzymes and the heavy metals combined with digestive enzymes become insoluble. It can be known that the effects of digestive enzymes addition are mainly dependent on the difference between the amounts of heavy metals extracted by digestive enzymes and deposited with digestive enzymes. Besides, the role of digestive enzymes played in simulated digestive juice is different with methods. Compared to the results of gastric digestion of PBET, the bioaccessibility of soil Cu after entered the intestinal phase stayed constant or even increased because of the addition of digestive enzymes. The results of SGET when all digestive enzymes (pepsin, bile salts and pancreatin) were added reflected more differences resulting from soil and element types. And considering the effects of digestive enzymes on the bioaccessibility of soil heavy metals were mostly seen in the intestinal phase, it is concluded the addition of digestive enzymes is indispensable to the gastrointestinal digestion methods, such as PBET and SGET, but for gastric digestion methods, such as SBET, the addition of digestive enzymes is not important.(3) The extraction results for soil heavy metals by PBET and six commonly used single-extraction methods (0.1M HN03、0.4M HOAc、0.1M NaNO3、0.01M CaCl2、0.05M EDTA and0.5M DTPA) were compared. It was showed that the ability of PBET to extract Cu, Zn and Pb from soils is strong, obvious higher than neutral salt solution, and close to diluted acid solution and complex agents. The effects of soil properties on the extracted amounts of soil heavy metals by PBET were analyzed. The extractable Cu and Pb by the gastric phase and gastrointestinal phase of PBET were impacted by the similar soil properties. They decreased as the content of soil clay increased while as the total content of soil P, K, Mg, Ca and corresponding heavy metals reduced. The extracted amounts of soil Zn in the gastric and gastrointestinal phase of PBET were both significantly positively linearly correlated with the total content of soil Fe and Zn. According to the stepwise multiple linear regression analysis, the solubility of soil Cu, Zn and Pb in the gastric phase and gastrointestinal phase of PBET was both well explained by the total content of Cu, Zn and Pb in soils, respectively. The solubility of soil Cu, Zn and Pb in the gastrointestinal phase of PBET was determined to a great extent by their solubility in the gastric phase of PBET, but compared to the gastric digestion results, the gastrointestinal digestion results reflected more differences resulting from soil and element types. The coefficients of variation of bioaccessible soil Cu, Zn and Pb in the gastric phase of PBET were70.4,80.1and119.9%, respectively, but after the gastrointestinal digestion, the coefficients of variation of bioaccessible soil Cu, Zn and Pb increased to71.7,91.8and128.7%. Among six commonly used single-extraction methods, the extraction efficiency of EDTA was similar to that of PBET, and its results were well explained by the total contents of soil heavy metals, the same as PBET. Therefore, in some extent, the extraction results of PBET and EDTA represent the bioaccessibility and phytoavailability of soil Cu, Zn and Pb at the same time.(4) The effects of the compositions of simulated gastric juice of PBET (1.25g pepsin,0.50g citrate,0.50g malate,500μl acetic acid and420μl lactic acid) on the solubility of soil heavy metals were studied. Compared with the results of original PBET, the changes of soil heavy metal bioaccessibility were analyzed when the simulated gastric juices only contained1.25g pepsin,0.50g citrate,0.50g malate,500μl acetic acid or420μl lactic acid at a constant pH, respectively. The results suggested that the extraction ability of original PBET was highest and that of the simulated digestive juices including only0.50g citrate was next. The extraction procedure and mechanism of PBET was uncovered. Firstly, metal ions in solution combined with organic acids whose molecular weight was lower, such as acetic acid and lactic acid. When the organic ligands of lower molecular weight organic acids were almost combined with metal ions, the citrate acid can start to form complex with metal ions. Therefore, the citrate acid solution can mimic the extraction of soil heavy metals by organic acid (malate, acetic acid and lactic acid) in PBET. The gastric juice of PBET was simplified:the gastric juice contained1.25g pepsin and1.0g citrate. The extraction efficiency of PBET and simplified PBET was studied and found the simplified PBET can not replace the original PBET, because the extracted amounts of heavy metals in a certain soil by simplified PBET could be higher or lower than that by PBET. If the health risk of heavy metals in a certain soil was assessed based on the results of simplified PBET, then the risk could be overestimated or underestimated. Moreover, the results of original PBET reflected more differences resulting from soil types. However, the regression analysis showed that the extraction results for soil Cu, Zn and Pb by the simplified PBET had a significantly positive linear correlation with those by the original PBET. Therefore, the simplified PBET can be used to predict the bioaccessibility of soil Cu, Zn and Pb in PBET.(5) According to the Second Degree of Soil Environmental Quality implemented in China, the urban soils from Hangzhou city were not polluted by Cu, but they were contaminated by Zn and Pb in different degrees. The average concentrations of Cu, Zn and Pb in street dusts from Hangzhou city were221.67,478.70and105.31mg/kg, respectively. Compared to the contents of Cu, Zn and Pb in streets from other cities of China, the concentration of Cu in street dusts from Hangzhou city was a little high; the concentrations of Zn and Pb in street dusts were normal. The bioaccessibility of Cu, Zn and Pb in soils and dusts from Hangzhou city were evaluated using PBET and the limits of daily intake soils and dusts were calculated based on the bioaccessible heavy metals and the total content of heavy metals, respectively. The results showed:1) the sequence of heavy metals in soils and dusts from Hangzhou city was Pb>Cu>Zn;2) according to the bioaccessibility of Cu, Zn and Pb in soils, the lowest limits of daily intake soils were23.1g,84.5g and3.01g, respectively;3) according to the bioaccessibility of Cu, Zn and Pb in dusts, he lowest limits of daily intake dusts were25.54g,24.29g and3.39g. A child averagely intake0.208g soils/dusts per day. Therefore, it was known based on the extraction results of PBET, the Cu, Zn and Pb in soils and street dusts from Hangzhou city have no obvious negative effects on human health.