123 results about "Pyruvate oxidase" patented technology

Succinic acid is produced by using a coryneform bacterium which has succinic acid-producing ability and has been modified so that an activity of pyruvate oxidase is decreased.

The present invention belongs to the field of medical detection technology. The reagent kit for blood ammonia diagnosis includes buffering solution, adenosine triphophate, glutamic acid, pyruvic acid, alcohol, oxidized coenzyme, glutamine synthetase, pyruvate oxidase, hydrogen peroxidase, aldehyde dehydrogenase and stabilizer. Through mixing the sample and reagent in certain volume ratio to produce enzyme coupling reaction, and detecting in biochemical analyzer the main wavelength absorbency change speed, the blood ammonia content is measured. The present invention can obtain the measurement result in biochemical analyzer in high sensitivity, high precision and no contamination of various foreign and internal matters.

Methods and compositions are provided for determining ADP in the presence of ATP. These comprise including among the assay reagents at least one of the correcting components creatine phosphokinase and phosphocreatine, pyruvate kinase and phosphoenolpyruvate, peroxidase and a non-interfering peroxidase substrate, and catalase. One aspect of the method employs formation of hydrogen peroxide from the ADP by pyruvate kinase, phosphoenolpyruvate and pyruvate oxidase. The hydrogen peroxide is then determined. A combined reagent having all of the reagents may optionally include a peroxidase when the hydrogen peroxide is to be enzymatically determined. A peroxidase substrate is added to the sample in conjunction with the peroxidase substrate reagent, the mixture incubated and depending on whether the peroxidase substrate is a fluorescer or chemiluminescer, the mixture may be illuminated with excitation light and the emitted light determined as a measure of the ADP in the sample.

Purpose of the invention is to provide a reagent bar capable of half quantitative / quantitative detecting whether content of glutamic-pyruvic transaminase (GPT) inside human body is normal or not rapidly, and relevant fabricating method. The reagent bar includes PVC hard strip for supporting reaction. The hard strip possesses carrier film of adsorbing L-alanine, pyruvate oxidase, and alpha-ketoglutarate. Belonging to fast test in micro scale, the disclosed product is one-off consumption material for testing. Range for detecting GPT is 0-1000U / L.

Methods and compositions are provided for determining ADP in the presence of ATP. These comprise including among the assay reagents at least one of the correcting components creatine phosphokinase and phosphocreatine, pyruvate kinase and phosphoenolpyruvate, peroxidase and a non-interfering peroxidase substrate, and catalase. One aspect of the method employs formation of hydrogen peroxide from the ADP by pyruvate kinase, phosphoenolpyruvate and pyruvate oxidase. The hydrogen peroxide is then determined. A combined reagent having all of the reagents may optionally include a peroxidase when the hydrogen peroxide is to be enzymatically determined. A peroxidase substrate is added to the sample in conjunction with the peroxidase substrate reagent, the mixture incubated and depending on whether the peroxidase substrate is a fluorescer or chemiluminescer, the mixture may be illuminated with excitation light and the emitted light determined as a measure of the ADP in the sample.

The invention relates to a kit for diagnosing / measuring potassium (ions) by utilizing the technologies of the enzymic colorimetry and the enzyme linked immunosorbent assay. The invention further relates to a method, a principle and the composition and the components of a reagent for measuring the concentration of the potassium (ions), and belongs to the technical field of medical / food / environmental inspection and measurement. The main components of the kit include a buffer solution, reduced coenzyme, adenosine diphosphate, a phosphoenolpyruvic acid, coenzyme A, pyruvate kinase, pyruvate oxidase, NADH peroxydase and a stabilizer. Through mixing a sample and the reagent by a certain volume ratio, a series of enzymatic reactions occur, then the reactant is placed under an ultraviolet / visible light analyzer, and the velocity of the decrease in absorbance at 340 nm of the dominant wavelength is detected, thereby measuring the concentration of potassium (ions).

Succinic acid is produced by the use of coryneform bacteria having the capability of producing succinic acid and engineered so as to lower the pyruvate oxidase activity thereof.

The invention provides an immobilization method of pyruvate oxidase, which adopts a polylysine carrier to immobilize the pyruvate oxidase on an electrode. 20% of pyruvate oxidase activity can be increased by adopting that method of the invention, and the obtained immobilized pyruvate oxidase has high activity, good tolerance to pH and temperature, and important guiding significance for the development and application of subsequent biosensors.

The invention discloses a fed-batch fermentation process for producing pyruvate oxidase through fermentation, which comprises: culturing recombinant escherichia coli which contains a pyruvate oxidase expression cassette under the culture temperature about 37DEG C, thereby expressing the pyruvate oxidase, lowering the temperature from about 37DEG C to about 32 DEG C after the initial growth stage of the recombinant escherichia coli is finished, beginning feeding, controlling the dissolved oxygen level between 30% and 40%, then, lowing the temperature for 1DEG C each four hours, and keeping the temperature constant after the temperature is lowered to 32DEG C until the fermentation is finished. The method of the invention increases the output of the pyruvate oxidase under the condition that the cost is not changed, the fermentation process is easily controlled, and thereby the method of the invention has extremely good industry application value.

The invention discloses a method for preparing acetyl phosphate on the basis of biological enzymes. The method comprises the following steps: carrying out enzyme catalysis reaction on lactic acid and / or lactate and phosphoric acid and / or phosphate and / or biphosphate by using an immobilized oxidase lactate-immobilized pyruvate oxidase composite enzyme as a catalyst; and after the reaction finishes, filtering and precipitating to obtain the high-purity acetyl phosphate. The method is simple to operate, has the advantages of mild reaction conditions, high raw material conversion rate, low cost and environmental protection, and satisfies the industrial production.

The invention discloses a glutamic oxalacetic transaminase electrochemical detection composition, comprising a first composition and a second composition, wherein the first composition comprises water, first buffer solution, first electrolyte, a first surface active agent, a first electronic mediator and pyruvate oxidase, and the second composition comprises water, second buffer solution, second electrolyte, a second surface active agent, a second electronic mediator, L-aspartic acid, L-alanine, oxaloacetic acid decarboxylase and pyruvate oxidase. The invention also discloses application of the glutamic oxalacetic transaminase electrochemical detection composition. The invention also discloses a glutamic oxalacetic transaminase electrochemical sensor and a glutamic oxalacetic transaminasedetection method.

The present invention discloses a glutamic-pyruvic transaminase electrochemical detection composition. The composition comprises a first composition and a second composition. The first composition comprises water, a first buffer solution, a first electrolyte, a first surface active agent, a first electron mediator and pyruvate oxidase. The second composition comprises water, a second buffer solution, a second electrolyte, a second surface active agent, a second electron mediator, [alpha]- ketoglutaric acid, L- alanine and the pyruvate oxidase. The present invention also discloses an application of the glutamic-pyruvic transaminase electrochemical detection composition. The present invention also discloses a glutamic-pyruvic transaminase electrochemical sensor and a glutamic-pyruvic transaminase detection method.

The invention belongs to the field of bioengineering and provides a culture medium mainly using glycerol as a carbon source. The invention also provides a method for producing pyruvate oxidase by using the culture medium. The present invention discloses for the first time that glycerol is used as a carbon source in the production of pyruvate oxidase, which enables recombinant Escherichia coli to continuously express pyruvate oxidase during the whole fermentation process, greatly improving the yield of pyruvate oxidase; and, adopting the present invention The culture medium is fermented, and the pH value is stable throughout the fermentation process, and the fermentation process is greatly simplified.

The invention is about the measuring method of inorganic Phosphates and its diagnosis reagent box. Producing hydroperoxide by reacting pyruvate oxidase with pyruvate under the existence of Inorganic Phosphates , then causing enzyme-coupled reaction with peroxidase and oxidating the colorless reduced chromogen combination to quinoneimine chromogen or indamide chromogen dyer with color, testing the variation of dominant wave-length 400ú¡600nm absorbance during the reaction and finally measuring the content of Inorganic Phosphates . This method has high specificity and would not be contaminated by material of internal and exogenous sources, and the result is precise and accurate. Diving the diagnosis reagent box into double-dose or three-dose can reduces the cross interaction of each element, keeps the stability of the reagent and deposits chronically. Using this method can realize the fast testing in common ultraviolet / visible light analyzer or semiautomatic / automatic analyzer and doesní»t require special or additional apparatus, so the cost is low. Thus, this method can be easily promoted and applied in the whole industry.

The invention relates to a method for improving photosynthetic hydrogen production efficiency of algae. The method expresses pyruvate oxidase and catalase in hydrogen production algae with no influence on normal metabolism and growth of the hydrogen production algae, so that oxygen concentration in the hydrogen production algae can be decreased effectively; photorespiration function is reduced; and photosynthetic hydrogen production efficiency of the hydrogen production algae is increased greatly. The method provides novel approaches for improving photosynthetic hydrogen production efficiency, and provides novel ideas for bioenergy research and industrialization.

The invention belongs to the technical field of clinical in vitro diagnostic reagents and relates to a dry chemical reagent tablet for quantitatively measuring the activity of alanine aminotransferaseand a preparation method of the dry chemical reagent tablet. The dry chemical reagent tablet comprises four laminations which are respectively a support layer, a color developing layer, a reagent layer and a diffusion layer distributed sequentially; reagents in the reagent layer include a buffer system, a hydrophilic colloid, a pyruvate oxidase, peroxidase, thiamine pyrophosphate, a chelating agent, an activator and a surfactant; and reagents in the color developing layer include a chromogen, a hydrophilic colloid, and a surfactant. The dry chemical reagent tablet and the preparation method thereof of the invention have the advantages of simple operation process, high stability, high good repeatability and high accuracy of detection results, quantitative detection, wide linear range, simple and convenient preparation process, easiness in operation, low risk and low pollution. According to the dry chemical reagent tablet for quantitatively measuring the activity of the alanine aminotransferase and the preparation method of the dry chemical reagent tablet of the invention, the reagents do not need to be prepared, and samples can be directly added dropwise; the reagent tablet is dryand is free of moisture; and the diffusion layer has a filtering function, so that hemolysis, bilirubin and ascorbic acid samples exert no significant interference on measurement results.

The invention provides an application of a fluorescent gold nanocluster in fluorescent detection of pyruvic acid and / or pyruvate oxidase. The fluorescent gold nanocluster probe is used for detecting the pyruvate and / or the pyruvate oxidase, has the characteristics of high sensitivity, good selectivity, wide linear range and the like. According to the application of the fluorescent gold nanoclusterin the fluorescent detection of the pyruvic acid and / or the pyruvate oxidase, the detection of the pyruvic acid by a method for emitting fluorescence by the fluorescent nanocluster not only avoids interference of other factors of biology, but also selectively detects the concentration of the pyruvic acid in blood and urine, and can detect the concentration of the pyruvic acid non-invasively. A fluorescence emission method is used to detect the concentration of the pyruvic acid instead of using pre-prepared fluorescent metal nanocluster to minimize the interference that may be caused by falsesignals in the experiment. The fluorescent gold nanocluster generated simultaneously during the detection process has excellent fluorescence properties and great significance in applications in the fields of optical materials, biomedicine and the like.

The invention provides a method for obtaining pyruvate oxidase in recombinant escherichia coli. The method comprises the steps as follows: inoculating recombinant escherichia coli pET28a-pod containing an expressed gene of pyruvate oxidase onto a solid culture medium for activation to obtain activated recombinant escherichia coli pET28a-pod; inoculating the activated recombinant escherichia coli pET28a-pod onto an LB culture medium placed on a shaker and shaking to obtain a seed solution of the activated recombinant escherichia coli pET28a-pod; inoculating the seed solution onto a TB culture medium for culture, adding an inducer for induction after the concentration is cultured to be a first cell concentration, continuously adding a hydrochloric acid solution for fermentation until a pH value is 5.5-6.5 when the concentration reaches a second cell concentration, and then performing induction continually to obtain a bacteria solution I; taking the bacteria solution I for centrifugation,pouring out supernatant, then washing twice with a PBS solution, and then resuspending with an equal volume of the PBS solution to obtain a resuspension solution; performing ultrasonication on the resuspension solution to obtain a ultrasonicated bacteria solution II; and taking the bacteria solution II for centrifugation, and collecting the supernatant containing the pyruvate oxidase.

The present invention relates to a method for measuring lactic acid with high sensitivity, also relates to a lactic acid detection kit, and belongs to the technical fields of medical inspection and lactic acid bacterium fermented food measurement. The lactic acid detection kit is an important detection index for high altitude reaction, athlete training, severe diabetes and body acidosis caused byother reasons, and also an index for detecting lactic acid fermented food. The method uses an original lactate oxidase method, then pyruvate oxidase is introduced, one molecule of lactic acid reacts with the lactate oxidase and pyruvate oxidase to generate two molecules of hydrogen peroxide, then a Trinder reaction is conducted to generate red color, absorbance at a wavelength of 500-600 nm is detected, and content of the lactic acid is calculated. The method has advantages of high sensitivity, good repeatability, wide linearity and good stability, and is very practical for routine laboratoryapplication.

The invention relates to an inorganic phosphorus (phosphate radical) diagnosis / determination kit using the technology of an enzyme colorimetric method and an ELISA method, a method for determining inorganic phosphorus (phosphate radical) concentration and composition and components of a reagent, and belongs to the technical field of test and determination of medical science, food, and environment. The kit comprises the following main components: buffer solution, coenzyme, pyruvic acid, acetaldehyde, pyruvate oxidase, alcohol oxidase, alcohol dehydrogenase and a stabilizer. The method comprises the following steps: mixing a sample and the reagent according to a certain volume ratio to perform a series of enzymic reaction, and then placing reactants under a UV / visible light analyzer to test the ascending speed of absorbance at the position where the dominant wave length is 340nm to determine the inorganic phosphorus (phosphate radical) concentration.

The invention relates to a kit for diagnosing / measuring kalium (ions) by utilizing the technologies of the enzymic colorimetry and the enzyme linked immunosorbent assay. The invention further relates to a method, a principle and the composition and the components of a reagent for measuring the concentration of kalium (ions), and belongs to the technical field of medical / food / environmental inspection and measurement. The main components of the kit include a buffer solution, coenzyme, adenosine diphosphate, enolphosphopyruvate, coenzyme A, pyruvate kinase, pyruvate oxidase, NAD(P)H oxidase and a stabilizer. Through mixing a sample and the reagent by a certain volume ratio, a series of enzymatic reactions occur, then the reactant is placed under an ultraviolet / visible light analyzer, and the velocity of the increase in absorbance at 340 nm of the dominant wavelength is detected, thereby measuring the concentration of kalium (ions).

The invention is about the measuring method of Inorganic Phosphates and its diagnosis reagent box. Producing carbon dioxide by reacting pyruvate oxidase with pyruvate under the activation ofInorganic Phosphates in the sample of plasma or serum and so on, and producing oxaloacetic acid by reacting carbon dioxide and phosphoenolpyruvate under the existence of phosphoenolpyruvate carboxylase, and then transferring oxidized coenzyme to reduced coenzyme by reacting oxaloacetic acid and malic acid dehydrogenase. Testing the descending range of dominant wave-length340nm absorbance and finally measuring the content of Inorganic Phosphates. This method has high specificity and would not be contaminated by material of internal and exogenous sources, and the result is precise and accurate. Diving the diagnosis reagent box into double-dose or three-dose can reduces the cross interaction of each element, keeps the stability of the reagent and deposits chronically. Using this method can realize the fast testing in common ultraviolet / visible light analyzer or semiautomatic / automatic analyzer and doesní»t require special or additional apparatus, so the cost is low. Thus, this method can be easily promoted and applied in the whole industry.

The invention provides a glutathione reductase determination kit, and a preparation method and applications thereof, and relates to the technical field of biochemical reagent determination. The kit includes a reagent R1 and a reagent R2; the reagent R1 includes a phosphate buffer solution, ascorbic acid oxidase, pyruvate oxidase, potassium ferrocyanide, oxidized glutathione, MgCl2, FAD, a preservative and a surfactant; and the reagent R2 includes a glycine buffer solution, NADPH, the preservative and a protective agent. Through the adding of EGTA and dithiothreitol in the reagents, the linearrange of the kit can be enlarged, and the stability of the kit can be enhanced. The preparation method and applications of the kit are also provided; and the kit can accurately determine glutathione reductase.

The invention relates to a carbon dioxide diagnosis / determination kit using the technology of an enzyme multiplication method, an enzyme colorimetric method and an ELISA method, a method for determining carbon dioxide concentration, and composition and components of a reagent, and belongs to the technical field of test and determination of medical science, food, and environment. The kid comprises the following main components: buffer solution, reduced coenzyme, hydrogen peroxide, acetylcoenzyme A, ferricytochrome b1, pyruvate oxidase, pyruvate dehydrogenase, aldehyde dehydrogenase, and a stabilizer. The method for determining carbon dioxide concentration comprises the following steps: mixing a sample and the reagent according to a certain volume ratio to perform a series of enzymic reaction, and then placing reactants under a UV / visible light analyzer to test the descending speed of absorbance at the position where the dominant wave length is 340nm to determine the concentration of carbon dioxide.

The invention relates to a kit for diagnosing / mensurating carbon dioxide by utilizing the technologies of the enzymic colorimetric method and the enzyme linked immunosorbent assay. The invention further relates to a method, a principle and the composition and the components of a reagent for mensurating the concentration of the carbon dioxide, and belongs to the technology field of medical / food / environmental inspection and measurement. The main components of the kit include a buffer solution, reduced coenzyme, pyruvate oxidase, alanine dehydrogenase, hydrogen peroxide, acetyl coenzyme A, ammonia ions and a stabilizer. Through mixing a sample and the reagent by a certain volume ratio, an enzymatic reactions occurs, then the reactant is placed under an ultraviolet / visible light analyzer, and the degree / velocity of the decrease in absorbance at 340nm of the dominant wavelength is detected, thereby mensurating the concentration of the carbon dioxide.

The invention relates to a monoamine oxidase diagnostic / measuring reagent (kit) which utilizes enzyme colorimetry and enzyme-linked method technologies, and meanwhile, the invention also relates to a method for measuring monoamine oxidase active concentration and compositions and components of the reagent, which belong to the technical field of medical inspection and measurement. The main components of the reagent (kit) of the invention comprise a buffer solution, coenzyme, an amine compound, sodium bicarbonate, acetyl coenzyme A, acetaldehyde, pyruvate oxidase, acetaldehyde dehydrogenase and a stabilizer; and the method comprises the following steps of: enabling a sample and the reagent to generate a series of enzymatic reactions through mixing the sample and the reagent according to a certain volume ratio, then placing the reactants under an ultraviolet / visible light analyzer and detecting the rising speed of the absorbance at a position with the dominant wavelength of 340nm to measure and calculate the active concentration magnitude of monoamine oxidase.

The present invention belongs to the field of medical detection technology. The reagent kit for blood ammonia diagnosis includes buffering solution, adenosine triphophate, glutamic acid, pyruvic acid, phosphoenolpyruvic acid, reduced coenzyme, glutamine synthetase, pyruvate oxidase, phosphoenolpyruvate carboxylase, malate dehydrogenase and stabilizer. Through mixing the sample and reagent in certain volume ratio to produce enzyme coupling reaction, and detecting in biochemical analyzer the main wavelength absorbency change speed, the blood ammonia content is measured. The present invention can obtain the measurement result in biochemical analyzer in high sensitivity, high precision and no contamination of various foreign and internal matters.

The invention relates to a monoamine oxidase diagnostic / measuring reagent (kit) which utilizes enzyme colorimetry and enzyme-linked method technologies, and meanwhile, the invention also relates to a method for measuring monoamine oxidase active concentration and compositions and components of the reagent, which belong to the technical field of medical inspection and measurement. The main components of the reagent (kit) of the invention comprise a buffer solution, reduced coenzyme, an amine compound, sodium bicarbonate (carbon dioxide), acetyl coenzyme A, arginine, pyruvate oxidase, octopine dehydrogenase and a stabilizer; and the method comprises the following steps of: enabling a sample and the reagent to generate a series of enzymatic reactions through mixing the sample and the reagent according to a certain volume ratio, then placing the reactants under an ultraviolet / visible light analyzer and detecting the descending speed of the absorbance at a position with the dominant wavelength of 340nm to measure and calculate the active concentration magnitude of monoamine oxidase.