56 results about "Multifunctional Enzymes" patented technology

The present invention provides a no-whey process for preparing a low-fat hard cheese product with completed capture of whey proteins using a modified milk protein concentrate. The modified milk protein concentrate comprises about 82 to about 96 percent milk protein concentrate and about 4 to about 18 percent multifunctional enzyme, wherein the multifunctional enzyme modifies the function of dairy proteins so that the completed capture of whey proteins into a cheese product will not interfere with the alignment of casein molecules into the fibers structure required for a good texture of hard cheeses such as mozzarella. The present invention solves problems associated with the whey generated from conventional making cheese plant and creates a new way of making low-fat hard cheese products.

The invention provides a method for preparing ferulic acid by utilizing corn barn hydrolyzed by a multifunctional enzyme. The method comprises the following steps of: adding the corn bran into NaOH and soaking darkly, sterilizing at high pressure, adding the multifunctional enzyme with peptidase and esterase activities for hydrolyzing ester bonds and peptide bonds on the cell wall, centrifugallytaking supernatant, treating the supernatant by adopting a column chromatography and eluting, concentrating eluent, extracting by adopting ethyl acetate and rotatably evaporating to obtain the ferulic acid. The ferulic acid is produced by utilizing the corn bran as a by-product in the process of grain processing or fuel ethanol production and the hydrolysis of the multifunctional enzyme with peptidase and esterase activities, therefore, the invention has simple process, time and energy saving and increased additional value of the corn by-products.

The invention relates to an analyzing method for detecting activity of soil xylanase, which comprises the following steps: firstly, weighting n sieved air dry soil samples into n thick test tubes, adding acetic acid buffer solution into each test tube, oscillating by a vortex oscillator, getting soil suspension into 96 micropore plates under the oscillation condition, adding 4-MUB-7-Beta-D-xyloside substrate solution into n-1 holes, and adding water with equal quantity into the other one hole so as to be used as non-substrate contrast, adding substrate solution with equal quantity and water with equal quantity into the (n+1)th hole so as to be used as soli-free contrast, oscillating and culturing under constant temperature; secondly, adding NaOH into the micropore plates to terminate the reaction after the culture is finished; thirdly, performing the fluorimetric determination to resultant of reaction by a multifunctional microplate reader; and fourthly, calculating the activity of the xylanase. Compared with the traditional method, the invention shortens the culture time, omits the operation procedures of filtration, and the like, and simplifies the operation steps; meanwhile, the determination data of fluorescent materials in the micropore plate can be obtained within 15s through the multifunctional microplate reader, huge samples are allowed to be simultaneously determined; and moreover, the invention has high accuracy and easy operation, stable and reliable result and good reproduction quality.

The present invention relates to a method for high-sensitivity detection of fumonisin B1 through a hydrogel silica photon crystal microsphere chemiluminescence method. According to the present invention, the gel is subjected to ultraviolet polymerization in the silica photon crystal microsphere gap so as to reduce the porous structure inside the microsphere, reduce the non-specific biological reaction background signal, and improve the detection sensitivity and the detection accuracy; and the fumonisin B1 probe carrier on the microsphere surface is used to construct the competitive fumonisin B1 chemiluminescent immunodetection system on the microsphere surface, the chemiluminescent signal is detected by using the multifunctional enzyme-labeling measuring instrument, the detection limit of the method on the fumonisin B1 is 0.0001 ng/mL, the linear detection range is 0.0001-1 ng/mL, and the method has advantages of high detection sensitivity and low background signal.

The invention relates to a method for rapidly and efficiently screening a rhamnolipid producing bacteria nutrition system. The method comprises the following steps: synthesizing a primer of rhamnolipid producing gene rhlAB, performing gene amplification by using PCR, synthesizing rhlAB gene fragments, connecting the rhlAB gene fragment to the position in front of lux luminescence gene on pMS402 plasmid by utilizing gene engineering shearing and splicing technology, then transferring the recombinant plasmid into pseudomonas aeruginosa DN1, coating the recombinant strain onto an LB solid plate containing 200mg/L-300mg/LTmp, screening out positive clone, naming the strain as rhlAB-lux strain, picking out positive monoclonal colonies to be enriched on the solid LB culture medium, refrigeratingat at 4DEG C for later use; taking out the refrigerated rhlAB-lux strain, and enriching the thallus overnight; and screening a nutrient optimized system. According to the method, the luminescence value can be determined by a multifunctional microplate reader, and the method has the characteristics that the flux is high, and the screening process is simplified and rapid.

The invention relates to an enzyme preparation method and a product thereof, wherein natural fruits are used as raw materials, preferably 6-9 parts by mass of apple, 5-8 parts by mass of pear, 3-4 parts by mass of lemon, 4-7 parts by mass of grapefruit, 7-10 parts by mass of carrot, 3-4 parts by mass of orange, 2-3 parts by mass of hawthorn, and 2-3 parts by mass of ficus carica linn are mixed andcrushed to prepare a liquid fruit juice, 5-8 parts by mass of the liquid fruit juice, 3-5 parts by mass of rock candy, 0.01-0.07 part by mass of lactic acid bacteria and 0.01-0.09 part by mass of yeast are uniformly mixed, fermenting is performed for 3-6 months at a room temperature in a closed and dark environment, the supernatant is extracted, and filtering, disinfecting and filling are performed to obtain the enzyme finished product or the product thereof, wherein the filling is bottling or boxing. According to the present invention, the enzyme and the environmental protection multifunctional enzyme soap have advantages of no pigment addition during the manufacturing, mild property and no irritating, and are suitable for a variety of people; and the obtained product is the pure naturalenzyme beverage, and can further be used for washing, shampooing, face washing, bathing, tooth brushing and the like.

According to the invention, the change of the level of 3',5'-cyclic adenosine monophosphate (cAMP) is taken as a detection index and a cAMP-depending type protein kinase A active analysis method is used for determining the biological activity of liraglutide. Rat insulinoma cells RIN-m5F are cultivated and a diluted liraglutide comparison product solution and a diluted test solution are added; a kit is used for detecting the change of the level of the cAMP in stimulated cells; a super-sensitive chemiluminiscence detection module of a multifunctional enzyme linked immunosorbent assay is used for reading a relative chemiluminiscence unit (RLU) and statistical software is used for fitting experimental data; the biological activity of a product to be detected is calculated by a formula. The method can be used for simply, conveniently, rapidly and accurately detecting the change of the biological activity of the liraglutide; the accuracy and the repeatability accord with the requirements of a biological activity determination method of biological products.

The invention relates to a method for detection of P-glycoprotein (P-gp) mediated rhodamine 123 (Rh123) transport in a 3D type organ and an application thereof in screening of P-gp inhibitors. The method comprises the steps: firstly, sorting small intestinal crypts of C57BL/6 mice, inoculating a petri dish containing a matrix gel, culturing in an Advanced DMEM/F12 culture medium, and forming the 3D type organ; and then carrying out qualitative and quantitative detection of P-gp mediated Rh123 transport and influence of the P-gp inhibitor verapamil on Rh123 transport by using the 3D type organ model. The method particularly comprises the steps of respectively co-incubating the 3D type organ with (1) the P-gp substrate Rh123 and with (2) the Rh123 and the verapamil, releasing the Rh123 in the 3D type organ by using an ultrasonic crushing method, and finally detecting the concentration of the Rh123 on a multifunctional microplate reader. The method has the advantages of being simple, rapid, and high in sensitivity, can be combined with the 3D type organ model for carrying out research of the P-gp mediated drug transport, and also can perform research of in-vitro screening of the P-gp inhibitors.

The invention provides a preparation method of dietary fiber by adopting corn bran hydrolyzed by a multifunctional enzyme. The preparation method comprises the following steps of: adding the corn bran into NaOH and soaking darkly, sterilizing at high pressure, adding the multifunctional enzyme with peptidase and esterase activities for hydrolyzing ester bonds and peptide bonds on the cell wall, centrifugally taking precipitate, decoloring and rinsing, drying at the temperature below 60 DEG C, crushing and screening by adopting a 80-100 mesh sieve to obtain the dietary fiber. The dietary fiber which has the expansion force up to 10-15ml/g and the water-holding capacity of 9-11g/g and is superior to other methods is prepared by utilizing the corn bran as a by-product in the process of grain processing or fuel ethanol production and the hydrolysis of the multifunctional enzyme with peptidase and esterase activities. The invention utilizes the enzyme, has simple process, saves time and energy and increases the additional value of the corn by-products.

The invention discloses a novel multifunctional enzyme gene HG32 and application, the enzyme is derived from Chinese rhizomyidae enteric microorganisms, can be compatible with an eukaryotic cell expression system, expresses cellulase with high biological activity, and can be applied to the fields of livestock and poultry feed enzyme preparations, transgenic animal preparation, food and medicine industries and the like; the invention solves the problems that plant cellulose is difficult to degrade, the feed nutrition utilization rate is low and the like are solved, and pollution of nitrogen, organic matter and the like in animal excrement. According to the invention, the application comprises the following steps: sequencing a bamboo rat intestinal microbe macro transcriptome, and splicing the sequences; performing homology comparison with the existing glycoside hydrolase family on NCBI, and screening high-expression suspected cellulase genes with homology higher than 50%; predicting the signal peptide, and replacing the original signal peptide with the pig parotid gland secreted protein signal peptide; optimizing according to pig codon preference, and cloning to an eukaryotic expression vector; transfecting into pig kidney epithelial cells through lipidosome for expression; collecting a cell culture supernatant; and carrying out enzyme activity analysis and comparison.

By adopting methods of fluorescent quantitation PCR, fluorescent quantitation RT-PCR, HBV DNA quantitation, HBsAg quantitation, HBeAg quantitation, cccDNA quantitation, Northern blot, Southern blot, Western blot, immunohistochemistry, and the like in the research and using the maintenance dose within the treating dose range for a long time (30-60 days), supernatant HBV DNA HBsAg HBeAg cultured by HepG-2.2.15 cells can completely disappear, HBsAg, HBeAg, HBcAg, and the like in the cells are completely turned to be negative, HBV DNA is in a high inhibited state, HBV cccDNA is completely negative, fluorescent quantitation RT-PCR detection finds that the mRNA for expressing HBsAg and HBcAg antigens in the cells is completely negative, and in addition, the curative effect of the atabrine is 30 times that of lamivudine as a first-line drug for resisting HBV in current clinic. In order to illustrate the action mechanism of the atabrine and the pyronaridine as a drug belonging to the same kind with the atabrine, an HBV genome is divided into 3 segments which are respectively inserted into an Xb1 position on a luciferase report carrier PGL3, a multifunctional microplate reader detects and finds that the light production value of the luciferase is remarkably reduced, which shows that the molecules of the atabrine and the pyronaridine as a drug belonging to the same kind with the atabrine can be nonspecifically combined with the HBV DNA in the cells, thereby inhibiting the copying of the virus and ensuring that the HBV DNA content of the virus in cells copied by the filial generation is reduced till to disappear. The patent requires protecting the application of 3 linked benzyl structures (named as ethyleneimine) and radicals such as CH3O-,-NH-, CL-, and the like for resisting HBV virus in clinic.

The invention discloses a method for accurately and quantitatively detecting an autophagy flux. The method comprises the following steps: (1) constructing an EGFP/LC3HIBIT co-expression vector, and respectively applying EGFP and LC3HIBIT to independent promoters without mutual influence; (2) obtaining co-expressed cells of EGFP and LC3-HIBIT proteins by a transient transfection or monoclonal screening method; (3) after screening the monoclonal stably transfected cells, carrying out various genetic operations related to autophagy on the basis of the cells, and screening autophagy related drugsand the like; and (4) setting programs on a multifunctional microplate reader to respectively measure luminescence and fluorescence readings, and calibrating a HIBIT reading caused by cell differencesby taking the EGFP as an internal reference in the experiment so as to accurately reflect the autophagy degree of the cells.

The invention discloses a continuous endo-cellulase as well as a coding gene and application thereof, and belongs to the technical field of bioengineering. The nucleotide sequence of the gene is shown as SEQ ID NO.3. The amino acid sequence of the continuous endo-cellulase coded by the gene is shown as SEQ ID NO.4. The continuous endo-cellulase SmCel5A is a multifunctional enzyme of a single structural domain, the molecular weight of the continuous endo-cellulase SmCel5A is less than 40 kD, heterologous expression is easy to conduct, active protein with the activity of endo-cellulase and excision-cellulase enzyme is obtained, and the problems that the growth cycle of sporocytophaga is long, enzyme components are complex, and the cellulase is not easy to directly separate and purify are effectively solved. The enzyme has wide application potential in the industries of biological energy, feed and the like.

The invention discloses a protein for catalytic synthesis of amyrin, and a preparation method and an application for the protein, and relates to the technical field of biomolecules. The protein has anamino acid sequence as shown in SEQ ID No. 2, or has a sequence which has 90% or above of sequence homology with the sequence as shown in the SEQ ID No. 2 and has the same protein function as the sequence as shown in the SEQ ID No. 2. According to the protein, 2,3-squalene oxide is used as a substrate; alpha-amyrin, beta-amyrin and delta-amyrin can be catalytically synthesized simultaneously; thealpha-amyrin is mainly synthesized; and a new direction is provided for research of multifunctional OSC enzymes.