Soil properties influence kinetics of soil acid phosphatase in response to arsenic toxicity.

Soil phosphatase, which plays an important role in phosphorus cycling, is strongly inhibited by Arsenic (As). However, the inhibition mechanism in kinetics is not adequately investigated. In this study, we investigated the kinetic characteristics of soil acid phosphatase (ACP) in 14 soils with varied properties, and also explored how kinetic properties of soil ACP changed with different spiked As concentrations. The results showed that the Michaelis constant (Km) and maximum reaction velocity (Vmax) values of soil ACP ranged from 1.18 to 3.77mM and 0.025-0.133mMh-1 in uncontaminated soils. The kinetic parameters of soil ACP in different soils changed differently with As contamination. The Km remained unchanged and Vmax decreased with increase of As concentration in most acid and neutral soils, indicating a noncompetitive inhibition mechanism. However, in alkaline soils, the Km increased linearly and Vmax decreased with increase of As concentration, indicating a mixed inhibition mechanism that include competitive and noncompetitive. The competitive inhibition constant (Kic) and noncompetitive inhibition constant (Kiu) varied among soils and ranged from 0.38 to 3.65mM and 0.84-7.43mM respectively. The inhibitory effect of As on soil ACP was mostly affected by soil organic matter and cation exchange capacity. Those factors influenced the combination of As with enzyme, which resulted in a difference of As toxicity to soil ACP. Catalytic efficiency (Vmax/Km) of soil ACP was a sensitive kinetic parameter to assess the ecological risks of soil As contamination.

Catalytic efficiency is a better predictor of arsenic toxicity to soil alkaline phosphatase.

Arsenic (As) is an inhibitor of phosphatase, however, in the complex soil system, the substrate concentration effect and the mechanism of As inhibition of soil alkaline phosphatase (ALP) and its kinetics has not been adequately studied. In this work, we investigated soil ALP activity in response to As pollution at different substrate concentrations in various types of soils and explored the inhibition mechanism using the enzyme kinetics. The results showed that As inhibition of soil ALP activity was substrate concentration-dependent. Increasing substrate concentration decreased inhibition rate, suggesting reduced toxicity. This dependency was due to the competitive inhibition mechanism of As to soil ALP. The kinetic parameters, maximum reaction velocity (Vmax) and Michaelis constant (Km) in unpolluted soils were 0.012-0.267mMh-1 and 1.34-3.79mM respectively. The competitive inhibition constant (Kic) was 0.17-0.70mM, which was lower than Km, suggesting higher enzyme affinity for As than for substrate. The ecological doses, ED10 and ED50 (concentration of As that results in 10% and 50% inhibition on enzyme parameter) for inhibition of catalytic efficiency (Vmax/Km) were lower than those for inhibition of enzyme activity at different substrate concentrations. This suggests that the integrated kinetic parameter, catalytic efficiency is substrate concentration independent and more sensitive to As than ALP activity. Thus, catalytic efficiency was proposed as a more reliable indicator than ALP activity for risk assessment of As pollution.

Using Qmsax* to evaluate the reasonable As(V) adsorption on soils with different pH.

As a toxic metalloid element, arsenic (As) derived from human activities can pose hazardous risks to soil and water. The bioavailability of arsenic is influenced by its behavior, in particular its adsorption-desorption in the soil environment. The maximum adsorption amount (Qmax) calculated from Langmuir equation is an important parameter to estimate the adsorption capacity of adsorbents. However, the soil is a more complicated system compared with specific adsorbents. Thus, in this study, we tried to find a more reasonable parameter (Qmax*) to evaluate the adsorption capacity of soils. Eighteen Chinese soil samples with different pH were used for adsorption-desorption experiments. The maximum As(V) adsorption capacity calculated through Langmuir fitting for 18 samples were ranged from 50.25 (S13) to 312.50 (S4) mg kg-1. Besides, Qmax was highly related with soil pH. Using the difference value of adsorption amount and desorption amount to indicate the amount of non-electrostatic adsorption of As(V) onto soils, calculated the maximum adsorption amount of non-electrostatic adsorption (Qmax*). The average Qmax* of acidic and neutral soils was 162.18 mg kg-1 whereas that for alkaline soils it was only 79.52 mg kg-1. The result from multiple linear regression analysis showed Qmax* was strongly influenced by Feox and clay contents. Furthermore, hysteresis index (HI) in the As(V) desorption varied from 0.83 (S13) to 1.82 (S6). The results further indicated the risk of secondary pollution originating from the desorption process cannot be ignored.

How different are the arsenic fractions inhibit alkaline phosphatases on aggregates scale?

Arsenate [As(V)], in general, is associated with various aggregates and exists as different species in soil, which in turn influences its toxicity and potential contamination. Previous studies have demonstrated the usefulness of alkaline phosphatases (ALP) to evaluate As(V) pollution. However, the effect of different arsenic fractions on ALP among soil aggregates is still unclear. Thus, the distribution of As fractions and ALP kinetics was determined in four-month As-aged paddy soil aggregates. Results revealed the two major fractions of As in aggregates were humic-bound and Fe and Mn oxides-bound [both around 30% under 800 mg kg-1 of As(V)]. Besides, it was observed that available soil phosphorus could positively affect the relative content of water-soluble, exchangeable and carbonate-bound arsenic. In the kinetics experiment, both the Michaelis-Menten constant (Km) and maximum reaction velocity (Vmax) of ALP increased with increasing As(V) concentration under four months ageing for each size aggregate. Multiple linear stepwise regression analysis between kcat and the relative content of arsenic fraction indicated that carbonate-bound arsenic is the main fraction that inhibited the kcat for macroaggregates (> 0.25 mm size). For soil aggregates of 0.1-0.25 mm size, kcat increased with an increase in arsenic residual fraction. As for aggregates <0.1 mm size, Fe and Mn oxide-bound fraction is the main fraction that inhibited the kcat. Overall, this study suggests carbonate-bound and Fe and Mn oxide-bound arsenic fractions could decrease the ALP activities via a decrease in the catalytic efficiency in macroaggregates and <0.1 mm size aggregates, respectively. Besides, available phosphorus should be considered as the main factor when assessing As biotoxicity and mobility.

Tinospora cordifolia and arabinogalactan exert chemopreventive action during benzo(a)pyrene-induced pulmonary carcinogenesis: studies on ultrastructural, molecular, and biochemical alterations.

The aim of the present study was to unveil the chemopreventive potentials of aqueous Tinospora cordifolia stem extract and its active component viz. Arabinogalactan against Benzo(a)pyrene-induced pulmonary carcinogenesis. Animals were divided into six groups: (I) Control, (II) aqueous Tinospora cordifolia (200 mg/kg b.wt, p.o.), (III) arabinogalactan (7.5 mg/kg b.wt, p.o.), (IV) benzo(a)pyrene (50 mg/kg b.wt, i.p.) at second and fourth week of study, (V) benzo(a)pyrene + aqueous Tinospora cordifolia, and (VI) benzo(a)pyrene + arabinogalactan. The benzo(a)pyrene treatment resulted in severe alterations in the cellular arrangement and morphology of the alveolar tissue in benzo(a)pyrene group. However, benzo(a)pyrene + aqueous Tinospora cordifolia and benzo(a)pyrene + arabinogalactan groups revealed classical features of apoptosis including chromatin condensation and formation of apoptotic bodies. Furthermore, Fourier transform Infrared spectroscopy analysis showed disturbed phospholipid saturation and protein secondary structures in benzo(a)pyrene treated animals. Depletion in relative glycogen and enhancement in total nucleic acid content was observed in benzo(a)pyrene treated animals, and the same was found to be restored upon arabinogalactan and aqueous Tinospora cordifolia supplementation. Benzo(a)pyrene insult also upregulated the phase I carcinogen metabolizing enzymes and differentially modulated the phase II metabolizing enzymes during pulmonary carcinogenesis. Also, depleted (reduced glutathione) and increased lipid peroxidation levels were observed in benzo(a)pyrene treated animals, which was found to be normalized upon aqueous Tinospora cordifolia and arabinogalactan administration. Clastogenic damage inflicted by benzo(a)pyrene was also reversed in benzo(a)pyrene + aqueous Tinospora cordifolia and benzo(a)pyrene + arabinogalactan group. Thus, the present study infers that aqueous Tinospora cordifolia and arabinogalactan showed promising anticancer activity against lung tumorigenesis in terms of ultrastructural, biochemical, and biomolecular aspects.

The effect of arsenic on soil intracellular and potential extracellular ß-glucosidase differentiated by chloroform fumigation.

Arsenic (As) contamination of soil is a global issue of serious ecological and human health concern. For better use of soil enzymes as biological indicators of As pollution, the response of soil ß-glucosidase in different pools of soil (total, intracellular and potential extracellular) to As(V) stress was investigated. Chloroform fumigation method was employed to distinguish the intracellular and potential extracellular ß-glucosidase in three soils. The intracellular and potential extracellular ß-glucosidase accounted about 79% and 21% of the total ß-glucosidase activity in the tested soils. Moreover, it was found that the response of these three enzyme pools to As(V) pollution was different. Under the stress of 400 mg kg-1 As(V), the ß-glucosidase activities decreased by 69%, 79%, and 28% for the total, intracellular and potential extracellular pools, respectively. The calculated median ecological dose (ED50) showed the highest value for potential extracellular ß-glucosidase (19.55-27.63 mg kg-1 for total, 18.49-27.42 mg kg-1 for intracellular, and 32.27-52.69 mg kg-1 for potential extracellular ß-glucosidase). As(V) exhibited an uncompetitive inhibition for total and intracellular ß-glucosidase and non-competitive inhibition for potential extracellular enzyme. The inhibition constant (Kiu) is biggest for potential extracellular ß-glucosidase among the three enzyme pools (0.61-0.79 mmol L-1 for total, 0.34-0.36 mmol L-1 for intracellular, and 4.01-23.90 mmol L-1 for potential extracellular ß-glucosidase). Thus, compared to potential extracellular ß-glucosidase, the total and intracellular ß-glucosidases are more suitable for their use as sensitive indicators of As(V) pollution.

The distribution of arsenic fractions and alkaline phosphatase activities in different soil aggregates following four months As(V) ageing.

Soil as a heterogeneous mass is composed of different size aggregates. The distribution of different arsenic (As) fractions in soil aggregates is vital to assess the potential risk of As pollution. In this study, soil samples were aged for 4 months with different arsenate [As(V)] concentrations. Dry sieving method was used to obtain five different size aggregates and the content of As in these fractions was determined. The results showed that P4 (0.1-0.25 mm) contained the highest organic matter (OM) than other size aggregates. After 4 months of ageing, available phosphorus (AP) content increased with the increase of As(V) concentration among 5 aggregates. The distribution of different arsenic fractions among 5 aggregates was similar. The relative contents of water-soluble (F1), exchangeable (F2) and carbonate (F3) fractions increased with the increase in As concentration, while the residual fraction (F7) decreased sharply. Humic-bound (F4), and Fe and Mn oxide bound fractions (F5) were about 35% and 20% respectively, after 4 months of As(V) ageing. Generally, the alkaline phosphatase (ALP) activities of P4 were lowest among five aggregates under each concentration of As(V). Moreover, F2 and F3 exhibited a strong inhibition of ALP activity. This study demonstrates that not only water-soluble and exchangeable arsenic but also humic-bound fraction should be considered when assessing As bioavailability and toxicity.

Differences in the response of soil dehydrogenase activity to Cd contamination are determined by the different substrates used for its determination.

Dehydrogenase activity (DHA) is an important indicator of heavy metal toxicity in contaminated soils. Different instances of DHA were determined using various substrates and which could affect the description of heavy metal toxicity. Currently, too few investigations have been done on selecting appropriate substrates. This study employed indoor simulation to determine soil DHA and its response to external cadmium (Cd) using two substrates (TTC and INT). Hormesis for DHA obtained using the TTC method (DHA-TTC) in low Cd concentration was observed which was quickly inhibited in high Cd concentration. While DHA obtained using the INT method (DHA-INT) decreased slowly when Cd concentration increased. The DHA-TTC and DHA-INT in soils at Cd concentration of 500 mg kg-1 decreased 86% and 53%, respectively, compared to the control. The dose-response relationship of Cd to DHA can be well simulated using the logistic model (p < 0.01), which indicated DHA could be used to indicate soil Cd toxicity. Multiple stepwise regression analysis revealed that total organic matter (TOC) is the major factor influencing the toxicity of Cd to DHA-TTC, while TOC, pH and cation exchange capacity (CEC) are major factors influencing the toxicity of Cd to DHA-INT. The different responses of soil DHA-TTC and DHA-INT to Cd are due to the differences in electron transport chain characteristics between TTC and INT, as well as the influence of soil properties. Although both DHA-TTC and DHA-INT can monitor soil Cd contamination, DHA-INT is recommended as a superior bio-indicator to indicate and assess contamination of Cd in soil.

Kinetics of soil dehydrogenase in response to exogenous Cd toxicity.

Soil dehydrogenase plays a role in the biological oxidation of soil organic matter and can be considered a good measure of the change of microbial oxidative activity under environmental pollutions. However, the kinetic characteristic of soil dehydrogenase under heavy metal stresses has not been investigated thoroughly. In this study, we characterized the kinetic characteristic of soil dehydrogenase in 14 soil types, and investigated how kinetic parameters changed under spiked with different concentrations of cadmium (Cd). The results showed that the Km and Vmax values of soil dehydrogenase was among 1.4-7.3mM and 15.9-235.2µMh-1 in uncontaminated soils, respectively. In latosolic red soil and brown soil, the inhibitory kinetic mechanism of Cd to soil dehydrogenase was anticompetitive inhibition with inhibition constants (Ki) of 12 and 4.7mM, respectively; in other soils belonged to linear mixed inhibition, the values of Ki were between 0.7-4.2mM. Soil total organic carbon and Ki were the major factors affecting the toxicity of Cd to dehydrogenase activity. In addition, the velocity constant (k) was more sensitive to Cd contamination compared to Vmax and Km, which was established as an early indicator of gross changes in soil microbial oxidative activity caused by Cd contamination.