OBJECTIVES

In response to the need for a simple and fast way of ensuring that generic drugs especially those that contain rifampicin are bioequivalent with reference drugs, this study validated an ultra-high-performance liquid chromatography (UHPLC) method of quantifying rifampicin in human plasma. The study also validated the method's selectivity, linearity, sensitivity, accuracy, precision, and the absence of a carry-over effect adhering to the Philippine Food and Drug Administration guidelines.

Plasma samples were prepared via protein precipitation using methanol containing ascorbic acid. Three microliters (3 uL) of the prepared samples were then analyzed in a Waters Acquity H-Class UPLC® system coupled to a tunable ultraviolet (TUV) detector with an attached UPLC® BEH C-18 column using a developed and optimized method. Briefly, the column temperature was set to 40°C and the sample temperature was set to 10°C. Elution was done using a linear gradient flow of a water-acetonitrile mixture that started with 45% acetonitrile increasing to 60% acetonitrile at 0.5 minutes and back to 45% acetonitrile at 3 minutes and having a constant flow rate of 0.5 mL/min. Detection was done at 340 nm. Method validation was performed following the ICH guidelines for Bioanalytical Method Validation, the same guidelines referenced by the ASEAN Guideline for Harmonisation of Standards and the Philippine Food and Drug Administration (FDA).

The method had an analysis time of 3 minutes wherein rifampicin eluted at 1.4 minutes while the internal standard, rifapentine (IS) eluted at 1.7 minutes. Since no co-eluting endogenous materials were observed for the rifampicin and the internal standard, the method was confirmed to be selective. Its linearity over the range of 2 ug/mL to 25 ug/mL has been validated where it has a limit of detection (LOD) and limit of quantification (LOQ) values of 0.64 ug/mL and 1.94 ug/mL, respectively. The interday and intraday precision, reported as % coefficient of variance (%CV), and interday and intraday accuracy, reported as %error all within the limits of ±20% for the LLOQ and ±15% for the rest indicating its reliability and reproducibility. Lastly, due to the nature of the injection of the sample into the system, wherein a blank immediately follows the highest concentration standard, the method has been cleared of a carry-over effect.

The study successfully validated a UHPLC method of quantifying rifampicin in human plasma. Due to the sample processing method used and the chromatographic conditions set, the method can prepare and analyze samples in a simple yet fast, sensitive, reliable, and reproducible manner. The method can be applied in bioavailability and bioequivalence studies of rifampicin.

Background: Rifampicin, isoniazid, and pyrazinamide are oral essential anti-tuberculosis drugs on single or combined preparations. Worldwide research has shown that the plasma concentration of anti-tuberculosis drugs with daily therapeutic doses were seen significant lower than permitted in tuberculosis patients, especially for rifampicin and isoniazid. Objective: To investigate plasma concentration of rifampicin, isoniazid, and pyrazinamide in pulmonary tuberculosis and pleural tuberculosis patients. Methods: Determine plasma concentration of rifampicin, isoniazid, and pyrazinamide at 2 hours after administration in 168 tuberculosis patients by the HPLC method. Identify prevalence of low plasma concentrations of anti-tuberculosis drugs. Results: There was a wide range of plasma concentration of rifampicin, isoniazid, and pyrazinamide in the tuberculosis patients. The mean plasma concentration of rifampicin was 6.13 \xb1 4.66 microgram/ml, of isoniazid was 2.99 \xb1 1.94 microgram/ml, pyrazinamide was 38.98 \xb1 18.39 microgram/ml. There was no significant differences in the plasma concentration of rifampicin, isoniazid, and pyrazinamide in groups of pulmonary tuberculosis and pleural tuberculosis patients. Percentage of patients with plasma concentration below therapeutic concentration was 76.83% of rifampicin, 51.85% of isoniazid, 10.13% of pyrazinamide. There were 12.03% of patients who had pyrazinamide concentration higher than the therapeutic range. Conclusions: There was a wide range of plasma concentration in rifampicin, isoniazid, and pyrazinamide of tuberculosis patients. Low plasma concentration of rifampicin and isoniazid are common. It may be necessary to optimize the drug dose by therapeutic drug monitoring, especially in patients with an inadequate clinical response to chemotherapy.
tuberculosis ; rifampicin ; isoniazid ; pyrazinamide

Introduction: Mycobacterium tuberculosis resists rifampicin (RIF) because of mutations in the rpoB (the p subunit of RNA polymerase) gene, mostly in the 81 bp region. \r\n', u'Objectives: Identify the frequency and characteristics relative to drug - resistant rpoB gene mutation in RIF - resistant M. tuberculosis strains. \r\n', u'Subjects and method: 40 M. tuberculosis strains including 11 RIF - sensitive strains and 29 RIF - resistant strains. Some bio molecular techniques were used such as extracting mycobacterial DNA, PCR, cloning, sequencing and analyzing mutation related RIF - resistance on rpoB gene. \r\n', u'Results: No mutation was found on the 81 bp region of rpoB gene of the RIF - sensitive M. tuberculosis strains. The rate of mutation on rpoB gene of 29 RIF - resistant M. tuberculosis strains is 96.6%. We found 12 mutation codon positions on the 81 bp region of the rpoB gene, and the mutation codon positions with high frequency were 531 (51.7%) and 526 (31%). The mutation position found in only one strain is codon 519 (3.4%) but not found in other reports. There are 15 types of drug resistant mutations in which TCG531 TCG is the most common with 50%. Multi - drug resistance was seen in mutable and none mutable cases, with all codon positions and mutable forms. \r\n', u'Conclusion: No mutation was found on the 81 bp region of the rpoB gene of RtF - sensitive M. tuberculosis strains. The rate of mutation on the rpoB gene of RIF - resistant M. tuberculosis strains is 96.6%. The new mutation position found is codon 519. The mutation on the rpoB gene does not determine the multi - drug resistance of M. tuberculosis. \r\n', u'
Mutation ; rpoB gene ; Rifampicin - resistant M. tuberculosis

Primary study showed hypoglycemic effect of gynostema pentaphyllum (GP) on mice. Objective: (I) To determine the hypoglycemic effect of (GP) and (2) To look for effective mechanism of GP on normal and diabetic mice and rats. Method: The ethanol extract of GP were introdiced to normal and pre diabetic & diabectic mice or rats. The blood glucose levels were measured at just (0h) before and at 2h, 4h and 6h after oral use or 1h, 2h and 3h after intraperitoneal use of GP. Results: On normal mice, 300 mg/kg-ip of GP decreased the blood glucose level with max. 33% at 3rd hour; 1500 mg/kg-po. with 20% at 6th hour; the dose of 500 mg/kg x 7 days consecutive decreased this level with 24%. On normal Wistar rats 500 mg/kg-po, diminished glycemia with max. 20% at 4th hour. But on genetic diabetic rats (GK rats), the doses 500 mg/kg-po, and 1000 mg/kg-po, decreased the blood glucose concentrations by 22% and 36%, respectively at the same time. In the glucose toleance test on mice the dose 1000 mg/kg-po, inhibited the hyperglycemic effect with 55% (after 30 minutes) and 63% (after 60 minutes) in comparison to control group. Conclusion: GP has the mild hypoglycemic effect on normal mouse/rat, but it causes more higher effect on mouse/rat having hyperglycemia. Thus, beside the insulin secreting stimulation, GP may sensitive the target tissues to insulin.
Tuberculosis, Rifampin

No abstract available.
Purpura, Thrombocytopenic* ; Rifampin*

Report one case of a 45 years old male patient lived in Hoc Mon district, Ho Chi Minh City, and admitted in November 24th 2004. His tuberculosis (TB) was first diagnosed in the year 2000 and he was received anti-TB drugs with 2SHR/6HE regime at the anti-TB station. During hospitalization, after 10 days of SHRZE regime (SM 1/2g, Rif 300mg PO, INH 200mg PO, PZA 750mg PO, EMB 600mg PO), he experienced fatigue, dyspnea, jaundice, fever of 39 degree C, blood pressure: 8/5cmHg, SpO2 = 90%. Hematological analysis revealed low hemoglobin and Hct levels, and increase of reticulocyte count and white blood cell count; a positive direct Coombs test. Biochemical tests revealed elevated total, direct and indirect bilirubin levels, increased LDH; decreased haptoglobin; LE cell and ANA were all negative. Patient was diagnosed immune haemolytic anemia and was treated by blood transfusion with the same blood group. He discharged at December 17th 2004 with the last diagnosis was recurrent tuberculosis M(+), rifampicin-induced immune haemolytic anemia
Tuberculosis ; Rifampin ; Therapeutics

In this prospective study, 56 re-treatment pulmonary tuberculosis patients with smear-positive were examined to investigate possible variations in plasma rifampicin concentration using fixed dose combination drugs. 2-hour and 3-hour plasma rifampicin concentration were measured by HPLC method among 56 smear-positive re-treatment pulmonary tuberculosis patients. Results: Plasma rifampicin concentrations were generally low: 2-hour and 3-hour plasma rifampicin concentration below 8mg/ml (93% and 86% of patients, respectively), 55% and 46% of patients had plasma concentration below 4mg/ml (at 2-hour and 3-hour time point, respectively). Although treatment at the same dose levels, there is high variable between individual patients in plasma rifampicin concentration. Plasma rifampicin concentrations at 3 hours after dosed were higher than at 2 hours in most of patients.
Rifampin ; Tuberculosis ; Pulmonary

57 rifampicin-resistant tuberculosis strains isolated from Ha Noi Institute of Tuberculosis and Lung Diseases and Pham Ngoc Thach Tuberculosis Center of Ho Chi Minh City were involved in this study. The regimen included 2-month SHRZ and 4-month HR. All of these strains were resistant to rifampicin. It was found that rifampicin-resistant tuberculosis strains had mutations on rpoB gene. The mutation frequency of tuberculosis trains isolated from Ha Noi and Ho Chi Minh City was highest on 526 locus (55% and 41.6%, respectively) and 531 locus (30% and 33%, respectively).
Tuberculosis ; Mutation ; Rifampin

Study on 12 healthy volunteers using 2 types of medication: rifampicin, isoniazid, and pyrazinamid standard separate tablets and in fixed dose combination of rifampicin + isoniazid + pyrazinamid (3- FDC). Results: rifampicin is one of essential anti-tuberculosis drugs that have most advantages of pharmacodynamics on both intra- and extra-cellular bacteria. Cmax (maximum concentration in plasma) and AUC 0-∞ (Area Under the Concentration-Time Curve) of rifampicin in 3- FDC is lower than that in standard separate tablet, Tmax (Time to Maximum Plasma Concentration) of rifampicin in 3- FDC is more slowly than in standard separate tablet. These findings showed that the bioavailability of rifampicin in FDC tablet that was using in treating tuberculosis is much lower than in standard separate tablet.
Biological Availability ; Rifampin ; Tablets

No abstract available.
Nephritis, Interstitial ; Rifampin