32 results about "6-APA" patented technology

The invention relates to an immobilized enzyme biocatalyst and the manufacturing method and application. The immobilized enzyme biocatalyst regards tubular hollow silica dioxide dielectric hole material as a carrier external surface, inner surface and micropore in the wall of which are fixed biological enzyme molecule. The method includes physical adsorption cast investment or chemical coupling/ crosslinking method; the biocatalyst is provided with good enzyme dispersibility and high carrying quantity, high recovery ratio of enzymatic activity, low enzyme flow rate. The invention can fix penicillin acylating enzyme, glucose oxidase, peroxidase, cytase and so on, which also can be used for removing sugar in protein, removing sugar in total egg, removing oxygen in food , microbiological sensing device, antibiosis and disinfection reaction, wherein immobilized penicillin acylating enzyme biocatalyst can be used to hydrolyze penicillin potassium and manufacture 6-APA.

The invention discloses a preparation method for 6-amino penicillanic acid, which comprises the following steps: a, performing ultrafiltration membrane separation and nanofiltration membrane concentration on a penicillin fermentation liquor to obtain a concentrated filter liquor; b, placing the concentrated filter liquor into a reaction tank, adding an immobilized penicillin acylase 4MU/m<3> concentrated filter liquor and performing conversion reaction to obtain a 6-amino penicillanic acid conversion solution; c, performing actived carbon decoloration and filtering on the conversion solution to obtain a 6-amino penicillanic acid filter liquor; and d, adding seed grain into the 6-amino penicillanic acid filter liquor obtained through the procedures in the step c, growing the grain, crystallizing, filtering, washing and drying. The preparation method has the advantages of simple process flow, easiness for operation, safety, environmental protection, and capabilities of effectively improving the yield of 6-APA, reducing the production cost and improving the labor productivity.

The invention discloses a process for preparing straight-through 6-aminopenicillanic acid, which comprises the following steps: a, filtering and acidizing penicillin fermentation solution, extracting the penicillin fermentation solution by using butanol, and concentrating and decoloring the extract to obtain butyl ester extracting solution of penicillin; b, back extracting the butyl ester extracting solution of the penicillin by using alkali solution to obtain brine solution of penicillin (heavy phase or RB for short); c, continuously injecting the brine solution of the penicillin into a degreasing tower in a vacuum pressure reduction state to convert the butyl ester into a gas phase from the brine solution of the penicillin, discharging the degreased brine solution of the penicillin out of a pressure reduction system from the bottom of the tower to a storage tank with a cooling device, and cooling the degreased brine solution of the penicillin for later use; and d, performing enzymatic conversion on the degreased brine solution of the penicillin, then adding 6-APA crystal seeds into the solution, growing the crystals, crystallizing the solution, and drying the crystals.

The invention provides a mutant of penicillin G acylase (PGA) and a preparation method and application of the mutant. The non-rationally and semi-rationally designed enzyme engineering reconstruction technology is adopted for mutation of penicillin G acylase obtained from Escherichia coli ATCC 11105, so that a PGA mutant with higher reactivity, higher reaction rate, better conversion rate, stronger in penicillihe concentration tolerance, less substrate residue, and higher in phenyl acetic acid concentration tolerance; meanwhile, the mutant is subjected to recombinant expression, bacteria strain construction, fermenting cultivation, immobilization and application to prepare 6-amino-penicillanic acid (6-APA). The activity of the PGA-6 mutant prepared by the invention is increased by 102 times, the substrate penicillihe concentration tolerance is increased to 30%, and the phenyl acetic acid concentration tolerance is increased to 20 mmol/L; meanwhile, the immobilized PGA-6 mutant is used to decompose penicillihe with a concentration of 25% under the condition of pH 8.0 and 25 DEG C so as to prepare 6-APA, and the reaction time is shortened to 55 minutes, the substrate conversion rate is 98% or above, and after being used for 600 batches and above, the activity is not lost obviously, therefore, good operation stability is achieved.

The invention discloses a process for preparing 6-aminopenicillanic acid, which comprises the following steps: (1) filtering the penicillin fermentation liquid, acidifying, extracting and concentrating with butanol, decoloring to obtain butyl extract of penicillin, (2) subjecting butyl extract of penicillin to back extraction with alkaline solvent, obtaining aqueous solution of penicillin salts, (3) degreasing the aqueous solution of penicillin salts, (4) loading the degreased aqueous solution of penicillin salts into penicillin acylated enzyme retort for enzyme conversion, charging 6-APA seeds, cultivating quartz, crystallizing and drying.

The invention provides a penicillin G acylase mutant for synthesis and an application thereof in the preparation of amoxicillin. Penicillin G acylase of Achromobacter xylosoxidans origin is mutated by computer aided design in connection with semi-rational design of site-saturation mutagenesis technique and enzyme engineering modification of orthogenesis, thus acquiring the penicillin G acylase mutant lower in hydrolytic activity, better in synthetic activity, higher in synthesis-hydrolysis ratio (S/H), higher in acid resistance and better in stability, and amoxicillin can be catalytically synthesized more effectively and quickly. Immobilized enzyme hydrolytic activity of the mutant SPGA-4 obtained is decreased by 8.7 times, synthetic activity is increased by 5.6 times, the S/H ratio is increased by 8 times, the activity remains at 79% for 60 min under the condition of pH 2.0, amoxicillin is catalytically synthesized by a solid method at 10 DEG C or 20 DEG C, substrates 6-APA and D-HPM are directly charged in a solid form without dissolving, reaction pH need not be controlled, substrate conversion rate is higher than 99%, continuous use is available in more than 300 batches, and good operational stability is given.

The present invention discloses a kind of immobilized penicillin amidase carrier and its preparation process. The immobilized pencillin amidase carrier is prepared with surfactant as dispersant and vinyl compound as monomer and through crosslinking polymerization in reverse suspension technology. The carrier immobilized pencillin amidase has an apparat enzyme activity in preparing 6-APA with benzyl pencillin potassium as high as 125 IU/g. The polymer bead carrier containing epoxy group of the present invention has high apparent enzyme activity, simple technological process, easy operation andcheap material and may be used in industrial production.

A synthetic method for the general artificial antigen of a beta-lactam medicine belongs to the biochemical technical field. The invention adopts 6-amino penicillanic acid as hapten, couples the hapten with the carrier protein of bovine serum albumin BSA through an N-(m-maleimide group benzene methanoyl) succinimides MBS method, and determines the coupling ratio of the coupled matter through a gel electrophoresis method. The method successfully synthesizes the 6-APA general artificial antigen with a simple and effective synthesis process. The method which can completely be used for immune analysis provides necessary artificial antigen for future study and meets the requirements of domestic study on the artificial antigen.

The invention relates to a novel method for preparing broad-spectrum penicillin antibiotic ticarcillin sodium. The method is characterized in that the method comprises the following steps of: (1) directly carrying out chlorination on 3-thiophenemalonic acid used as raw material and thionyl chloride in the presence of a catalyst, without protecting carboxyl, and chloridizing one carboxyl to obtian a compound (II); (2) carrying out condensation reaction with 6-aminopenicilanic acid (6-APA), controlling the reaction temperature and pH value, dripping the compound (II) and sodium bicarbonate solution, dispensing liquid after reaction, adjusting pH value to wash, adjusting pH value again to 2.0, extracting reaction solution, and drying to obtain a compound (IV); using sodium iso-octoate to salify, and refining crude products, to obtian the ticarcillin sodium salt.

The invention discloses a recycling process of amoxicillin in enzymatically synthesized amoxicillin mother liquor. The process comprises the following steps: concentrating the amoxicillin mother liquor through the adoption of nanofiltration membrane high-power concentration equipment, feeding the concentrated liquor in a reaction tank, sufficiently reacting the 6-APA in the concentrated liquor with methyl D-(-)-4-hydroxy-phenylglycinate under the effect of penicillin G acylase so as to generate the amoxicillin. The recycling process disclosed by the invention has the advantages that through the nanofiltration concentration, not only the amoxicillin in the mother liquor is recycled, but also the 6-APA contained in the mother liquor participates the enzymatic synthesis reaction to be translated into the amoxicillin, the aim of extremely recycling the amoxicillin in the mother liquor is achieved, and the recycling process has more economy.

The present invention discloses the application of cloud point system in enzymati hydrolysis of penicillin to prepare 6-amino cefo alkanoic acid. By means of two phase distributing bioreactor technology with cloud point system, penicillin acylating enzyme is used in enzymati hydrolysis of penicillin G to prepare semi-synthesized 6-amino cefo alkanoic acid as the key intermediate for beta-lactam antibiotic. The process has direct extraction of penicillin from the fermented liquid without need of penicillin crystallization, direct extraction of phenylacetic acid in hydrolysis without needing addition acid or alkali to control pH value, and less produced wastes.

The invention provides a penicillin V acylase mutant, an encoding array, a recombinant expression vector, a genetically engineered bacterium and application. Penicillin V acylase derived from Bacillussphaericus is mutated, the specific activity of the obtained mutant PVA-3 is improved by nine times, the immobilization activity is improved to 820U/g from 92U/g, and after being incubated for one hour under a condition of 30% penicillin V sylvite, the mutant has activity residue of 89.1%; the reaction time that the immobilized mutant PVA-3 is adopted to prepare 6-APA (6-Aminopenicillanic Acid) by splitting 25% penicillin V under a condition that the pH value is 6.0 at 25 DEG C is shortened to 60 minutes from 180 minutes, a substrate conversion rate is increased to 99.5% or greater, the activity is not greatly lost after continuous use for 600 times or greater, and good operation stability can be achieved. By adopting the mutated PVA-3, the production cost can be greatly reduced, the production efficiency can be improved, and the mutant is applicable to industrial application.

Ampicillin is produced in a batch process by enzymatic acylation of 6-aminopenicillanic acid (6-APA) with the aid of phenylglycine derivative such as D-phenylglycine amide. High conversions of phenylglycine derivative may be achieved by having the total concentration in the reaction mixture of 6-APA and ampicillin greater than 250 mM and the molar ration of total quantity of phenylglycine derivative to total quantity of 6-APA less than 2.5. Higher yields of ampicillin may be achieved when the amount of dissolved 6-APA is kept low, e.g. below 300 mM.

The invention provides a preparation method of 6-APA (6-amino-penicillanic acid). 6-APA is prepared from a potassium carbonate reversed-extraction liquid of benzylpenicillin potassium directly, a molecular formula of 6-APA is C8H12N2O3S, and the molecular weight is 216.26. With the adoption of the process, a crystallization step can be omitted, and crystallization yield loss is avoided; extracting and washing processes of a penicillin solvent are optimized in a technical process; with the adoption of the advanced preparation process, the preparation method is higher in yield and lower in cost.

The present invention discloses isolation of Penicillin Acylase (PA) from Achromobacter sp CCM 4824 expressed in recombinant strain E. coli BL21 CCM 7394 bearing the recombinant plasmid pKXIP1 and processing of PA into biocatalyst useful for the industrial synthesis of antibiotics. More particularly the invention discloses a synthesis of semi-synthetic β-lactam antibiotics in the reaction mixture consisting of activated acyl-donor (D-p-hydroxyphenylglycine methyl ester or amide for Amoxicillin and Cefadroxil; D-phenylglycine methyl ester or amide for Ampicillin and Cephalexin) and nucleophile (6-APA or 7-ADCA) catalyzed by PA obtained from recombinant E. coli BL21 CCM 7394 as the biocatalyst.