Sensitivity of Commercial Enzyme Inhibition Colorimetric Pesticide Residue Rapid Test Kit to a Variety of Pesticides

[Objectives]To fully understand the quality of commercial enzyme inhibition-colorimetric pesticide residue rapid detection kits,so that they can play a greater role in the detection and supervision of agricultural products.[Methods]The sensitivity of 28 kinds of pesticides was determined by using the commercially available enzyme inhibition colorimetric rapid detection kit with Hendu brand.[Results]There was a significant difference in the sensitivity of the kit to each pesticide,and the kit was more sensitive to dichlorvos among the 28 pesticides tested.The sensitivity to methyl isosalifos,dimethoate,isocarbophos,fenthion and phorate was poor,and the sensitivity to quinalphos was different between 3.0 and 2.5 mL.[Conclusions]The large difference of the sensitivity of the enzyme inhibition-colorimetric rapid detection kit for pesticide residues to different kits is a reason for the false positive and false negative test results of the kit,which needs to be considered by relevant personnel.

Enzyme Inhibition Rate Method for Rapid Detection of Organophosphorus and Carbamate Pesticides in Cowpea

[Objectives ] The paper was to explore enzyme inhibition rate method for rapid detection of organophosphorus and carbamate pesticides in cowpea. [ Methods ] Acetylcholinesterase （ACHE） was added to cowpea extract, to determine the inhibition rate of extract against enzyme. The influences of different sampiing methods and sampling parts on detection results were compared. [ Results] The positive rate of standard sampling was 18.18% higher than that of non-stand- ard sampling, and the positive rate of samples collected from cowpea tail was 16.67% higher than that collected from other parts. [ Condmions] Enzyme inhibi- tion rate method is suitable for rapid detection of organophosphorus and carbamate pesticides in cowpea.

Enzyme inhibition assay for pyruvate dehydrogenase complex: Clinical utility for the diagnosis of primary biliary cirrhosis

Primary biliary cirrhosis (PBC) is usually diagnosed by the presence of characteristic histopathological features of the liver and/or antimitochondrial antibodies (AMA) in the serum traditionally detected by immunofluorescence. Recently, new and more accurate serological assays for the detection of AMA, such as enzyme-linked immunosorbent assay (ELISA), immunoblotting, and enzyme inhibition assay, have been developed. Of these, the enzyme inhibition assay for the detection of anti-pyruvate dehydrogenase complex (PDC) antibodies offers certain advantages such as objectivity, rapidity, simplicity, and low cost. Since this assay has almost 100% specificity, it may have particular applicability in screening the at-risk segment of the population in developing countries. Moreover, this assay could be also used for monitoring the disease course in PBC. Almost all sera of PBC-suspected patients can be confirmed for PBC or non-PBC by the combination results of immunoblotting and enzyme inhibition assay without histopathological examination. For the development of a "complete" or "gold standard" diagnostic assay for PBC, similar assays of the enzyme inhibition for anti-2-oxoglutarate dehydrogenase complex (OGDC) and anti-branched chain oxo-acid dehydrogenase complex (BCOADC) antibodies will be needed in future.

In vitro and in vivo cytochrome P450 3A enzyme inhibition by Aframomum melengueta and Dennettia tripetala extracts

Objective: To evaluate the in vitro and in vivo inhibitory effects of two commonly used herbs, Aframomum melengueta(A. melengueta) and Dennettia tripetala(D. tripetala) on CYP 3A enzymes. Methods: In vitro inhibition of the enzymes were assessed with microsomes extracted from female albino rats using erythromycin-N-demethylation assay(EMND) method while their in vivo effects were measured by estimating simvastatin plasma concentrations in rats. Pharmacokinetic parameters were determined using non-compartmental anaysis as implemented in Win Nonlin pharmacokinetic program. Results: EMND assay with intestinal microsomes indicated that aqueous extracts of D. tripetala and A. melengueta significantly(P < 0.05) inhibited intestinal CYP 3A activity at both 50 μg and 100 μg concentrations. Petroleum ether extract of D. tripetala and ethanol extracts of A. melengueta inhibited intestinal CYP3 A activity at 100 μg but not at 50 μg concentrations. All the extracts showed an in vitrodose dependent CYP 3A inhibition with liver microsomes. In vivo analysis showed that pretreatment with the extracts enhanced systemic absorption of simvastatin with reductions in metabolizing enzymes activity as indicated in significant increases in maximal concentration, area under curve, area under moment curve and mean resident time of simvastatin(P < 0.05). Conclusions: Herbal preparations containing these plants' extracts should be used with caution especially in patients on CYP450 3A substrate medications.

Purification and Characterization of Angiotensin I Converting Enzyme Inhibition Peptides from Sandworm Sipunculus nudus

Three angiotensin I converting enzyme(ACE) inhibition peptides were isolated from sandworm Sipunculus nudus protein hydrolysate prepared using protamex. Consecutive purification methods, including size exclusion chromatography and reverse-phase high performance liquid chromatography(RP-HPLC), were used to isolate the ACE inhibition peptides. The amino acid sequences of the peptides were identified as Ile-Asn-Asp, Val-Glu-Pro-Gly and Leu-Ala-Asp-Glu-Phe. The IC_(50) values of the purified peptides for ACE inhibition activity were 34.72 μmol L^(-1), 20.55 μmol L^(-1) and 22.77 μmol L^(-1), respectively. These results suggested that S. nudus proteins contain specific peptides that can be released by enzymatic hydrolysis. This study may provide an experimental basis for further systematic research, rational development and clinical utilization of sandworm resources.

Mobola plum seed methanolic extracts exhibit mixed type inhibition of angiotensinⅠ-converting enzyme in vitro

Objective: To explore the possible inhibitory potentials and mechanism by Mobola plum(Parinari curatellifolia) seeds crude methanol(CE) and flavonoid-rich(FE) extracts on angiotensin-1-converting enzyme(ACE Ⅰ).Methods: The sensitivity and kinetic model of inhibition of CE and FE on ACE Ⅰ using N-[3-(2-furyl)-acryloyl]-Phe-Gly-Gly as enzyme substrate for ACE Ⅰ was evaluated by Michealis Menten approach. The inhibition mechanism was explored from Lineweaver–Burk model and IC_(50) was determined from Cheng–Prusoff empirical analysis.Results: The IC_(50) of CE and FE were 13.54 and 39.38 mg/m L, respectively. Both extracts exhibited mixed type inhibition with the inhibitory constant(K_i) of CE was between0.38 and 0.37 mg/m L while that of FE showed a two-fold increase(1.62 mg/mL and0.28 mg/mL). FE on ACE Ⅰ demonstrated positive cooperativity with a Hill's coefficient of1.89.Conclusions: The study reveals the superior ACE Ⅰ inhibitory potential of CE over FE and suggest that mixed inhibition pattern of the enzyme might be the underlying mechanism of antihypertensive activity.

Synthesis of Novel 4-Thiazolidinone and Bis-Thiazalidin-4-One Derivatives Derived from 4-Amino-Antipyrine and Evaluated as Inhibition of Purine Metabolism Enzymes by Bacteria

Novel 4-thiazolidione and 1,4-bis-thiazolidinone derivatives bearing antipyrine moiety have been obtained from condensation of 4-aminoantipyrine 1 with aromatic/heteroaldehydes followed by cycloaddition with mercaptoacetic acid in nonpolar solvents. Structure of the products has been deduced upon their elemental analysis and spectral measurements. Most of the targets evaluated as enzymatic effect towards some bacteria (E. coli) in compare with Xanthine oxidase (from buttermilk) where the role of compounds is an inhibition of purine metabolism enzymes caused by E. coli.

Studies on Reduced Extent Method for Inhibited Thermokinetics Of Enzyme－Catalyzed Reaction──Inhibition of the laccase－catalyzed oxidation of o－dihydroxybenzene by m－dihydroxybenzene

The thermokinetic reduced extent equations of reversible inhibitions for Michaiels-Menten enzymatic reaction were deduced, and then the criteria for distingushing inhibition type was given and the methods for calculating kinetic parameters, KM,Ki and Urn were suggested. This theory was applied to inverstigate the inhibited thermokinetics of laccase-catalyzed oxidation of o-dihydroxybenzene by m-dihydroxybenzene. The experimental results show the inhibition belongs to reversible competitive type, KM=6.224×10-3 mol L-1, Ki=2. 363 × 10-2 mol. L-1.

Phenolic composition and inhibitory activity of Mangifera indica and Mucuna urens seeds extracts against key enzymes linked to the pathology and complications of type 2 diabetes

Objective:To investigate the phenolic compounds composition and the inhibitory activity of Mangifera indica(M.indica) and Mucuna urens(M.urens) seeds extracts against some key enzymes(-amylase,-glucosidasc and aldose reductase) implicated in the pathology and complications of type 2 diabetes in vitro.Methods:Reverse phase chromatographic quantification of the major flavonoids and phenolic acids in the seeds extracts was carried out using high performance liquid chromatography coupled with diode array detection.The inhibitory activities of the seeds extracts against-amylase and-glucosidase were estimated using soluble starch and p-nitrophenylglucopyranosidc as their respective substrates.Inhibition of aldose reductase activity by the extracts was assayed using partially purified lens homogenate of normal male rat as source of enzyme;inhibition of Fe^(2+)-induced lipid peroxidation by extracts was tested in rat pancreas homogenate.Results:The chromatography result revealed that extracts of both seeds had appreciable levels of some major flavonoids and phenolic acids of pharmacological importance,including gallic acid,chlorogenic acid,caffeic acid,ellagic acid,catechin,rutin,quercitrin.quercetin and kaempferol.Extracts of both seeds effectively inhibited-amylase,-glucosidase and aldose reductase activities in a dose-dependent manner,having inhibitory preference for these enzymes in the order of aldose reductase>-glucosidase>-amylase.With lower half-maximal inhibitory concentrations(IC50)against-amylase,-glucosidase,and aldose reductase,M.indica had stronger inhibitory potency against these enzymes than M.urens.Extracts of both seeds also inhibited Fe2+-induced lipid peroxidation in a dose-dependent pattern,with M.indica being more potent than M.urens.Conclusions:The results obtained provide support for a possible use of M.indica and M.urens seeds in managing hyperglycemia and preventing the complications associated with it in type 2diabetes.

Metabolic enzyme inhibitory abilities, in vivo hypoglycemic ability of palmleaf raspberry fruits extracts and identification of hypoglycemic compounds

The unripe palmleaf raspberry,namely Fupenzi(FPZ),is an important medicinal and edible food.This study aims to evaluate the potential of FPZ extracts prepared with different approaches in attenuating hyperglycemia,gout,Alzheimer’s disease,and pigmentation,to obtain the enriching fraction and to identify the major active compounds.Results indicated that FPZ extracts showed weak activity against acetylcholinesterase,considerable ability against tyrosinase and xanthine oxidase,but excellent inhibition onα-glucosidase.Ultrasound-assisted 40%ethanol extract(40EUS)gave the highest phenolics content,and the bestα-glucosidase inhibition(IC_(50)=0.08μg/mL),which is 877-fold higher than that of positive control acarbose.The 40%ethanol eluting fraction of 40EUS showed the strongestα-glucosidase inhibition with the IC_(50) value of 37.79 ng/mL,it could also effectively attenuate the fasting blood glucose level and oral glucose tolerance of C57BL/6 mice.Twenty-six compounds were identified from 40%ethanol fraction by using HPLC-QTOF-MS/MS,hydrolysable tannins(including 11 ellagitannins and 4 gallotannins)were the major compounds,phenolic acids came to the second.Above results could provide important technical supporting for the further application and research of FPZ in health foods and drugs against diabetes.

Purification and Molecular Docking Study of Angiotensin-I Converting Enzyme (ACE) Inhibitory Peptide from Alcalase Hydrolysate of Hazelnut (<i>Corylus heterophylla</i>Fisch) Protein

Although a number of bioactive peptides are capable of angiotensin I-converting enzyme (ACE) inhibitory effects, little is known regarding the mechanism of hazelnut peptides using molecular simulation. In the present study, gel filtration chromatography, reverse phase-high performance liquid chromatography, and liquid chromatography-electrospray ionization-tandem mass (LC-ESI-MS/MS) were employed for purifying and identifying the ACE inhibitory peptides from hazelnut. To understand the mode of action of these peptides, the interaction between the inhibitory peptides and ACE was investigated. The results identified novel ACE inhibitory peptides Asp-Asp-Glu-Leu-Arg-Gln-Ala (DDELRQA), Asp-Asp-Glu-Leu-Arg-Ala-Ala (DDELRAA), and Asp-Gly-Glu-Leu-Arg-Glu (DGELRE). The binding free energies of DDELRQA, DDELRAA, and DGELRE for ACE were -10.2, -9.0, and -8.8 kcal/mol, respectively. This study proves the high stability of ACE inhibitory peptides derived from hazelnut against different temperature and pH of processing.

Enzymes inhibitory property,antioxidant activity and phenolics profile of raw and roasted red sorghum grains in vitro

Whole grain cereals are important dietary sources for management of metabolic diseases due to the bioactive components they contain.Hence,this study investigated enzymes(pancreatic lipase,-amylase,-glucosidase,xanthine oxidase and angiotensin 1-converting enzyme)inhibitory property,antioxidant activity and phenolics profile of raw and roasted red sorghum(Sorghum bicolor)grains in vitro.Extracts of flours of raw and roasted(150◦C and 180◦C,for 20 min)grains were assayed for enzymes inhibitory and antioxidant activities using spectrophotometric methods;while their phenolic constituents were characterized using HPLC-DAD.The raw grains exhibited strong enzymes inhibitory and antioxidant activities,and contained phenolic acids(gallic,chlorogenic,caffeic,ellagic and p-coumaric acids)and flavonoids(quercetin,luteolin and apigenin).However,whereas the enzymes inhibitory activity and levels of the phenolic compounds in the grains decreased significantly(p<0.05)with increasing roasting temperature,the antioxidant activity increased.Hence,roasting at high temperature may not be recommended for the optimum retention of the enzymes inhibitory property and phenolic compounds of red sorghum grains.

Methanol extracts of Brachystegia eurycoma and Detarium microcarpum seeds flours inhibit some key enzymes linked to the pathology and complications of type 2 diabetes in vitro

The inhibitory effect of methanol extracts of Brachystegia eurycoma and Detarium microcarpum seeds flours on some key enzymes[α-amylase,α-glucosidase and aldose reductase(AR)]linked to the pathology and complications of type 2 diabetes(T2D);and their antioxidant properties were evaluated.The antioxidant properties evaluated were DPPH•and ABTS•^+scavenging abilities,reducing power,and antioxidant phytochemicals(total phenolics,tannins,total flavonoids and total saponins).Extracts of both flours inhibitedα-amylase,α-glucosidase and AR in a dose-dependent manner.The half-maximal inhibitory concentrations(IC50)of B.eurycoma onα-amylase,α-glucosidase,AR and lipid peroxidation were lower than those of D.microcarpum,indicating that it had stronger inhibitory potency than D.microcarpum.B.eurycoma also had significantly(P<0.05)higher DPPH•and ABTS•^+scavenging abilities,and reducing power than D.microcarpum.The antioxidant phytochemicals(total phenolics,tannins,total flavonoids and total saponins)were also significantly(P<0.05)higher in B.eurycoma than D.microcarpum.The inhibitory effect of B.eurycoma and D.microcarpum extracts onα-amylase,α-glucosidase and AR activities may be attributed to the combined action of their polyphenols and total saponins,and this may be a possible mechanism of action providing support for their use in managing hyperglycemia and the complications of T2D.

Inhibitory effect of Cr(Ⅵ) on activities of soil enzymes

To evaluate the influence of various Cr(Ⅵ) concentrations (0.05, 0.25, 0.50, 1.00 and 2.00 g/kg) on the activity of soil enzymes, the activities of catalase, polyphenol oxidase, dehydrogenase, alkaline phosphatase in soils were investigated in the incubation experiment with a period of 35 d. The results indicate that all the tested Cr(Ⅵ) concentrations significantly inhibit dehydrogenase activity by over 70% after 35 d. The activity of alkaline phosphatase is slightly inhibited during the whole experiment except for on the day 7. Cr(Ⅵ) has no obvious effect on the activity of catalase in soil. On the contrary, Cr(Ⅵ) stimulates the activity of polyphenol oxidase. The results suggest that dehydrogenase activity can be used as an indicator for assessing the severity of chromium pollution.

Kinetic and binding equilibrium studies of dihydroflavonol 4-reductase from Vitis vinifera and its unusually strong substrate inhibition

Dihydroflavonol 4-reductase (DFR), a member of the short-chain dehydrogenase family, catalyzes the last common step in the biosynthesis of flavan-3-ols and condensed tannins. Initial rates of DFR were measured by monitoring the 340-nm absorbance decrease resulting from the joint consumption of dihydroquercetin (DHQ) and NADPH, as a function of pH, temperature and ionic strength. At pH 6.5 and 30o C, substrate inhibition was observed above 30 μM DHQ. At lower/non-inhibitory DHQ concentrations, NADP+ behaves as a competitive inhibitor with respect to NADPH and as a mixed inhibitor with respect to DHQ, which supports a sequential ordered mechanism, with NADPH binding first and NADP+ released last. Binding-equilib-rium data obtained by means of the chromatographic method of Hummel and Dreyer at pH 7.5 and by isothermal calorimetric titration at pH 6.5 led to the conclusion that ligands of the apoenzyme included NADPH, NADP+ and DHQ. The mechanism which best accounts for substrate inhibition at pH 6.5 in the absence of product involves the formation of a binary non-productive E.DHQ complex. Thus, a productive ternary complex cannot be formed when DHQ binds first. This mechanism of inhibition may prevent the accumulation of unstable leucoanthocyanidins within cells.

Inhibition of Porcine Small Intestinal Sucrase by Validamine

As an important medicinal intermediate with broad uses, validamine, an aminocyclitol, isolated from the enzymolysis broth of validamycins, has gained more and more attention. The absolute configuration of validamine is similar to that of a-D-glucose, and it demonstrates powerful inhibition activity on glycosidase. In this paper, the inhibitory effect of validamine on porcine small intestinal sucrase was investigated. Validamine was found to be a potent, competitive inhibitor to porcine small intestinal sucrase in vitro with an IC50 value of 6.85 X 10(-4) mol center dot L-1. Validamine exhibited a dose-dependent inhibition effect on porcine small intestinal sucrase, whereby the inhibition interaction of validamine and porcine small intestinal sucrase was a fast binding process. The inhibition of validamine on porcine small intestinal sucrase was pH-dependent.

THE INHIBITION EFFECT OF ^(211)At-Te COLLOID AND Na^(211) At INJECTIONS ON MURINE EHRLICH ASCITES TUMOR CELLS

Na211 At and 211 At-Te colloid injections are prepared. It has been demonstrated that the 211 At-Te colloid is stable in vivo and vitro, and can be applied in the study of biology and medicine. In the report, the model of Murine Ehrlich Ascites Cells cultured in vivo and vitro is elected for a series of experiments. It has been proved that Na 211At and 211At-Te colloid injections possess an inhibition effect on tumor cells. The inhibition effect was expressed in surviving of the mice and inhibiting growth of tumor as well as the changes of enzyme activity. Meanwhile, it was also noticed that Na 211 At and 211 At-Te colloid injections of various dose inhibited the absorb of pyrimidine nucleosides in Murine Ehrlich Ascites Cells. And the effect isn’t reversible. It is closely related to the dose administratered and 50% inhibition rate needs about 1.48×105 Bq/ml culture.

Studies on Enzyme Kinetics by Microchip and Related Techniques

Both conventional and microchip-based capillary electrophoresis(CE) technologies have been used for the analysis of enzymes. Practical procedures of using CE to determine the Km and Vmax values of an enzyme have been developed. By studying the inhibition to the enzyme, it is possible to select a suitable drug candidate. When compared with the conventional CE method, single lane microchip-based method can improve the speed for the assay three times. By using multiple lane-based microchip, the speed can be further increased.

Toxicity of Tagetes erecta Essential Oil Based Combinatorial Formulations on Various Metabolic Enzymes in Indian White Termite Odontotermes obesus

Plant essential oils and their constituents have proven to be very effective against insects, especially termites. They are the best alternative to synthetic pesticides that are harmless to the environment and human health. In the present study, different enzymes, namely alkaline phosphatase, acid phosphatase, glutamate-oxaloacetate transaminase, glutamate-pyruvate transaminase and acetylcholinesterase, were evaluated in Indian termite Odontotermes obesus in a combination preparation based on Tagetes erecta essential oil. For the study of anti-termite effects against worker termites were treated topically with 40% and 80% of the 24-hour LD50 values of various combination formulations. Subsequent bioassays at 40% and 80% of LD50 of combinatorial preparations S-AST-A, S-AST-B, and S-AST-C resulted in a significant (p > 0.05) decrease in glutamate-oxaloacetate transaminase. After 16 hours of treatment, they were 87.92%, 80.17%, 89.30%, 79.17%, 81.92% and 73.17% respectively. In vivo exposures of 40% and 80% of the LD50 of the combination formulation caused a highly significant (p > 0.05) reduction (p > 0.05) of all test enzymes tested compared to controls. The effects of different oil formulations exhibited time- and dose-dependent responses, resulting in physiological changes in the concentrations of various enzymes. The combined mixture of Tagetes erecta essential oils has significantly better anti-termite ability compared to inorganic insecticides. Findings from this study will help support termite control in fields, gardens and homes in a sustainable way, without the downsides of insecticide resistance and pollution. These could potentially be used to produce commercial formulations for use against pests.

Toxic Effects of Citrus maxima Based Combinatorial Formulations on Important Metabolic Enzymes in Indian White Termite Odontotermes obesus

In the present study, Citrus maxima essential crude oil extract was used to prepare combinatorial formulations and workers of Indian white termite Odontotermes obesus were treated topically with 40% and 80% of 24 hr LD50 values of these formulations. In subsequent bioassays levels of various enzymes i.e. alkaline phosphatase, acid phosphatase, glutamate oxaloacetate transaminase, glutamate pyruvate transaminase and acetylcholinesterase were evaluated to determine the anti-termite efficacy of Citrus maxima essential oil based combinatorial formulations. S-RET-A, S-RET-B and S-RET-C caused significant (p > 0.05) decrease in glutamate oxaloacetate transaminase i.e. 87.47%, 86.81% and 81.77% & 82.04%, 79.39% and 74.75% respectively at 16 h treatment. In vivo exposure of 40% and 80% of LD50 of combinatorial formulations caused very significant (p > 0.05) reduction in all the test enzymes i.e. alkaline phosphatase, acid phosphatase, glutamate oxaloacetate transaminase and glutamate pyruvate transaminase expect acetylcholinesterase levels after 16 h all tested treatments in comparison to control. Both dose-response and time period were found important in physiological alteration in levels of various enzymes. Combinatorial mixtures of Citrus essential oils have shown synergistic activity against termites. The research findings of the present study would help termite control in crop fields, gardens and houses in a sustainable way.