Maia Alexandra S, Castro Paula M L, Tiritan Maria Elizabeth
CESPU, Instituto de Investigação e Formação Avançada em Ciências e Tecnologias da Saúde, Rua Central de Gandra, 1317, 4585-116 Gandra PRD, Portugal; Universidade Católica Portuguesa, CBQF - Centro de Biotecnologia e Química Fina - Laboratório Associado, Escola Superior de Biotecnologia, Rua Arquiteto Lobão Vital, Apartado 2511, 4202-401 Porto, Portugal.
Universidade Católica Portuguesa, CBQF - Centro de Biotecnologia e Química Fina - Laboratório Associado, Escola Superior de Biotecnologia, Rua Arquiteto Lobão Vital, Apartado 2511, 4202-401 Porto, Portugal.
J Chromatogr B Analyt Technol Biomed Life Sci. 2016 Sep 1;1029-1030:174-183. doi: 10.1016/j.jchromb.2016.06.026. Epub 2016 Jun 16.
Ofloxacin is a chiral fluoroquinolone commercialized as racemate and as its enantiomerically pure form levofloxacin. This work presents an integrated liquid chromatography (LC) method with fluorescence detection (FD) and exact mass spectrometry (EMS) developed to assess the enantiomeric biodegradation of ofloxacin and levofloxacin in laboratory-scale microcosms. The optimized enantioseparation conditions were achieved using a macrocyclic antibiotic ristocetin A-bonded CSP (150×2.1mm i.d.; particle size 5μm) under reversed-phase elution mode. The method was validated using a mineral salts medium as matrix and presented selectivity and linearity over a concentration range from 5μgL(-1) (quantification limit) to 350μgL(-1) for each enantiomer. The method was successfully applied to evaluate biodegradation of ofloxacin enantiomers at 250μgL(-1) by an activated sludge inoculum. Ofloxacin (racemic mixture) and (S)-enantiomer (levofloxacin) were degraded up to 58 and 52%, respectively. An additional degradable carbon source, acetate, enhanced biodegradation up to 23%. (S)-enantiomer presented the highest extent of degradation (66.8%) when ofloxacin was supplied along with acetate. Results indicated slightly higher biodegradation extents for the (S)-enantiomer when supplementation was done with ofloxacin. Degradation occurred faster in the first 3days and proceeded slowly until the end of the assays. The chromatographic results from LC-FD suggested the formation of the (R)-enantiomer during levofloxacin biodegradation which was confirmed by LC-MS with a LTQ Orbitrap XL.
氧氟沙星是一种手性氟喹诺酮类药物，以消旋体和对映体纯形式左氧氟沙星上市。本文介绍了一种集成液相色谱（LC）方法，该方法结合荧光检测（FD）和精确质谱（EMS），用于评估实验室规模微观世界中氧氟沙星和左氧氟沙星的对映体生物降解情况。使用大环抗生素瑞斯托菌素A键合的手性固定相（CSP）（内径150×2.1mm；粒径5μm）在反相洗脱模式下实现了优化的对映体分离条件。该方法以矿物盐培养基为基质进行了验证，对每种对映体在5μg L⁻¹（定量限）至350μg L⁻¹的浓度范围内具有选择性和线性。该方法成功应用于评估活性污泥接种物对250μg L⁻¹氧氟沙星对映体的生物降解情况。氧氟沙星（外消旋混合物）和（S）-对映体（左氧氟沙星）的降解率分别高达58%和52%。额外的可降解碳源乙酸盐可将生物降解率提高至23%。当氧氟沙星与乙酸盐一起供应时，（S）-对映体的降解程度最高（66.8%）。结果表明，用氧氟沙星进行补充时，（S）-对映体的生物降解程度略高。降解在最初3天内发生得更快，之后进展缓慢直至试验结束。LC-FD的色谱结果表明，左氧氟沙星生物降解过程中形成了（R）-对映体，这通过LTQ Orbitrap XL的LC-MS得到了证实。
