Page 35 - 《中国药房》2021年第4期

Effects of Rat Intestinal Flora on the Pharmacokinetic Parameters of Pyrazinamide and Its Active
Metabolite Pyrazinoic Acid
LIU Qingxiang ，WU Zhenghua ，LAI Yalin ，FAN Guorong ，FAN Qi （1. School of Pharmacy，Chongqing
2
1
2
1
1
Medical University，Chongqing 400016，China；2. Dept. of Clinical Pharmacy，Shanghai General Hospital
Affiliated to Shanghai Jiao Tong University，Shanghai 200080，China）
ABSTRACT OBJECTIVE：To study the effects of rat intestinal flora on the pharmacokinetic parameters of pyrazinamide and its
active metabolite pyrazinoic acid. METHODS：Totally 16 SD rats were randomly divided into trial group and control group，with 8
rats in each group. Trial group was given mixed antibiotics（streptomycin sulfate+neomycin sulfate）intragastrically to construct
pseudoaseptic rat model. After modeling，both groups were given pyrazinamide intragastrically（150 mg/kg）. Before and 0.167，
0.333，0.667，1，1.5，2，3，4，6，9 h after administration，0.1 mL blood sample was collected from orbital venous plexus，and
0.3 mL blood sample was collected from orbital venous plexus 12，24 h after administration. Using phenacetin as internal standard，
LC-MS/MS method was adopted to determine the plasma concentration of pyrazinamide and pyrazinoic acid. The determination was
performed on Agilent ZORBAX SB-Aq column with mobile phase consisted of 0.2％ formic acid（containing 8 mmol/L ammonium
acetate）-methanol（gradient elution）at the flow rate of 1 mL/min. The column temperature was set at 30 ℃，and sample size was
10 μL. The ion source was ESI and the temperature of ion source was 500 ℃. The collision gas was nitrogen and the pressure was
10 psi. The temperature of mass transfer interface was 100 ℃. The mass spectrum monitoring mode was multi reaction monitoring，
and the collection mode was positive ion mode. The monitoring transition ion-pairs were m/z 124.0→79.0（pyrazinamide），m/z
125.1→79.1（pyrazinic acid）and m/z 180.0→110.2（internal standard）. The de-clustering potential and collision voltage were 55，
26 and 85 V，24，23 and 28 V，respectively. The pharmacokinetic parameters were calculated and compared by using DAS 2.1.1
software. RESULTS：The linear ranges of pyrazinamide and pyrazinoic acid were 25-5 000 ng/mL（r＝0.997 6）and 100-12 500
ng/mL（r＝0.999 0）. The lower limits of quantification were 25 and 100 ng/mL，respectively. Intra-batch and inter-batch accuracy
were 92.93％-100.50％，and RSDs of intra-batch and inter-batch precision and matrix effect tests were all lower than or equal to
8.42％（n＝6 or n＝3）. Compared with control group，tmax of pyrazinamide in trial group was prolonged significantly（P＜0.01）；
there was no statistical significance in other pharmacokinetic parameters between 2 groups （P＞0.05）. CONCLUSIONS：The
absorption of single dose pyrazinamide is delayed with the change of intestinal flora in rats.
KEYWORDS Pyrazinamide；Pyrazinoic acid；Rat；Intestinal flora；Pharmacokinetics；LC-MS/MS

[1]
结核病是全球最严重的传染病之一 。吡嗪酰胺
（化学结构式见图 1A）是治疗结核病的一线药物，对药
物敏感性和多药耐药性结核病均表现出独特、优良的治
疗作用，其不但能够降低结核病复发率，还能够缩短结
核病治疗周期，常用于对异烟肼和利福平耐药的结核病 A.吡嗪酰胺 B.吡嗪酸
[2]
患者的治疗 。有研究表明，吡嗪酰胺经肝微粒体酰胺 图1 吡嗪酰胺和吡嗪酸的化学结构式
酶代谢产生的主要活性代谢产物吡嗪酸（化学结构式见 Fig 1 The chemical structures of pyrazinamide and
图1B）可与天冬氨酸脱羧酶结合，中断结核分枝杆菌辅 pyrazinoic acid
酶A的生物合成，从而起到杀菌作用 。但是，相较于其 菌 SD 大鼠模型，建立测定大鼠血浆中吡嗪酰胺及其活
[3]
它抗结核药物，吡嗪酰胺被证实具有更强的肝毒性 [4－6] 。 性 代 谢 产 物 吡 嗪 酸 浓 度 的 液 相 色 谱 - 串 联 质 谱 法
已有研究报道，联用吡嗪酰胺酶抑制剂可以减轻吡嗪酰 （LC-MS/MS），研究大鼠肠道菌群对吡嗪酰胺和吡嗪酸
胺诱导的 SD 大鼠肝损伤，由此推测该药引起的肝毒性 药动学参数的影响，旨在为提高吡嗪酰胺的疗效及降低
[7]
可能与其代谢产物吡嗪酸有关 。可见，关注吡嗪酰胺 其毒副作用提供参考。
的体内代谢，对提高抗结核病的疗效、降低毒副作用均 1 材料
具有重要意义。 1.1 主要仪器
口服药物在胃肠道被吸收入血前，可能被寄居在肠 LC-20A型液相色谱系统（配有LC-20AD型二元泵、
道内的菌群转化为具有生物活性、生物惰性或毒性的代 DGU-20A3 型脱气机、SIL-20A 型自动进样器、CTO-
谢产物，然后再吸收入血 。本课题组前期研究显示，灌 20AC 型柱温箱）、AUW220D 型电子分析天平均购自日
[8]
胃吡嗪酰胺后，SD 大鼠体内吡嗪酰胺的胆汁浓度远高 本 Shimadzu 公司，API 4000 型三重四极杆质谱仪（配有
于粪便浓度，据此推测吡嗪酰胺可能在肠道内发生了生 Analyst software 1.6 工作站）购自美国 AB Sciex 公司，
物转化。基于此，本文拟通过灌胃混合抗生素复制伪无 MX-S 型涡旋混合仪购自美国 Scilogex 公司，SK7200H

中国药房 2021年第32卷第4期 China Pharmacy 2021 Vol. 32 No. 4 ·413 ·