高效液相色谱法对N-乙酰-L-酪氨酸生产工艺的中间物监控分析

Monitoring and analysis of intermediates in N-acetyl-L-tyrosine production by high performance liquid chromatography

为了探讨在生产N-乙酰-L-酪氨酸时,中间物料对产品质量的影响,试验研究了在采用乙酸酐乙酰化生产N-乙酰-L-酪氨酸过程中,对中间产生物料L-酪氨酸、N-乙酰-L-酪氨酸和N,O-二乙酰-L-酪氨酸的中控分析并测定它们的含量。具体方法是:把样品用超纯水稀释100倍后,在色谱柱(C_(18),150mm×4.6mm,5μm)上进行分离,以10mmol·L^(-1)磷酸盐(pH3.0)-甲醇(60:40,V/V)为流动相进行等度洗脱,在波长270nm处进行紫外检测。结果表明:L-酪氨酸、N-乙酰-L-酪氨酸、N,0-二乙酰-L-酪氨酸的质量浓度分别在10~1000mg·L^(-1),10~1000mg·L^(-1),50~1000mg·L^(-1)内与对应的峰面积呈现线性关系,检出限(3S/N)分别为1.1mg L^(-1),1.2mg·L^(-1),14.0mg·L^(-1);对实际样品进行加标回收试验,回收率分别在98.4%~105.0%,98.5%~100.8%,98.9%~100.5%之间,测定值的相对标准偏差RSD(n=6)分别为1.8%、0.7%和0.9%,均小于2.0%。说明所建立的高效液相方法切实可行,重现好,精密度高,可用于乙酰酪氨酸生产工艺中间物的监控分析。

In order to investigate the influence of intermediate materials on the quality of N-acetyl-L-tyrosine,the intermediate mate-rials L-tyrosine,n-acetyl-L-tyrosine and N,O-diacetyl-L-tyrosine in the process of n-acetyl-L-tyrosine production by acetic anhydride acetylation were analyzed and their contents were determined.The specific method was:the sample was diluted 100 times with water,it was separated on a chromatographic column(C_(18),150mm×4.6 mm,5μm).The isocratic elution was performed with 10 mmol:L^(-1)phosphate(pH 3.0)-methanol(60:40,V/V)as the mobile phase.The UV detection was performed at the wavelength of 27Onm.The results showed that linear relationships were obtained between peak area and mass concentrations of L-tyrosine,N-ace-tyl-L-tyrosine,N,0-diacetyl-L-tyrosine in the ranges of 10~1000mg·L^(-1),10~100mg·L^(-1)0,50~1000mg·L^(-1),respectively.The detection limits(3S/N)were 1.Img·L-l,1.2mg·L^(-1),and 14.0mg·L^(-1),respectively.Tests for recovery was made by standard addition method to the actual samples,giving values of recovery between 98.4~105%,98.5~100.8%,98.9~100.5%,respectively.The relative standard deviations RSD(n=6)of the measured values werel.8%,0.7%,and 0.9%,all of which were less than 2.0%.The results showed that the HPLC method was feasible,reproductable and accurate,and could be used to monitor and ana-lyze the intermediate of acetyltyrosine production process.