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Beta-lactamase protective agent and preparation method

A technology of lactamase and protective agent, which is applied in the field of medicine and biology, can solve the problems of increasing the amount of enzyme, large loss of activity, and inability to purchase and use enzyme plates, so as to reduce the cost of use, reduce the loss of enzyme activity, and maintain relatively stable effect

Inactive Publication Date: 2018-12-07
广州穗珩生物技术有限公司 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The enzyme added to the enzyme plate medium used for the clean environment monitoring of the production of β-lactam antibiotics in pharmaceutical factories is β-lactamase, and the activity of this enzyme is greatly lost after the enzyme plate medium is irradiated and sterilized Therefore, it is necessary to double the amount of enzyme in the production of enzyme-added plates, which makes it difficult for pharmaceutical users to bear the cost of using enzyme-added plates, and even cannot purchase and use qualified enzyme-added plates.

Method used

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  • Beta-lactamase protective agent and preparation method
  • Beta-lactamase protective agent and preparation method
  • Beta-lactamase protective agent and preparation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Example 1 Prepare a φ90mm TSAc sterile plate (200,000 units / 20g.dish).

[0019] (Enzyme raw material: Hangzhou XX Biotechnology Co., Ltd., cephalosporin enzyme batch number 7041630D, the enzyme activity unit measured by the Chinese Pharmacopoeia method is 10.098 million units / g.)

[0020] 1. Prepare 6000ml of TSA solution, after sterilization, keep it warm at 60°C for later use.

[0021] 2. A β-lactamase protective agent and its preparation method, including Dextran20 0.1mg / wu, trehalose 0.5mg / wu, sorbitol D 0.1mg / wu, L- Histidine 0.2mg / wu, L-arginine 0.2mg / wu, L-ascorbyl phosphate sodium salt 0.01mg / wu, Tween 20 0.02mg / wu, ZnSO 4 0.002mg / wu.

[0022] The β-lactamase protective agent and its preparation method comprise the following steps:

[0023] (1) Prepare 6000ml of TSA solution, after sterilization, keep warm at 60°C for later use.

[0024] (2) Accurately weigh the required amount of Dextran20 0.5g, trehalose 2.5g, sorbitol D 0.5g, L-histidine 1g, L-arginine 1...

Embodiment 2

[0030] Example 2 Preparation of φ90mmTSAc sterile plate (200,000 units / 20g.dish)

[0031] (Enzyme raw material: Hangzhou XX Biotechnology Co., Ltd., cephalosporin enzyme batch number 7041630D, the enzyme activity unit measured by the Chinese Pharmacopoeia method is 10.098 million units / g.)

[0032] 1. Prepare 6000ml of TSA solution, after sterilization, keep it warm at 60°C for later use.

[0033] 2. A β-lactamase protective agent and its preparation method, including Dextran20 0.5mg / wu, trehalose 2mg / wu, sorbitol D 0.5mg / wu in terms of enzyme activity units (ten thousand units, wu), L-group Amino acid 1mg / wu, L-arginine 1mg / wu, L-ascorbyl phosphate sodium salt 0.05mg / wu, Tween 20 0.05mg / wu, ZnSO 4 0.006mg / wu.

[0034] The β-lactamase protective agent and its preparation method comprise the following steps:

[0035] (1) Prepare 6000ml of TSA solution, after sterilization, keep warm at 60°C for later use.

[0036] (2) Accurately weigh the required amount of Dextran20 2.5g,...

Embodiment 3

[0042] Embodiment 3 prepares phi 90mmTSAm aseptic petri dish (200,000 units / 20g.dish)

[0043] (Enzyme raw material: Hangzhou XX Biotechnology Co., Ltd., metallo-β-lactamase batch number: 6021639D, the enzyme activity unit measured by the Chinese Pharmacopoeia method is 7.302 million units / g.)

[0044] 1. Prepare 6000ml of TSA solution, after sterilization, keep it warm at 60°C for later use.

[0045] 2. A β-lactamase protective agent and preparation method, including Dextran20 as 0.5mg / wu, trehalose 1.5mg / wu, sorbitol D 0.2mg / wu in terms of enzyme activity units (ten thousand units, wu), L -Histidine 0.6mg / wu, L-Arginine 0.4mg / wu, L-Ascorbyl Phosphate Sodium Salt 0.03mg / wu, Tween 20 0.04mg / wu, ZnSO 4 0.004mg / wu.

[0046] The β-lactamase protective agent and its preparation method comprise the following steps:

[0047] (1) Prepare 5000ml of TSA solution, after sterilization, keep warm at 60°C for later use.

[0048] (2) Accurately weigh the required amount of Dextran20 2....

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Abstract

The invention discloses a beta-lactamase protective agent. The beta-lactamase protective agent is prepared from the following components by enzyme activity unit (wu): 0.1-0.6mg / wu Dextran 20, 0.5-2mg / wu trehalose, 0.1-0.6mg / wu sorbitol D, 0.2-1mg / wu L-histidine, 0.2-1mg / wu L-arginine, 0.01-0.06mg / wu L-sodium ascorbate phosphate, 0.02-0.06mg / wu Tween 20 and 0.002-0.006mg / wu ZnSO4. The components are dissolved in a defined amount of PH7.0 citrate buffer solution. In use, 1wu beta-lactamase is added. In the protective agent disclosed by the invention, polysaccharides, alcohols and enzyme form a polyhydroxy hydrogen bond structure, for substituting water molecules and maintaining the space structure stability of enzymes; amino acids can protect the active centers of serine, histidine and cysteine of enzymes; glycerinum is used as a common water-retaining agent; Zn2+ is an activating agent of the enzymes; Tween 20 is a nonionic surfactant and is generally used as an enzymatic activity protective agent for enzymatic reaction. Through verification for enzyme activity stability of enzyme-containing products, the protective agent disclosed by the invention is capable of greatly reducing theloss of enzyme activity and maintaining the relative stability of enzyme activity.

Description

technical field [0001] The invention relates to the fields of medicine and biotechnology, in particular to a β-lactamase protective agent and a preparation method. Background technique [0002] β-lactamase is an inactivating enzyme produced by bacteria that catalyzes the hydrolysis of the β-lactam ring of β-lactam antibiotics. Mainly divided into four categories: (1) penicillinase, the hydrolysis substrate is mainly penicillin antibiotics; (2) cephalosporinase, the hydrolysis substrate is mainly cephalosporin antibiotics; (3) metallo-β-lactamases, The hydrolysis substrate is mainly carbapenem antibiotics; (4) Oxacillin-like penicillinase (OXA type), the hydrolysis substrate is mainly oxacillin, cloxacillin, methicillin, dicloxacillin and other penicillin-resistant enzymes antibiotic. Most of the active centers of β-lactamases are mostly serine (serine active enzyme), while the active centers of metallo-lactamases are mostly histidine and cysteine, which require Zn 2+ or M...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/96
CPCC12N9/86C12N9/96C12Y305/02006
Inventor 不公告发明人
Owner 广州穗珩生物技术有限公司
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