First Line Anti-Tuberculosis Drugs Resistance Patterns of <i>Mycobacterium tuberculosis</i>Isolates from Newly Diagnosed Cases of Tuberculosis

Introduction: Tuberculosis is a major cause of mortality and morbidity world-wide. Anti-tuberculosis drugs have been used for many decades but resistance to them is now widespread. Globally 5% of tuberculosis cases and in India 3% among new TB cases. This study was planned to know the pattern of first line anti-tuberculosis drug resistance in south Gujarat, Surat region in newly diagnosed patients of tuberculosis. Material and Methods: 350 samples were processed for homogenisation and concentration using 4% NAOH-2.9% trisodium citrate. Processed samples were inoculated in liquid medium that is MGIT (Mycobacterial growth indicator tube). Positive samples for M. tbwere processed further for first line anti-tuberculosis drugs sensitivity testing (DST). Reading was taken by using MicroMGIT system. Result: Out of 350 samples 59 (17%) were positive samples, of which 48 (13%) were M. tb and 11 (3%) were non tuberculous mycobacteria. Out of 48 samples 2% (1 isolate) was resistant to isoniazid and Rifampicin while 2% were monoresistant to isoniazide, 2% monoresistant to streptomycin. No rifampicin monoresistant was detected. Conclusion: Such study may help in control of tuberculosis at regional and national level which would in turn help in planning of measures to control Multi-drug resistance tuberculosis. Continuous surveillance should be applied to know the periodic changing patterns and trend in Drug resistant tuberculosis.

Use of Cost Effective Semi-Automated (Mannual/Micro) MGIT System over BACTEC 960 to Perform First Line Anti-Tuberculosis Drugs Sensitivity Testing

Introduction: Multi-drug resistant tuberculosis (MDR-TB) that is the tuberculosis that is resistant to at least 2 of the first line anti-tuberculosis drugs is fatal infectious disease. Cases of MDR-TB are now increasing with 30,000 cases of MDR-TB reported in 2013 by national TB programme. Rapid diagnosis of MDR-TB is extremely important for rapid treatment of patient and to prevent spread of MDR-TB to other. BACTEC 960 system helps in rapid diagnosis but purchase of expensive instrument for the same is the limitation. However, the same purpose can be solved by use of semi-automated MGIT system. Aims and Objectives: Aim of this study is to do drug sensitivity testing of the first line anti-tuberculosis drugs with the use of semi-automated MGIT systems. 350 newly registered and suspected cases of tuberculosis in tertiary care hospital were included. Samples were processed for digestion and decontamination and inoculated in MGIT tubes and also on LJ medium. Reading was taken using semi-automated MGIT system. Positive tubes were confirmed by rapid test for M. tuberculosis and then drug sensitivity was performed. Result: Out of 350 samples, 62% were sputum;33% were pleural fluid and rest 5% were lymph node, Ascetic fluid, CSF, pus. Average day of positivity by MGIT was 13 - 20 days as compared to 25 - 37 days by solid medium, which was statistically significant with p value Conclusion: Manual MGIT System is a simple, efficient, safe to use diagnostic system. It does not require any expensive/special instrumentation other than the UV lamp for detection of fluorescence. The rapidity by which mycobacteria are detected is the most important advantage of the Manual MGIT. In areas with limited resources where purchase of expensive instruments such as the MGIT960 is out of scope, the use of manual MGIT for rapid susceptibility testing for MDR-TB could be a possibility.

NAT2＊6A, a haplotype of the N-acetyltransferase 2 gene, is an important biomarker for risk of anti-tuberculosis drug-induced hepatotoxicity in Japanese patients with tuberculosis

AIM: To investigate an association between N -acetyltransferase 2 (NAT2 )-haplotypes/diplotypes and adverse effects in Japanese pulmonary tuberculosis patients. METHODS: We studied 100 patients with pulmonary TB treated with anti-TB drugs including INH. The frequencies and distributions of single nucleotide polymorphisms, haplotypes, and diplotypes of NAT2 were determined by the PCR-restriction fragment length polymorphism method, and the results were compared between TB patients with and without adverse effect, using multivariate logistic regression analysis.RESULTS: Statistical analysis revealed that the frequency of a variant haplotype, NAT2*6A , was signifi cantly increased in TB patients with hepatotoxicity, compared with those without hepatotoxicity [P = 0.001, odds ratio (OR) = 3.535]. By contrast, the frequency of a wild-type (major) haplotype, "NAT2*4", was signif icantly lower in TB patients with hepatotoxicity than those without hepatotoxicity (P < 0.001, OR = 0.265). There was no association between NAT2-haplotypes and skin rash or eosinophilia. CONCLUSION: The present study shows that NAT2 is one of the determinants of anti-TB drug-induced hepatotoxicity. Moreover, the haplotypes, NAT2*4 and NAT2*6A, are useful new biomarkers for predicting anti- TB drug-induced hepatotoxicity.

Pyrrolidine dithiocarbamate alleviates the anti-tuberculosis drug-induced liver injury through JAK2/STAT3 signaling pathway:An experimental study

Objective:To study the effect of pyrrolidine dithiocarbamate(PDTC) on the anti-tuberculosis drug-induced liver injury and the molecular mechanism. Methods:Clean male SD rats were selected as experimental animals and randomly divided into normal group,model group,PDTC group and AG490 group. Animal model of anti-tuberculosis drug-induced liver injury was established by intragastric administration isoniazid + rifampicin. PDTC group received intraperitoneal injection of PDTC,and AG490 group received intraperitoneal injection of AG490. Twenty-eight days after intervention,the rats were executed,and the liver injury indexes,inflammation indexes and oxidative stress indexes in serum as well as JAK2/STAT3 expression,liver injury indexes,inflammation indexes and oxidative stress indexes in liver tissue were determined. Results:p-JAK2,p-STAT3,TNF-α,IL-1β,IL-6,ROS,8-OHdG and MDA expression in liver tissue as well as TBIL,ALT,AST,γ-GT,TNF-α,IL-1β,IL-6,ROS,8-OHdG and MDA levels in serum of model group were significantly higher than those of normal group while p-JAK2,p-STAT3,TNF-α,IL-1β,IL-6,ROS,8-OHdG and MDA expression in liver tissu as well as TBIL,ALT,AST,γ-GT,TNF-α,IL-1β,IL-6,ROS,8-OHdG and MDA levels in serum of PDTC group and AG490 group were significantly lower than those of model group. Conclusions:PDTC can inhibit the inflammation and oxidative stress mediated by JAK2/STAT3 signaling pathway to alleviate the anti-tuberculosis drug-induced liver injury.

Drug Resistance Pattern in Pulmonary Tuberculosis Patients and Risk Factors Associated with Multi-Drug Resistant Tuberculosis

Introduction: Anti-tuberculosis drug resistance is a major problem in tuberculosis (TB) control programme, particularly multi-drug resistance TB (MDR-TB) in Nepal. Drug resistance is difficult to treat due to its associated cost and side effects. The objective of this study was to assess the drug resistance pattern and assess risk factor associated with MDR-TB among pulmonary tuberculosis patients attending National Tuberculosis Center. Methodology: The comparative cross sectional study was conducted at National Tuberculosis Center during August 2015 to February 2015. Early morning sputum samples were collected from pulmonary tuberculosis suspected patients and subjected to Ziehl-Neelsen staining and fluorochrome staining and culture on Lowenstein-Jensen (LJ) medium. Drug Susceptibility test was performed on culture positive isolates by using proportion method. Univariate and multivariate analysis was computed to assess the risk factors of MDR-TB. Results: Out of 223 sputum samples, 105 were fluorochrome staining positive, 85 were ZN staining positive and 102 were culture positive. Out of 102 culture positive isolates, 37.2% were resistance to any four anti-TB drugs. 11 (28.9%) were initial drug resistance and 28 (43.7%) were acquired drug resistance. The overall prevalence of MDR-TB was 11.7%, of which 2 (5.3%) were initial MDR-TB and 10 (15.6%) were acquired MDR-TB. Univariate and multivariate analysis showed female were significantly associated (P = 0.05) with MDR-TB. Conclusion: Drug resistance TB particularly MDR-TB is high. The most common resistance pattern observed in this study was resistance to both isoniazid and rifampicin. Female were found to be associated with MDR-TB. Thus, early diagnosis of TB and provision of culture and DST are crucial in order to combat the threat of DR-TB.

Research progress on the drug action and resistance mechanism in Mycobacterium tuberculosis

Tuberculosis(TB)is a chronic infectious disease caused by Mycobacterium Tuberculosis(MTB).It is the second largest single cause of death besides novel coronavirus pneumonia.Along with the abuse of antibiotics and extensive use of anti-tuberculosis drugs,multidrug-resistant(MDR)TB,drug-resistant(XDR)TB and totally drug-resistant(TDR)TB became obstacles to the tuberculosis eradication worldwide.According to the World Health Organization(WHO)statistics,China is not only a high burden tuberculosis country in the world,but also a country with a serious epidemic of MDR.Traditional drugs fail to meet the needs of tuberculosis control.Therefore,it is urgent to find new targets of anti-tuberculosis drugs and develop new anti-tuberculosis drugs.Hence,this paper systematically summarizes the mechanism of traditional and newly developed anti-tuberculosis drugs,in which stressing the research progress of drug resistance mechanisms.This work provides us with new insights of new anti-tuberculosis drug developments,and may contribute to a reduction in the harm that tuberculosis brings to society.

Endogenous peroxynitrite activated fluorescent probe for revealing anti-tuberculosis drug induced hepatotoxicity

Py^(+)razinamide (PZA), isoniazid (INH) and rifampicin (RFP) are all commonly used anti-tuberculosis drugs in clinical practice, and long-term medication may cause severe liver damage and toxicity. The level of peroxynitrite (ONOO^(-)) generated in liver has long been regarded as a biomarker for the prediction and measurement of drug-induced liver injury (DILI). In this article, we constructed a BODIPY-based fluorescent probe (BDP-Py^(+)) that enabled quickly and sensitively detect and image ONOO^(-) in vivo. Utilizing this probe, we demonstrated the change of ONOO^(-) content in cells and mice model of DILI induced by acetaminophen (APAP), and for the first time revealed the mechanism of liver injury induced by antituberculosis drug PZA. Moreover, BDP-Py^(+) could be applied to screen out and evaluate the hepatotoxicity of different anti-tuberculosis drugs. Comparing with the existing serum enzymes detection and H&E staining, the probe could achieve early diagnosis of DILI before solid lesions in liver via monitoring the up-regulation of ONOO^(-) levels. Collectively, this work will promote the understanding of the pathogenesis of anti-tuberculosis drug induced liver injury (ATB-DILI), and provide a powerful tool for the early diagnosis and treatment of DILI.

防治阿尔茨海默病多靶点药物研究进展

阿尔茨海默病(AD)是一种复杂的神经退行性疾病,临床上用于AD治疗的药物均为单靶点药物。但AD可引起全身性病理变化,其发病过程涉及多种调控因子和调控网络且囊括多种疾病假说。因此,对于AD的治疗策略目前更倾向于多靶点治疗。本文将近期报道的各类多靶点AD防治药物及化合物区分为单一化学实体多靶点药物、多组分药物和联合用药3类,分别对其研究进展进行综述。

偏最小二乘法用于药物分析

本文研究了偏最小二乘法(PLS)用于药物的紫外可见多组份光度分析。当混合物中诸组份存在相互作用、光谱加和性受到扰动时,PLS法较之常规最小二乘(OLS)及多元线性回归(MLR)法有显著优点。本法用于去痛片的分析,得到了良好的回收率。

中药成药性本质分析及其与重大新药创制的关系

药物成药性在药物开发中至关重要,是药物研究的源头。中药是多成分药物,由于其存在复杂性,在成药性研究中必然面临更大的挑战。本文主要结合中药多成分的化学物质基础本质,提出了研究中药成药性分析的一些观点和思路方法。

自乳化释药系统对丹参酮类多组分增溶作用的研究

目的:探讨自乳化释药系统(SEDDS)对丹参酮类组分增溶的效果。方法:采用D-最优混料设计优化SEDDS处方,以隐丹参酮、丹参酮Ⅰ和丹参酮Ⅱ_A含量、SEDDS的粒径和乳化时间为指标对SEDDS进行优化并对其进行综合评价。结果:优化得SEDDS最佳处方为:油酸∶吐温-20∶二乙二醇单乙基醚(0.25∶0.52∶0.23),SEDDS粒径为(204±5)nm,乳化时间为(59±2)s。37℃时,SEDDS对隐丹参酮、丹参酮Ⅰ和丹参酮Ⅱ_A的最大载药量分别为5.075、3.397和6.559 mg/g,在水中的溶解度分别为0.138、0.238和0.367 mg/mL。结论:SEDDS对丹参酮类组分的增溶效果显著。

HPLC波长切换法同时测定理气散结颗粒中4个成分的含量

目的建立同时测定理气散结颗粒中氧化芍药苷、芍药内酯苷、芍药苷和α-香附酮的HPLC波长切换联合梯度洗脱法。方法采用Venusil MP C18色谱柱(4.6 mm×250 mm,5μm),以乙腈(A)-0.1%磷酸溶液(B)为流动相,梯度洗脱(0~15 min,12.0%A;15~31 min,12.0%A→26.0%A;31~39 min,26.0%A→40.0%A;39~45 min,40.0%A→12.0%A),流速0.9 mL/min,波长切换(0~31 min,在230 nm波长下检测氧化芍药苷、芍药内酯苷和芍药苷;31~45 min,在242 nm波长下检测α-香附酮),柱温30℃,进样量为20μL。结果氧化芍药苷、芍药内酯苷、芍药苷和α-香附酮4个成分的质量浓度分别在5.15~103.00μg/mL(r=0.999 6)、8.99~179.80μg/mL(r=0.999 5)、19.83~396.60μg/mL(r=0.999 9)、6.35~127.00μg/mL(r=0.999 3)呈良好线性关系;平均加样回收率及相应的RSD(n=6)分别为96.83%(0.93%)、97.85%(1.31%)、99.71%(0.80%)、98.77%(1.59%)。结论所建立的方法灵敏度高、快速、专属性好、准确度高,为理气散结颗粒的质量控制提供了依据。

人工神经元网络用于复方氯丙嗪的含量测定

将误差反向传播（ＢＰ）的人工神经元网络（ＡＮＮ）模型，用于复方氯丙嗪紫外重叠光谱分析。对ＡＮＮ模型的参数进行了优化。采用ｆ（ｘ）＝１／（１＋ｅ－ｘ）作为网络节点的输入输出转换函数的三层神经网络具有较佳性能，当取隐含节点数为１５时，该网络预测结果的最小平均相对误差为１．２２％，将研制的ＡＮＮ模型用于复方氯丙嗪含量测定，其结果与药品标准法和偏最小二乘法（ＰＬＳ）一致。

基于多成分药物代谢的秦艽质控成分遴选研究

目的:基于中药多成分作用特点,探讨秦艽多成分在体内的代谢情况,遴选秦艽质控成分。方法:制备秦艽水提物,采用相应的实验操作手术,收集秦艽的肠代谢、肝代谢以及综合代谢样品,结合超高效液相色谱-四级杆/静电场高分辨质谱联用技术(UPLC-Q-Exactive MS),对秦艽水提物、肠代谢样品、肝代谢样品及综合代谢样品中的成分进行鉴别,遴选秦艽的质控成分。结果:秦艽水提物中共鉴定出33个化学成分,其中有12个成分在经肠代谢样品中可检测到,有9个成分在先经肠代谢后经肝代谢样品中可检测到,有11个成分在经灌胃给药后的综合代谢样品中检测到。结论:根据质谱数据分析结果,遴选出秦艽中9个可以以原型的形式进入血液循环的成分作为该药材的质控成分,包括马钱苷酸、secologanoside、6′-O-β-葡萄糖龙胆苦苷、獐牙菜苦苷、龙胆苦苷、獐牙菜苷、异牡荆素、淫羊藿苷、齐墩果酸,作为秦艽的质控成分。

生物医学新兴学科与中药现代化——现代组合成分药物的研究

中药的特色是通过多成分、多靶点的整体调控作用来系统纠正疾病造成的机体失衡。中药在治疗多基因、多因素引起的某些复杂疾病方面相对西药(通常是单一化学成分药物)有独到优势。但以天然动植物为来源的中药在质量控制和疗效科学依据方面的不足严重影响了中药在现代社会的应用。目前,组合治疗现代医学和系统生物学的发展为中药现代化提供了新的机会。提出应用现代组合成分药物和组合治疗新概念,科学阐明中药药效的学术观点;并提出综合应用蛋白质组学和化学生物学等生物医学新兴科技从中药中开发现代组合成分新药的技术。

HPLC波长切换法同时测定金嗓散结胶囊中的(R,S)-告依春、羟基红花黄色素A、丹酚酸B、木蝴蝶苷B和木蝴蝶苷A

目的:建立金嗓散结胶囊中(R,S)-告依春、羟基红花黄色素A、丹酚酸B、木蝴蝶苷B和木蝴蝶苷A的同时测定方法。方法:采用HPLC法,应用Agilent HC-C18(250 mm×4.6 mm,5μm)色谱柱;以甲醇-乙腈(3∶1)与0.4%磷酸溶液为流动相,进行梯度洗脱;流速为1.0 m L·min^(-1);供试品采用50%甲醇超声提取。结果:(R,S)告依春、羟基红花黄色素A、丹酚酸B、木蝴蝶苷B和木蝴蝶苷A的线性范围分别为4.150~83.00、5.120~102.4、11.07~221.4、4.590~91.80、8.140~162.8μg·m L^(-1),相关系数分别为0.999 2、0.999 9、0.999 4、0.999 7、0.999 5;5种成分分离良好,阴性对测定无干扰,平均加样回收率及相应的RSD分别为99.06%(1.27%)、99.54%(1.15%)、98.19%(0.89%)、97.88%(1.71%)、97.22%(0.99%)。结论:所建立的方法可用于金嗓散结胶囊的质量控制。

多组分药物相互作用定量评价在抗艾滋病毒研究中的应用

目的定量评价四组分药物抗艾滋病毒作用的相互关系。方法用MTT法检测天然药物四组分药物对细胞生长的影响,在最大无毒剂量以下观察4种药物组分单用及合用的抗病毒作用,用中效原理计算2种及4种药物组分合用后的合用指数(combination index,CI)值,以CI值评价不同药物合用后抗病毒作用的相互关系。结果 AB药物组分合用时以协同作用为主,大剂量趋向作用相加;CD药物组分合用以拮抗作用为主,大剂量趋向相加;但四药合用10%抑制率以上均为协同作用。结论采用中效原理可以定量评价4组分药物之间抗艾滋病毒的相互作用关系。

基于多成分药物代谢的枸杞子质控成分遴选研究

目的:基于多成分序贯代谢的研究策略,探讨枸杞子在体内的代谢成分,遴选枸杞子的质控成分。方法:制备枸杞子水提液后,运用在体双向肠灌流技术和灌胃采血技术收集其肠代谢、肝代谢及综合代谢样品,结合超高效液相色谱-四级杆/静电场高分辨质谱联用技术(UPLC-Q-Exactive MS),对枸杞子水提物、肠代谢样品、肝代谢样品及综合代谢样品中的成分进行鉴别,遴选枸杞子的质控成分。结果:枸杞子提取物经LC-MS分析,共鉴定出36个化学成分。序贯代谢研究表明,枸杞子中有20个原形入血成分,其中有17个成分在经过肠代谢后可以检测到,16个成分在经过肝代谢后可以检测到,11个成分经过灌胃给药后,在腹主动脉血中可以检测到。结论:根据序贯代谢的研究结果,可以将这11个成分作为枸杞子药材的候选指标性成分,其分别为咖啡酸己糖偶联物、香豆酸己糖偶联物、N-caffeoyl,N′-dihydrocaffeoyl spermidine dihexose、绿原酸、N1,N10-bis(dihydrocaffeoyl)spermidine hexose、香豆素、槲皮素-O-芸香糖苷、阿魏酸、芦丁、N-乙酰色氨酸、水杨酸。

化学药及中草药的代谢与转运:从分子到疾病

Detailed ADME (absorption, distribution, metabolism and excretion) evaluation is required in the preclinical stage of drug development. The preclinical ADME data may be extrapolated to humans and some predictions may be made with the application of proper pharmacokinetic and pharmacodynamic principles. Novel drugs with validated drug targets and favorable ADME properties are being sought by drug scientists, and oral administration is becoming a major delivery approach for many drugs due to its great safety, reduced cost, improved quality of life, and the potential for improved efficacy. The incorporation of herbal therapy into the main stream medical-care system has been encouraged by World Health Organization (WHO). Chinese herbal medicines may represent an important group of multi-component therapeutics suitable for the management of diseases with networked abnormalities such as cancer and diabetes. However, the disposition pathways, pharmacological targets and safety profiles of most Chinese herbal medicines are unknown or not well characterized. The evidence for the use of Chinese herbal medicines as effective and safe multi-component therapeutics is not established. The quality control and standardization of these natural products still constitute a major challenge in their evidence-based clinical application. Recently, there is an increased interest in the incorporation of systems biology approaches and combined use of preclinical and clinical models in understanding the complex disposition pathways, multiple targets and toxicities of Chinese herbal medicines. Like synthetic drugs, Chinese herbal medicines are subject to similar ADME processes in the body like synthetic drugs. However, data are scant on the ADME of most Chinese herbal medicines. Thus, it is important to characterize ADME for both drugs and Chinese herbal medicines. My research team and collaborators have synthesized several small molecule novel compounds for cancer treatment, including 5,6-dimethylxanthenone-4-acetic acid (DMXAA, an antiangiogenic agent and TNF-( inducer) and SYUIQ-5, a novel telomerase inhibitor. The metabolic pathways and transport properties of both DMAA and SYUIQ-5 have been characterized. We have also investigated the metabolism and transport of the major active components of several commonly used Chinese herbal medicines, such as Danshen (Salvia miltiorrhiza) and ginkgo (Ginkgo biloba) using a combination of in vitro and in vivo models. It is believed that ADME studies are necessary and important for both synthetic drugs and Chinese herbal medicines in an attempt to establish their efficacy and safe-ty profiles.

基于多成分序贯代谢的石榴皮中枢神经系统保护作用物质基础研究

将序贯代谢法与超高效液相色谱-串联质谱(UPLC-MS/MS)相结合,研究石榴皮发挥中枢神经系统保护作用的物质基础。首先制备石榴皮提取液,通过肠灌流和封闭肠环法收集肠壁代谢、肠道菌群代谢的血浆样品,经液相色谱-串联质谱分析以原型吸收入血成分和代谢产物;综合代谢组大鼠灌胃给药后,鉴定其血中移行成分;再结合以上结果对脑脊液样品进行解析,鉴定出入脑成分。结果表明,从提取液中共检测到鞣质类、黄酮类、酚酸类和异香豆素类等38个化学成分。肠壁代谢、肠道菌群代谢情况基本一致,检测到30个原型入血成分和12个代谢产物;综合代谢组血浆中检测到7个原型入血成分和16个代谢产物;在脑脊液中发现2个原型成分和4个代谢产物,分别是柠檬酸、鞣花酸和2个甲基鞣花酸同分异构体、二甲基鞣花酸、二甲基鞣花酸葡萄糖醛酸化物。本研究基于多成分序贯代谢靶向识别出石榴皮及其代谢产物中入脑成分,可为阐明石榴皮中枢神经系统保护作用提供证据。