高效毛细管电泳法测定酶法合成阿莫西林反应体系

Analysis of the Composition in the Enzymic Synthesis of Amoxicillin by High Performance Capillary Electrophoresis

建立了用高效毛细管电泳法对酶法合成阿莫西林反应体系中5种物质[阿莫西林(Amox),D-对羟基苯甘氨酸甲酯(D-HPGM),D-对羟基苯甘氨酸(D-HPG),6-氨基青霉烷酸(6-APA),苯乙酸(PAA)]进行定量测定的分析方法.以0.1%苯甲酸为内标,在电压25kV、波长214nm、柱温25℃、进样压力3.45kPa、进样时间5s的条件下,以pH9.0、聚乙二醇(PEG)4000含量为6mg/mL、浓度为50mmol/L的硼酸-硼砂缓冲溶液为流动相,5种物质与内标实现了基线分离.D-HPGM,D-HPG,Amox,6-APA和PAA分别在0.1～0.6,0.1～1.6,0.1～1.0,0.1～2.0和0.1～2.0mg/L浓度范围内,样品浓度与样品峰面积/内标峰面积的比值线性关系良好,回收率分别为101.45%,97.5%,101.5%,99.9%和101.3%,相对标准偏差分别为1.33%,1.69%,2.44%,1.27%,0.35%.该方法简单、快速、准确、重现性好,可取代高效液相色谱用于定量检测酶法合成阿莫西林的反应体系中物质的变化.

A analysis method for the determination of amoxicillin （Amox）, D-p-hydroxyphenylglycine methyl ester （D-HPGM）, D-p-hydroxyphenylglycine （D-HPG）, 6-aminopenicillanic acid （6-APA） and phenylacetamide （PAA） in the reaction of enzymic synthesis of amoxicillin by high performance capillary electrophoresis was established. The optimal separation was carried out at 25 ℃, 25 kV, injection pressure 3.45 kPa, inject time 5 s, and detection wavelength 214 nm, using 0.1% benzoic acid as internal standard and 50 mmol/L borate （pH 9.0） buffer with 6 mg/mL PEG4000. For D-HPGM, D-HPG, Amox, 6-APA and PAA, there is a good I/near relationship between the ratio of peak areas and the concentration in the ranges of 0.1-0.6, 0.1-0.16, 0.1-1.0, 0.1-2.0 and 0.1-2.0 mg/L respectively. The average recovery rates of the above five analytes are 101.45% （RsD=1.33%）, 97.5% （RsD=1.69%）, 101.5% （RsD=2.44%）, 99.9% （RsD=1.27%） and 101.3% （RsD=0.35%） respectively. The results show that the HPCE method is convenient, economical, sensitive and satisfactory in determination of the analytes in enzyme-based production of amoxicillin.