214 results about "Phospholipase A2" patented technology

A universal sauce base, i.e. an edible composition, having a low pH for use in hot or cold food applications that is microbiologically stable, heat stable and freeze-thaw tolerant. The universal sauce base has an oil-in-water emulsion and comprises water, vegetable oil, starch, phospholipase A2 modified egg yolk and inorganic acid acidulent including at least phosphoric acid and other ingredients. The universal sauce base has a bland and non-sour flavor, can be used in a wide variety of food applications and can be combined with a wide range of flavors and other ingredients.

The invention relates to a lipoprotein phospholipase A2 assaying reagent and a preparation method thereof, and aims to ensure the characteristics of high reagent accuracy and convenience in preparation method. The invention adopts the technical scheme that the lipoprotein phospholipase A2 assaying reagent comprises the following components: a, a lipoprotein phospholipase A2 reagent 1; b, a lipoprotein phospholipase A2 reagent 2; and c, a liquid lipoprotein phospholipase A2 reference product; the preparation method comprises the following steps: (1) for the lipoprotein phospholipase A2 reagent 1: uniformly mixing; (2) for the lipoprotein phospholipase A2 reagent 2: (1) taking a suspension, (2) reacting with a mixture, (3) obtaining the suspension, (4) regulating the concentration, (5) adding the suspension in step (3) into the solution obtained in the substep (4), (6) reacting, (7) adding ethanol amine, and (8) performing centrifugal treatment; and (3) for the liquid LP-PLA2 (lipoprotein phospholipase A2) reference product: mixing according to a formulation, arranging according to contents, or adding a purified product in the mixed solution.

This invention relates to a method for measuring enzymatically active Lipoprotein Phospholipase A2 (Lp-PLA2) in a sample. Further, this invention relates to a Hybrid Immunocapture method for measuring enzymatically active Lp-PLA2 in a sample. Specifically, this invention relates to a Hybrid Immunocapture method for measuring enzymatically active Lp-PLA2 in a sample utilizing an enzymatically active Lp-PLA2 standard. In addition, this invention relates to a kit for measuring enzymatically active Lp-PLA2 in a sample. Specifically, this invention relates to a kit for measuring enzymatically active Lp-PLA2 in a sample containing an enzymatically active Lp-PLA2 standard.

The invention discloses a method for degumming vegetable fat by using phosphatidase A2. The method comprises the following steps of: A, heating crude oil to 40-50 DEG C; B, adding a 45 percent (W/W) citric acid solution till the final concentration is 0.01-0.02 percent, stirring (at the speed of more than or equal to 40 revolutions per minute) or homogenizing for 10 seconds, and preserving heat for 30 minutes; C, adding water in an amount of 1-5 percent based on the weight of the crude oil, and stirring uniformly; and D, adding enzyme in an amount of 0.005-0.015 percent (W/W) based on the weight of the crude oil, stirring and reacting for 1-3 hours, and centrifuging or naturally precipitating for separating degummed oil from a gum deposit. The method is practicable, and is easy and convenient to operate; when phosphatidase A2 is used for degumming, high-value enzyme hydrolyzed phospholipid can be obtained, so that energy consumption is lowered; and expressed (or leached) crude oil with very high phosphorus content or hydrated degummed oil with relatively low phosphorus content can be directly used for degumming in the method, so that the method has a wider application range.

The invention discloses an engineering bacterium for producing a PL A2 and applications thereof. A preparation method of the PLA2 comprises the following steps: A, preparing a PLA2 gene: designing a primer PCR and amplifying a PLA2 gene fragment, and cloning a DNA fragment containing the PLA2 gene (a sequence is SEQ ID NO.2) with an amplified fragment plaT as a probe; B, constructing a recombinant plasmid: cloning a Bam H1 restricted fragment with the size of 3 kb which contains the PLA2 gene to a high-copy plasmid pHZ132 to obtain a recombinant plasmid pLH 001; and C, constructing the engineering bacterium: introducing the plasmid pLH001 into a streptomyces lividans 1326 of a host bacterium through protoplast transformation to obtain a streptomyces lividans LH001 of the engineering bacterium. The engineering bacterium has good stability and the introduced plasmid is not easy to lose. The secretory active PLA2 (an amino acid sequence is SEQ ID NO.1) can be generated through liquid fermentation, the enzymatic activity of a fermentation liquid is equal to or more than 2,000 U/mL, and the enzyme production capability is better than present levels. The PLA2 can be applied to fields of vegetable oil degumming, lysophospholipid preparation and the like.

The invention discloses a lipoprotein-related phospholipase A2 content detection kit and a preparation method thereof, belonging to the field of detection of lipoprotein-related phospholipase A2 content. The kit consists of a reagent I and a reagent II which are independent of each other, wherein the reagent I comprises the following components: a biological buffer agent, a coagulant, a preservative, a chelating agent and water; the reagent II comprises the following components: latex particles coated with a lipoprotein-related phospholipase A2 antibody, a biological buffer agent, a surfactant, a preservative, a suspending agent and water. The latex particles coated with the lipoprotein-related phospholipase A2 antibody are prepared by adopting latex particles with the particle size of 120-180nm, and the stability, sensitivity and wide detection linear range of the kit can be effectively guaranteed. Meanwhile, the latex particles coated with the lipoprotein-related phospholipase A2 antibody are prepared by adopting a specific method, so that the bottle-opening stability and detection specificity of the kit are relatively good, and the product is suitable for popularization and application of clinical detection.

The invention provides an immunological detecting kit, a preparation method and a using method thereof. The kit comprises a protein antibody which is to be detected and is marked by a marker capable of directly detecting, wherein the protein to be detected is the protein related to heart cerebrovascular disease examination. The protein to be detected comprises one or more of fibrinogen, C-reactive protein, thrombus precussor protein, creatine kinase and human body lipoprotein related phospholipase A2. The abundance or the concentration of the protein to be detected in the sample to be detected can be detected in one step by using the kit, so the step and the time are saved; and compared with the traditional method requiring signal cascade amplification such as enzyme-linked immuno sorbent assay (ELISA) and the like, the accuracy is improved.

Described herein are peptides from secretory phospholipase A₂-IB and antibodies that can be used to reduce the contribution of the gastrointestinal tract to inflammatory processes including systemic inflammatory responses. Specifically, antibodies that bind specifically a peptide from secretory phospholipase A₂-IB prevent death in a mouse model of LPS-induced endotoxemia. The antibodies described herein are particularly useful to treat systemic inflammatory response syndromes, including sepsis.

The invention relates to a method for preparing phosphatidylserine (2-DHA-PS) with docosahexaenoic acid at the sn-2 bit through high-activity phospholipase A2 and high-activity phospholipase D. Directed evolution is achieved through the overlapping PCR technology so that the high-activity phospholipase A2 and the high-activity phospholipase D can be obtained; the 2-DHA-PS is prepared through catalysis by means of the high-activity phospholipase A2 and the high-activity phospholipase D, phosphatidylserine is generated through phosphatidylcholine and serine under the catalysis of the high-activity phospholipase D first, and then the 2-DHA-PS is generated through the phosphatidylserine and the docosahexaenoic acid under the catalysis of the high-activity phospholipase A2. The relative content of the 2-DHA-PS in the product synthesized through the method is high, and the defects of an existing synthesizing method are effectively overcome.

The invention discloses a preparation method suitable for industrialized continuous production of enzyme modified concentrated phospholipids. The preparation method comprises the following steps: a, adding phospholipase A1, phospholipase A2, and a 1, 3-lipase aqueous solution into concentrated phospholipids for enzymatic modification reaction; b, pumping the concentrated soybean phospholipids after enzymatic modification into a fixed scraper film evaporator for dehydration by continuous evaporation and enzyme inactivation through a screw pump, so as to enable the moisture to be decreased to 1% or below; c, concentrating, dewatering, cooling the inactivated enzyme modified phospholipids with a cooler, and introducing the cooled enzyme modified phospholipids into a vacuum stirring tank for removing oxygen in the modified phospholipids, so as to obtain the enzyme modified soybean phospholipids with good storage stability. The preparation method is suitable for continuous industrial production, the processing technology is easy and convenient, the production efficiency is remarkably improved, and the produced enzyme modified soybean phospholipids are good in dispersity, hydrophilia and heat resistance in water, quite good in emulsion stability in a low pH value range and hyper-salt concentration, and widely applicable to the industries, such as, food, medicine, daily-use chemicals, pesticides, paint coatings, leather and textile, petrochemical industry and fodder.

The invention relates to a kit for rapidly diagnosing lipoprotein-associated phospholipase A2 and a use method of the kit. The kit comprises a standard substance, a quality control substance, a liposome complex, a magnetic separation reagent, an assay buffer solution and a cleaning solution, wherein the inner part of the liposome complex is coated with N-hydroxysuccinimide ester of tris (bipyridine) ruthenium; an antibody for resisting the lipoprotein-associated phospholipase A2 is connected to the surface of the liposome complex by interaction of streptavidin and biotin; the magnetic separation reagent takes magnetic particulates containing amino terminals as a carrier and is connected with the antibody for resisting the lipoprotein-associated phospholipase A2 through the interaction of the streptavidin and the biotin. Compared with an enzyme linked immunosorbent assay kit produced in a foreign Diadexus company, the kit prepared by the invention has the advantages that sensitivity and precision for analyzing the lipoprotein-associated phospholipase A2 are greatly improved; in addition, the kit disclosed by the invention is simple and convenient to operate, quick and easier to popularize.

The invention discloses a lipoprotein-related phospholipase A2 enzyme-linked immunosorbent assay (ELISA) kit and a preparation method of the lipoprotein-related phospholipase A2 ELISA kit. The lipoprotein-related phospholipase A2 ELISA kit comprises standard substances, quality control substances, a coating carrier, an enzyme marker, a color-substrate solution, an analysis buffer solution and a concentrated cleaning solution. The preparation method of the lipoprotein-related phospholipase A2 ELISA kit includes the first step of preparing the lipoprotein-related phospholipase A2 standard substances and the lipoprotein-related phospholipase A2 quality control substances, the second step of preparing the carrier wrapped by lipoprotein-related phospholipase A2 antibodies, the third step of preparing the enzyme marker of the lipoprotein-related phospholipase A2 antibodies, the fourth step of preparing the analysis buffer solution, the fifth step of preparing the concentrated cleaning solution, and the sixth step of assembling the lipoprotein-related phospholipase A2 ELISA kit. The lipoprotein-related phospholipase A2 ELISA kit can be used for replacing other kits for quantitative detection of lipoprotein-related phospholipase A2, and is low in cost, easy and convenient to operate, high in sensitivity and capable of being widely popularized and used clinically on a large scale.

The invention relates to a strain for generating phospholipase D with high and stable yield by utilizing physical and chemical mutation, belonging to the technical field of microbial mutation. The strain is characterized in that the phospholipase D is prepared by fermenting streptomyces chromofuscus of high yield phospholipase D, which is obtained by taking the streptomyces chromofuscus as an original strain and sequentially carrying out composite mutation of ultraviolet rays and lithium chloride and composite mutation of atmosphere cold plasmas and the lithium chloride on a monospore bacterial suspension thereof. The invention has the advantages that the streptomyces chromofuscus of the high yield phospholipase D is obtained through mutation screening, thereby providing a high yield strain with higher application value to industrial production of the phospholipase D, laying a good foundation for fermentation culture and application of the phospholipase D, and also providing a good and feasible method for microbial mutation breeding. The invention adopts a mutation and screening method, has the advantages of simple operation, high efficiency and the like, and has a practical value on screening the strain with an industrial application value.

The present invention relates to a lipid-based drug delivery system for administration of a lysolipid derivative present in a prodrug from, said prodrug furthermore being a substrate for extracellular phospholipase A2 to the extent that an organic radical can be hydrolytically cleaved off, whereas the aliphatic group of the lysolipid derivative remains substantially unaffected, said system having included therein lipopolymers or glycolipids so as to present hydrophilic chains on the surface of the system. Particularly interesting lipid derivatives are lipids in which the head group is linked to the C-2 position and the organic radical (a drug substance) is covalently attached to the C-3 position of the glycerol moiety. Pharmaceutical compositions comprising the drug delivery system can be used in diagnosis and targeted treatment of various disorders, e.g. cancer, infectious, and inflammatory conditions, etc., i.e. disorders and diseases associated with or resulting from increased levels of extracel lular PLA₂ activity in the diseased tissue.

The invention relates to a phospholipase A2 mutant and a preparation method thereof. The technical scheme is as follows: the preparation method comprises the following steps: carrying out site-specific mutagenesis on the amino acid residues of Glu37 and Asp78 of wild type phospholipase A2 derived from streptomyces coelicolor by utilizing a recombinant DNA technology to obtain a phospholipase A2 gene with higher activity; then expressing the phospholipase A2 gene with higher activity in a bacillus subtilis expression system and a pichia pastoris expression system (which comprises a pichia pastoris free expression system and a pichia pastoris cell surface display system) to obtain a recombinant strain capable of generating high-activity phospholipase A2; after the phospholipase A2 gene with higher activity is expressed, detecting that the enzyme activity of the high-activity phospholipase A2 is enhanced by 13.7% compared with that of the wild type phospholipase A2, wherein the enzyme activity of the phospholipase A2 can be up to 53.5 U/ml after the bacillus subtilis high-activity phospholipase A2 recombinant bacteria is fermented, the enzyme activity of pichia pastoris free-expression high-activity phospholipase A2 recombinant bacteria can be up to 106.4 U/ml, and the enzyme activity of a pichia pastoris cell surface display high-activity phospholipase A2 whole-cell catalyst can be up to 260 U/(g.stem cell).

The invention provides a method for degumming with magnetic immobilized phospholipase A2. The method comprises the steps of adding magnetic immobilized PLA2 (phospholipase A2) into soybean raw oil for degumming, and recycling magnetic immobilized enzyme with a magnetic field after a reaction is finished. For determination of a phosphorus content after the degumming, oil yield and recycling times of immobilized PLA2, the obtained minimum phosphorus content after the degumming is 20ppm, and the relative activity of the immobilized enzyme keeps around 80% after the oil yield is increased by 1.2%, and magnetic immobilized PLA2 is continuously utilized for eight lots. After magnetic immobilized PLA2, phospholipid formation in degumming oil can be increased; separation of an oil phase and a gum phase is improved; formation of free fatty acid in the degumming oil can be controlled; and sterol can be re-esterified with free fatty acid to form sterol ester to become a part of edible oil. Therefore, physical refining is realized, and the yield of the degumming oil can be increased. The high quality degumming oil with a low phosphorus content can be produced, and the degumming oil contains higher-content functional plant sterol ester at the same time.

The invention relates to an anti-phospholipase A2 acceptor antibody IgG kit and a detection method. The kit comprises a micropore board prewrapped with human phospholipase A2 receptor antigens, and the following bottled reagents: an anti-phospholipase A2 acceptor antibody IgG calibrator, a positive control group, a negative control group, a concentrated washing liquid, a sample diluent, a standard enzyme solution, a substrate liquid and a stop solution. According to the detection method of the invention, the contents of anti-phospholipase A2 acceptor antibody IgG in serum and plasma can be quantitatively detected in a non-invasive, low-risk, safe, cheap and precise manner, the positive detection rate of patients suffering from idiopathic membranous nephropathy is high, the detection result is precise and objective, the method is simple and convenient, and the popularization and the application are easy. The anti-phospholipase A2 acceptor antibody IgG kit and the detection method have important significances in diagnosis, treatment guidance and curative effect evaluation of idiopathic membranous nephropathy, and important clinical reference values in treatment scheme making and method selection.

The invention provides a myocardial infarction early-warning colloidal gold kit and a preparation method thereof and relates to a medical detection device. According to the myocardial infarction early-warning colloidal gold kit and the preparation method thereof, the preparation method comprises the following steps of: preparing a colloidal gold mark protein solution of a colloidal gold mark protein related phospholipase A2 monoclonal antibody and detecting a reaction carrier. The preparation method is characterized in that PEG20000 (Poly Ethylene Glycol 20000) with the concentration of 0.24mg/ml, BSA (Bull Serum Albumin) with the concentration of 1.5% (w/w), cane sugar with the concentration of 3% (w/w), mycose with the concentration of 0.5% (w/w), casein with the concentration of 1% (w/w) and sodium dodecyl sulfate SDS are added into the colloidal gold mark protein solution. According to the improvement of the preparation method disclosed by the invention, two kinds of colloidal gold detection kits capable of rapidly detecting whole-blood lipoprotein related phospholipase A2 (Lp-PLA2) are provided; and the invention further provides the colloidal gold mark protein solution for optimizing the property of the kit.

The invention provides a lipoprotein phospholipase A2 detection kit which comprises reagent R1 and reagent R2, the reagent comprises a buffer solution, a stabilizing agent, electrolyte, a surface active agent and preservative; the reagent R2 comprises polystyrene latex particles coupled to an antibody resisting to the lipoprotein phospholipase A2, a buffer solution, a stabilizing agent, a surface active agent and preservative. A directional coupling method is adopted for the polystyrene latex particles sensitized by the antibody resisting to the lipoprotein phospholipase A2, and the preparation steps include activation of the polystyrene latex particles; oxidation of the antibody resisting to the lipoprotein phospholipase A2; coupling of the antibody resisting to the lipoprotein phospholipase A2 to the polystyrene latex particles. The kit is high in detection sensitivity, wide in detection range, good in accuracy and high in interference resistance.

A modified phospholipase D having an ability to synthesize phosphatidylinositol which contains: (A) an amino acid sequence derived from the amino acid sequence represented by SEQ ID NO:2 by substitution of at least one of the amino acid residues at the 187-, 191- and 385-positions by other amino acid residue(s); or (B) an amino acid sequence of a polypeptide derived from the amino acid sequence as described in the above (A) by substitution, deletion, insertion and/or addition of at least one amino acid residue at a position different from the 187-, 191- and 385-positions and having an ability to synthesize phosphatidylinositol. By using this modified phospholipase D, phosphatidylinositol can be efficiently produced from a phospholipid. It is also intended to provide a method of screening a phospholipase D having an ability to synthesize an acylglycerophospholipid having glycol group.

The invention discloses a strain of lactobacillus casei and application of the lactobacillus casei to feed. The Llactobacillus casei L-1 is obtained through screening and separation from soil, the preservation number is CCTCC (China Center for Type Culture Collection) M2012150. The strain can be fermented to produce phospholipase A2, the enzyme yield is high, the strain is applied to the feed field, oil crops containing plant phospholipid or processing byproducts of oil crops are used as fermentation substrates, ammonium sulfate, sodium chloride and corn starch are used as energy supply bodies, the strain is used as a fermentation strain, and feed additives are prepared through solid fermentation. The plant phospholipid conversion rate of the feed additives is high, good antioxidation and antibacterial capability is realized, and no toxic or side effect is caused on animals. Feeding experiments show that when the feed additives are added into a pig feed premix compound, the feed intake and the daily weight increase of pigs can be improved, the feed-weight ratio is reduced, and a growth promotion effect is realized.

The invention provides a method for immobilizing phospholipase A2 by using a sodium alginate-chitosan composite carrier as material through a physical embedding method. According to the invention, the influences of different immobilization conditions such as sodium alginate concentration, chitosan concentration, calcium ion concentration, glutaraldehyde concentration, cross-linking time and the like on the immobilization efficiency and catalysis effect of the phospholipase A2 are researched; and a response surface analysis method is used for optimizing the researched factors. Through immobilizing an SEM (Scanning Electron Microscope) image of the phospholipase A2, the advantage of fixing the phospholipase A2 by using sodium alginate-chitosan as a carrier is further explained. According to the embedding characteristic of the composite carrier in the invention, the phospholipase A2 is immobilized, so that the activity and stability of enzyme in a reaction system are improved, the activity and selectivity of the enzyme are adjusted and controlled and then benefit is brought to the recovery of enzyme and the production of products; and the activity recovery rate of the finally obtained immobilized enzyme is 74.8% and important guiding significance is provided to the enzymatic degumming process in oil and fat refining.

The invention belongs to the technical field of biomedicine test, and relates to a testing kit of Lp-PLA2 testing kit and its detecting method. The testing kit comprises reagent R2, reagents R2A and R2B, wherein the reagent R1 is composed of buffer solution, electrolyte, inhibitor, and preservative, and its pH value is 7.0-7.5; the reagent R2A is composed of buffer solution, inhibitor, protectionagent, and preservative, and its pH value is 2.6-6.0; the reagent R2B is composed of buffer solution, Lp-PLA2 substrate, co-solvent, and preservative, and its pH value is 7.0-7.5. the sample and the reagent R1 are evenly mixed and then hatched for 3-5 minutes at 35-40 DEG C; then the reagent R2 prepared by the reagents R2A and R2B is added, and hatched for 60 seconds at 35-40 DEG C; the light absorbency A1 is read; the reagent is hatched for 180 seconds, the light absorbency A2 is read; the differential value of A2 and A2 is calculated; the activity of Lp-PLA2 in the sample is calculated through the calibration curve.

The invention belongs to the technical field of plant molecular breeding and biology, and discloses application of a PLD (phospholipase D) epsilon gene in increasing nitrogen nutrition effective utilization and seed yield of crops. The nucleotide sequence of the PLD epsilon gene is shown as SEQ ID NO:1, or has a similarity more than or equal to 60 percent as the nucleotide sequence shown as SEQ ID NO:1. The invention also discloses a method for increasing nitrogen nutrition effective utilization and seed yield of rape. Tests prove that growth and occurrence of lateral roots of rape can be promoted due to overexpression of AtPLD epsilon either in nitrogen starvation or nitrogen sufficiency condition, increase of rape leaf tissue cell growth and biological yield can be reinforced, absorption transportation and assimilation metabolism processes of rape plants can be promoted, which are particularly indicated in that the expression quantities of genes NTR1.1 and NTR2.1 related to overexpressed plant N transportation are remarkably higher than that of a wild type, and the nitrogen absorption rate is remarkably higher than that of the wild type.

The invention relates to a method for predicting the inhibiting concentration of an inhibitor of cytosolic phospholipase A2alpha based on a support vector machine, and belongs to the cross field of chemometrics and cheminformatics. By establishing a relational model between the molecular structure and the inhibiting concentration of one inhibitor through the support vector machine, the inhibiting concentration corresponding to the inhibitor can be predicted as long as the molecular structure of the inhibitor is known. The prediction accuracy rate of the method can achieve over 90 percent, the model is stable, the risk of inhibitor development in a later period can be reduced, and the cost of research and development can be reduced.

The present invention relates to a method for preparing hydrolyzed phospholipids by utilizing ultrasonic treatment and enzymic hydrolysis modification, belonging to the field of phospholipids modification technology. Said invention uses soybean hydrous oil foot as raw material, uses the phosphatidase A2 as enzyme for enzymic hydrolysis and adopts ultrasonic treatment method to make the soybean hydrous oil foot be modified in water phase so as to prepare the hydrolyzed phospholipids.