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目的建立柱前手性衍生化-反相高效液相色谱法(HPLC)测定L-肌肽对映体的纯度。方法以邻苯二甲醛/N-乙酰基-L-半胱氨酸(OPA/NAC)为手性衍生化试剂,反应生成具有荧光吸收的一对非对映异构体衍生物,采用C18色谱柱(250mm×4.6mm,5μm),流速1.0mL·min-1,50mmol·L-1乙酸铵缓冲溶液(pH6.0)-甲醇(70:30,V/V)流动相,柱温35℃,λex=350nm,λem=450nm。结果L-肌肽与其光学异构体分离度大于3,在0.104～2.68mg·L-1范围内,D-肌肽衍生物色谱峰面积与其浓度呈良好线性关系,检测限浓度为0.02mg·L-1。结论建立的L-肌肽光学异构体(杂质)手性衍生化-HPLC拆分检查法方便准确,可用于L-肌肽的光学纯度控制。
Objective To establish a pre-column chiral derivatization-reversed phase high performance liquid chromatography (HPLC) for the determination of enantiomeric purity of L-carnosine. Methods O-phthalaldehyde / N-acetyl-L-cysteine (OPA / NAC) was used as a chiral derivatization reagent to generate a pair of diastereoisomer derivatives with fluorescence absorption. C18 chromatography (250 mm × 4.6 mm, 5 μm), a flow rate of 1.0 mL · min -1, a 50 mmol·L -1 ammonium acetate buffer (pH 6.0) -methanol (70:30, V / V) λex = 350 nm, λem = 450 nm. Results The separation of L-carnosine and its optical isomer was more than 3. The linear range of the peak area of D-carnosine derivative was 0.104 ~ 2.68 mg · L-1. The limit of detection was 0.02 mg · L- 1. Conclusion The establishment of L-carnosine optical isomer (impurity) chiral derivatization-HPLC resolution method is convenient and accurate and can be used for optical purity control of L-carnosine.