Africa drugs

Rapid Screening of Tuberculosis Drugs by Thin Layer Chromatography: 

Rapid Screening of Tuberculosis Drugs by Thin Layer Chromatography David Ndimbwa KA, Zambia Chipo Kuleya, Zimbabwe Howard Masiyachengo, Zimbabwe Zaveria Kimondo, Kenya

Introduction (1): 

Introduction (1) The World Health Organization declared tuberculosis (TB) to be a global emergency in 1993 Globally, TB incidence and prevalence, as well as TB drug resistance are increasing* HIV, increased poverty, migration, and travel are important factors responsible for these increases Factors contributing to drug resistance include poorly managed TB programs, poor patient compliance, drug shortages, and poor quality TB drugs *World Health Organization. Global Tuberculosis Control: WHO Report 2000. Geneva, Switzerland: World Health Organization, 2000; report no. WHO/CDS/CPC/TB/00.259.

Introduction (2): 

Introduction (2) To address the issue of assessing poor quality TB drugs, the WHO, FDA, CDC, and CENQAM together initiated a training and research course to learn and apply an FDA-developed thin layer chromatography technique, which is quick and low-cost, to the investigation of TB drug quality

Objectives: 

Objectives To train the participants to use TLC techniques To investigate the quality of TB drugs currently available in different African countries and India To generate an interest in determining quality of TB drugs through the use of TLC as a screening technique

Background: TB: 

Background: TB Tuberculosis is caused by Mycobacterium tuberculosis and is highly infectious Transmission is airborne Normal progression of disease without treatment 50% die, 25% spontaneous cure & 25% chronic excretors

World TB Situation: Estimated Incidence of TB 1998*: 

World TB Situation: Estimated Incidence of TB 1998* Country Cases (000’S) Rate /100K 1 India 1,828 186.1 2 China 1,414 112.6 3 Indonesia 591 286.6 6 Nigeria 259 243.4 8 S. Africa 172 437.9 9 Ethiopia 160 268.6 12 DRC 130 263.7 14 Tanzania 99 308.6 15 Kenya 86 296.8 19 Uganda 68 332.3 21 Zimbabwe 64 560.1 *World Health Organization. Global Tuberculosis Control: WHO Report 2000. Geneva, Switzerland: World Health Organization, 2000; report no. WHO/CDS/CPC/TB/00.259.

Background: Drug-resistant TB : 

Background: Drug-resistant TB Resistance to the conventional TB drugs is increasing* isoniazid, rifampicin, pyrazinamide, ethambutol, streptomycin There has also been an increase in multidrug- resistant TB (MDR-TB) (defined as resistance to INH/RIF)* *World Health Organization. Anti-tuberculosis Drug Resistance in the World: the WHO/IUATLD Global Project on Anti-tuberculosis Drug Resistance Surveillance, 2000. Report #2. Geneva, Switzerland: WHO Communicable Diseases Cluster, 2000; report no. WHO/CDS/TB/2000.278.

Background: Drug-resistant TB : 

Background: Drug-resistant TB As part of investigating increasing drug-resistant TB, there is a need to determine the quality of the TB drugs in the market If any of the drugs in the standard TB drug regimen are of low quality, this may lead to drug resistance, and the need for second-line TB drugs Drug resistant TB, especially MDRTB, is significantly more difficult and more expensive to treat

Background: TLC: 

Background: TLC The TLC kit technique was developed ~1995* TLC is a convenient, easy, quick, and inexpensive method to semi-quantitatively and qualitatively screen the quality of TB medications, and can identify different components in the TB drugs*† Both sensitivity and specificity of TLC exceed 90%** In previous studies, 1/10 (10%) INH samples and 4/30 (13%) RIF samples were found to be abnormal using TLC (confirmed by UV and HPLC)** *Roy, et al. Bull WHO 1997; 75:19-22 †Kenyon el al. J AOAC Intl 1998; 81: 44-50 **Laserson et al, 2000, unpublished data

Methods: Sample Collection: 

Methods: Sample Collection A convenience sample of TB drugs was collected from various African countries and the Indian government central medical stores, central hospitals, national quality control laboratory storerooms, and during inspections at premises not licensed to sell such drugs Drugs were collected as single units and in fixed-dose-combinations (FDC’s) Collection included different lots/manufacturers available

Methods: Sample Recording: 

Methods: Sample Recording Lot number, manufacturer, and expiration date recorded for each TB drug TB drugs were coded by CENQAM staff Analysis was done blind to country origin of samples

Methods: Procedures: 

Methods: Procedures Participants performed TLC on INH, RIF, PZA, ETH, STR, on single and FDC tablets and capsules CENQAM re-tested all single-dose rifampicin samples and isoniazid samples quantitatively using UV For interest sake, CENQAM tested 3 rifampicin tablets were using dissolution testing

Methods: Materials : 

Methods: Materials The TLC kit Solvents Standards UV light (hand-held battery-operated)

Performing the TLC Analysis:Procedures: 

Performing the TLC Analysis: Procedures Preparation of sample Preparation of standards Preparation of developer solvent (mobile phase) Plate marking Spotting of the plate Development of plate Visualization and interpretation Calculations of Rf values Acceptance and rejection of sample

Results: 

Results 7 countries sampled: India, Kenya, Malawi, Mozambique, Tanzania, Zambia, Zimbabwe

Descriptive Results:Country Results : 

Descriptive Results: Country Results Country Total Analyzed Pass Fail India 16 8 7 1 Kenya 8 4 4 0 Malawi 2 0 0 0 Mozambique 12 3 3 0 Tanzania 38 12 12 0 Zambia 23 3 3 0 Zimbabwe 10 4 4 0 Total 109 34 33 1 Analyzed: 34/109 = 31.2% Passed: 33/34 = 97.1 % Failed: 1/34 = 2.9 %

Descriptive Results: Drug Types: 

Descriptive Results: Drug Types Type Total Analyzed TLC Results Single INH 13 13 13 passed RIF 17 17 16 passed PZA 24 1 1 passed ETH 29 0 _____ STR 13 0 _____ FDC 2-drug [R/I] 13 1 1 passed 2-drug [E/I] 1 1 1 passed 3-drug[R/I/P] 2 2 2 passed

TLC Performance: 

Sensitivity: 0/0 = 100% [Can’t assess] Specificity: 29/30 = 97% TLC Performance

TLC Reproducibility: 

TLC Reproducibility Reproducibility = (0 + 13)/ 14 = 93%

Dissolution Testing: 

Dissolution Testing 3 Rifampicin tested (only 1 unit tested [6 units are required]): Mozambique (1) India (2) The RIF from Mozambique failed The 2 RIF from India passed (??-further testing may be required)

Conclusions: 

Conclusions TLC Method Works well for initial screening of TB drugs Sensitive and specific Very versatile: can be used in the laboratory or the field (no electricity needed) Fast, economical Reliable/ reproducible Available for other drugs Method modification and development is easy

Conclusions: 

Conclusions African and Indian TB drugs By TLC, the tested samples were found to comply with the label claim within the range of 85%-115%, with 1 exception By UV, the tested samples were found to lie within the same range By dissolution, we found 1 of 3 tablets failed Based on our testing, we discovered that the tested TB drugs may be of good strength (TLC + UV) but may not be bioavailable (dissolution)

Limitations: 

Limitations Sampling of TB drug samples was not representative Small number of drugs tested

Recommendations: 

Recommendations Due to our results, we encourage the use of this method for screening of TB drugs At present many countries do not have official monographs for fixed-dose-combinations, so TLC could be used for these combinations After TLC screening, it will be important to perform dissolution testing WHO should encourage screening of TB drugs and particularly the use of TLC because it is fast, economical, and versatile

Implications: 

Implications Is it better to give the patient nothing or TB drugs which you know that are of low quality? “is half a loaf better than nothing?”

Acknowledgements: 

Acknowledgements Thank you to the CENQAM staff!!!!!!!!! Thank you to CDC, FDA, WHO! Thank you to the instructors!