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Enzymatic Synthesis of Amoxicillin via a One-pot Enzymatic Hydrolysis and Condensation Cascade Process in the Presence of Organic Co-solvents

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Abstract

A cascade reaction combining the enzymatic hydrolysis of Penicillin G potassium salt (PGK) with the kinetically controlled enzymatic coupling of in situ formed 6-aminopenicillanic acid (6-APA) with p-hydroxyphenylglycine methyl ester (D-HPGM) to give amoxicillin as the final product by using a single enzyme has been demonstrated successfully. Ethylene glycol (EG) was employed as a component of reaction buffer to improve the synthesis yield. Reaction parameters, including different cosolvents, EG content, the loading of immobilized penicillin G acylase (IPA), and reaction temperature and time were studied to evaluate their effects on the reaction. The best result of 55.2% yield was obtained from the reaction which was carried out in the mixed media containing 40% sodium dihydrogen phosphate buffer (apparent pH 6.0) and 60% EG (v/v), with the initial concentration 150 mM and 450 mM of PGK and D-HPGM, respectively, catalyzed by 50 IU/mL IPA at 25 °C for 10 h. The IPA could be recycled for nine batches without obviously losing of catalytic activity. The important strategy will have potential application in the β-lactam antibiotics industry due to the advantages of saving the effort of isolating 6-APA, reducing usual enzymatic steps and the industrial cost of amoxicillin synthesis.

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Abbreviations

PGK:

Penicillin G potassium salt

6-APA:

6-Aminopenicillanic acid

D-HPGM:

p-Hydroxyphenylglycine methyl ester

EG:

Ethylene glycol

IPA:

Immobilized penicillin G acylase

PGA:

Penicillin G acylase

CLEAs:

Cross-linked penicillin acylase aggregates

7-ADCA:

7-Aminodeacetoxycephalosporanic Acid

HPGA:

p-Hydroxyphenylglycine amide

HPLC:

High-performance liquid chromatography

UV:

Ultraviolet

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Acknowledgments

The financial supports from National Natural Science Foundation of China (No. 20704037) and Zhejiang Provincial Science and Technology Council (No. 2005C11023-03) are gratefully acknowledged.

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Authors and Affiliations

  1. Department of Chemistry, Zhejiang University, Hangzhou, 310027, People’s Republic of China

    Qi Wu, Chun-Xiu Chen, Li-Li Du & Xian-Fu Lin

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  1. Qi Wu
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  2. Chun-Xiu Chen
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  3. Li-Li Du
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  4. Xian-Fu Lin
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Correspondence to Xian-Fu Lin.

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Wu, Q., Chen, CX., Du, LL. et al. Enzymatic Synthesis of Amoxicillin via a One-pot Enzymatic Hydrolysis and Condensation Cascade Process in the Presence of Organic Co-solvents. Appl Biochem Biotechnol 160, 2026–2035 (2010). https://doi.org/10.1007/s12010-009-8847-x

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