Metabolism and interactions of antileprosy drugs.

Leprosy is a chronic infectious disease caused my Mycobacterium leprae that primarily affects peripheral nervous system and extremities and is prevalent in tropical countries. Treatment for leprosy with multidrug regimens is very effective compared to monotherapy especially in multibacillary cases. The three major antileprosy drugs currently in use are 4, 4'-diaminodiphenyl sulfone (DDS, dapsone), rifampicin, and clofazimine. During multidrug therapy, the potent antibiotic rifampicin induces the metabolism of dapsone, which results in decreased plasma half-life of dapsone and its metabolites. Furthermore, rifampicin induces its own metabolism and decreases its half-life during monotherapy. Rifampicin upregulates several hepatic microsomal drug-metabolizing enzymes, especially cytochrome P450 (CYP) family that in turn induce the metabolism of dapsone. Clofazimine lacks significant induction of any drug-metabolizing enzyme including CYP family and does not interact with dapsone metabolism. Rifampicin does not induce clofazimine metabolism during combination treatment. Administration of dapsone in the acetylated form (acedapsone) can release the drug slowly into circulation up to 75 days and could be useful for the effective treatment of paucibacillary cases along with rifampicin. This review summarizes the major aspects of antileprosy drug metabolism and drug interactions and the role of cytochrome P450 family of drug metabolizing enzymes, especially CYP3A4 during multidrug regimens for the treatment of leprosy.

Novel electrochemical biosensor based on PVP capped CoFe2O4@CdSe core-shell nanoparticles modified electrode for ultra-trace level determination of rifampicin by square wave adsorptive stripping voltammetry.

This work introduces a new electrochemical sensor based on polyvinyl pyrrolidone capped CoFe2O4@CdSe core-shell modified electrode for a rapid detection and highly sensitive determination of rifampicin (RIF) by square wave adsorptive stripping voltammetry. The new PVP capped CoFe2O4@CdSe with core-shell nanostructure was synthesized by a facile synthesis method for the first time. PVP can act as a capping and etching agent for protection of the outer surface nanoparticles and formation of a mesoporous shell, respectively. Another important feature of this work is the choice of the ligand (1,10-phenanthroline) for precursor cadmium complex that works as a chelating agent in order to increase optical and electrical properties and stability of prepared nanomaterial. The nanoparticles have been characterized by field emission scanning electron microscopy (FESEM), transmission electron microscope (TEM), energy dispersive X-ray spectroscopy (EDX), X-ray diffraction (XRD), UV-vis, photoluminescence (PL) spectroscopy, FT-IR, and cyclic voltammetry techniques. The PL spectroscopy study of CoFe2O4@CdSe has shown significant PL quenching by the formation of CoFe2O4 core inside CdSe, this shows that CoFe2O4 NPs are efficient electron acceptors with the CdSe. It is clearly observed that the biosensor can significantly enhance electrocatalytic activity towards the oxidation of RIF, under the optimal conditions. The novelty of this work arises from the new synthesis method for the core-shell of CoFe2O4@CdSe. Then, the novel electrochemical biosensor was fabricated for ultra-trace level determination of rifampicin with very low detection limit (4.55×10-17M) and a wide linear range from 1.0×10-16 to 1.0×10-7M. The fabricated biosensor showed high sensitivity and selectivity, good reproducibility and stability. Therefore, it was successfully applied for the determination of ultra-trace RIF amounts in biological and pharmaceutical samples with satisfactory recovery data.

Bone distribution study of anti leprotic drug clofazimine in rat bone marrow cells by a sensitive reverse phase liquid chromatography method.

The aim of the present study was to investigate the distribution of clofazimine (CLF) in rat bone marrow cells by a validated reverse phase high performance liquid chromatography. CLF and chlorzoxazone (I.S) were extracted by liquid-liquid extraction from plasma and rat bone marrow cells. The chromatographic separation was performed in isocratic mode by the mobile phase consisting of 10mM ammonium formate (pH 3.0 with formic acid) and acetonitrile in a ratio of 50:50 (v/v). The method was accurate and precise in the linear range of 15.6-2000.0 ng/mL with a correlation coefficient (r(2)) of 0.996 and 0.995 in rat plasma and bone marrow cells, respectively. After single oral dose of 20mg/kg, the maximum concentration of CLF in plasma and bone marrow cells were obtained at 12h with the concentrations of 593.2 and 915.4 ng/mL, respectively. The AUC0-t and mean elimination half life (t1/2) of CLF in bone marrow cells were 54339.02 ng h/mL and 52.46 h, respectively, which signified the low body clearance and high distribution of CLF in bone marrow cells. The single oral dose pharmacokinetic investigation was confirmed the CLF endure for a long period in rat due to high distribution in various tissues. The developed method was successfully applied to the estimation of the pharmacokinetic parameters of CLF in plasma and bone marrow cells after administration of single oral dose of 20mg/kg to rats.

Dapsone and body mass index in subjects with multibacillary leprosy.

BACKGROUND: The physiological changes in obese subjects can modify the pharmacokinetic profiles of drugs influencing the therapeutic efficacy. METHODS: In this study, the authors compare plasma dapsone trough levels of multibacillary leprosy subjects stratified by body mass index (BMI) to evaluate if obesity plays a significant role on drug levels. The relationship between drug levels and BMI was also determined. Dapsone was measured by high-performance liquid chromatography and BMI based on World Health Organization criteria. RESULTS: At steady state, the median plasma dapsone trough level was significantly lower in obesity class 2 group, when compared with other groups, but they were similar between normal weight and preobesity groups. A weak association between drug levels and BMI was observed. CONCLUSIONS: Obesity promotes a significant reduction in plasma dapsone trough levels of subjects with multibacillary leprosy with a weak association between drug levels and BMI.

Safety and tolerability of dapsone for the treatment of patients with drug-resistant, partial-onset seizures: an open-label trial.

Dapsone has shown anti-convulsive properties in animal models of epilepsy. In the present study, we tested the safety and tolerability of dapsone as adjunctive therapy in adult patients with drug-resistant partial-onset seizures. Twenty-two adult patients with drug-resistant partial-onset seizures were included. After a 3-month baseline period, patients received dapsone 100 mg per day, for a 3-month evaluation period. Plasma concentrations of anti-epileptic drugs (AEDs) did not significantly change during the study. No alteration of mean clinical laboratory values was observed. The reported adverse events were: mild methemoglobinemia (50%), headache (31.8%), paleness (27.3%) and somnolence (4.5%).Sixteen of 22 patients reduced their seizure frequency in more than 50% as a result of dapsone treatment. Three subjects remained seizure-free during the entire dapsone treatment period. This open-label study of adjunctive dapsone therapy at 100 mg/day suggests that dapsone is safe, and well-tolerated in adults with drug-resistant partial-onset seizures.

[Thalidomide used by patients with erythema nodosum leprosum]. / Talidomida usada por pacientes com eritema nodoso hansênico.

INTRODUCTION: Thalidomide is a drug currently used in Brazil for treating erythema nodosum leprosum. METHODS: This was a prospective study to follow up clinical evolution, record adverse events and determine plasma thalidomide levels from a dose of 100 mg/day, among 20 patients with clinical manifestations of erythema nodosum leprosum, divided into two groups: during or after leprosy multidrug therapy. RESULTS: No significant differences between the groups were seen during the study, either in relation to favorable clinical evolution among the patients (70% and 90%), or in relation to the adverse events recorded, which were dizziness and somnolence. The plasma thalidomide levels on D7 and D14 were 0.82 + or - 0.4 microg/ml and 0.79 + or - 0.3 microg/ml in group 1 and 0.82 + or - 0.4 and 1.55 + or - 1.0 in group 2, respectively. CONCLUSIONS: In this sample, the multidrug therapy had no effect on the clinical evolution, incidence of adverse events and plasma thalidomide levels.

Talidomida usada por pacientes com eritema nodoso hansênico / Thalidomide used by patients with erythema nodosum leprosum

INTRODUÇÃO: A talidomida é um fármaco utilizado atualmente no tratamento do eritema nodoso hansênico no Brasil. MÉTODOS: Estudo prospectivo para acompanhar a evolução clínica, registrar os eventos adversos e determinar as concentrações plasmáticas de talidomida em dose diária de 100mg/dia, em 20 pacientes com manifestações clínicas de eritema nodoso hansênico, divididos em dois grupos: após ou em curso da poliquimioterapia para hanseníase. RESULTADOS: Não foram observadas diferenças significativas nos grupos no decorrer do estudo, tanto na evolução clínica favorável dos pacientes, de 70 por cento e 90 por cento, quanto nos eventos adversos registrados que foram tontura e sonolência. Os teores plasmáticos de talidomida em D7 e D14 foram de 0,82±0,4μg/mL e 0,79±0,3μg/mL no grupo 1 e de 0,82±0,4 e 1,55±1,0 no grupo 2, respectivamente. CONCLUSÕES: Na amostra estudada, a poliquimioterapia não interferiu na evolução clínica, na incidência dos efeitos adversos e nos níveis plasmáticos de talidomida.

INTRODUCTION: Thalidomide is a drug currently used in Brazil for treating erythema nodosum leprosum. METHODS: This was a prospective study to follow up clinical evolution, record adverse events and determine plasma thalidomide levels from a dose of 100 mg/day, among 20 patients with clinical manifestations of erythema nodosum leprosum, divided into two groups: during or after leprosy multidrug therapy. RESULTS: No significant differences between the groups were seen during the study, either in relation to favorable clinical evolution among the patients (70 percent and 90 percent), or in relation to the adverse events recorded, which were dizziness and somnolence. The plasma thalidomide levels on D7 and D14 were 0.82 ± 0.4μg/ml and 0.79 ± 0.3 μg/ml in group 1 and 0.82 ± 0.4 and 1.55 ± 1.0 in group 2, respectively. CONCLUSIONS: In this sample, the multidrug therapy had no effect on the clinical evolution, incidence of adverse events and plasma thalidomide levels.

Thalidomide levels in patients with erythema nodosum leprosum.

Thalidomide is used for the acute treatment and suppression of the cutaneous manifestations of erythema nodosum leprosum (ENL). In this study, comparisons were made regarding the plasma concentrations of thalidomide in patients with ENL on the course or after leprosy therapy in a prospective clinical trial. Thalidomide concentrations were measured by liquid chromatography on days 1, 3, and 14 of treatment. After 100 mg/d, the thalidomide concentrations ranged from 0.82 to 1.03 and 0.43 to 0.80 microg/mL, on the course or after leprosy therapy, respectively. No differences were observed in thalidomide concentrations between and within the groups. Our results suggested that leprosy multidrug therapy does not seem to affect the plasma concentrations of thalidomide in patients with ENL.

Antileprosy activity and preliminary toxicological characteristics of some dialkyldithiocarbamate derivatives.

Antileprosy activity of dialkyldithiocarbamate derivatives was studied in experiments on mice infected with M. leprae into paw pads. We found that 2-diethyldithiocarbamoyl-3-cyano-5-nitropyridine is the most promising antileprosy agent; it effectively suppresses multiplication of M. leprae and is well tolerated under conditions of chronic animal experiment.

Pharmacokinetics and relative bioavailability of clofazimine in relation to food, orange juice and antacid.

BACKGROUND: Clofazimine is potentially useful for the treatment of disease due to multidrug resistant Mycobacterium tuberculosis, as well as leprosy and certain chronic skin diseases. Its pharmacokinetics have been incompletely characterized. This study was conducted to explore issues relating to bioavailability in the presence of food, orange juice, and antacid. METHODS: A 5 drug regimen consisting of clofazimine, cycloserine, ethionamide, para-aminosalicyclic acid, and pyridoxime was administered to healthy subjects four times using a four period cross-over design with two weeks washout between treatments. Subjects also received orange juice, a high fat meal, aluminum/magnesium antacid, or only water in random order with the drug regimen. The pharmacokinetics of clofazimine were assessed using individual- and population-based methods and relative bioavailability compared to fasting administration was determined. RESULTS: Clofazimine exhibited a sometimes prolonged and variable lag-time and considerable variability in plasma concentrations. From the population analysis (one-compartment model), the mean oral clearance was 76.7 l/h (CV=74.2%) and mean apparent volume of distribution was 1470 l (CV=36.3%). The first-order absorption rate constant ranged from 0.716 to 1.33 h(-1) (pooled CV=61.7%). Residual (proportional) error was 49.1%. Estimates of bioavailability compared to fasting administration were 145% (90% CI, 107-183%) for administration with high fat food, 82.0% (63.2-101%) for administration with orange juice, and 78.5% (55.1-102%) for administration with antacid. CONCLUSION: Administration of clofazimine with a high fat meal provides the greatest bioavailability, however, bioavailability is associated with high inter- and intra-subject variability. Both orange juice and aluminum-magnesium antacid produced a reduction in mean bioavailability of clofazimine.

Liquid chromatography/tandem mass spectrometric bioanalysis using normal-phase columns with aqueous/organic mobile phases - a novel approach of eliminating evaporation and reconstitution steps in 96-well SPE.

Bioanalytical methods using automated 96-well solid-phase extraction (SPE) and liquid chromatography with electrospray tandem mass spectrometry (LC/MS/MS) are widely used in the pharmaceutical industry. SPE methods typically require manual steps of drying of the eluates and reconstituting of the analytes with a suitable injection solvent possessing elution strength weaker than the mobile phase. In this study, we demonstrated a novel approach of eliminating these two steps in 96-well SPE by using normal-phase LC/MS/MS methods with low aqueous/high organic mobile phases, which consisted of 70-95% organic solvent, 5-30% water, and small amount of volatile acid or buffer. While the commonly used SPE elution solvents (i.e. acetonitrile and methanol) have stronger elution strength than a mobile phase on reversed-phase chromatography, they are weaker elution solvents than a mobile phase for normal-phase LC/MS/MS and therefore can be injected directly. Analytical methods for a range of polar pharmaceutical compounds, namely, omeprazole, metoprolol, fexofenadine, pseudoephedrine as well as rifampin and its metabolite 25-desacetyl-rifampin, in biological fluids, were developed and optimized based on the foregoing principles. As a result of the time saving, a batch of 96 samples could be processed in one hour. These bioanalytical LC/MS/MS methods were validated according to "Guidance for Industry - Bioanalytical Method Validation" recommended by the Food and Drug Administration (FDA) of the United States.

High-performance liquid chromatographic method with ultraviolet detection for the determination of dapsone and its hydroxylated metabolite in human plasma.

A validated high-performance liquid chromatographic method with ultraviolet detection for the quantitative determination of dapsone (4,4'-diaminodifenyl sulfone, DDS) and a metabolite, hydroxylaminodapsone (4-amino-4-hydroxylaminodiphenyl sulfone, DDS-NOH), in human plasma is described. Human plasma was deproteinized with acetone and the clear supernatant solution after centrifugation was evaporated to dryness under a gentle stream of nitrogen at 70 degrees C. The residue was dissolved in a mixture of HPLC eluent and acetone (18:5 v/v) and an aliquot of this solution (50 microL) was injected onto the HPLC column. Dapsone, hydroxylaminodapsone and diazoxide as internal standard, were separated within 10 min by isocratic elution with water:acetonitrile:glacial acetic acid:triethylamine (80:20:1.0:0.5 by volume) as eluent. Detection was by ultraviolet at the wavelength of 295 nm. The within-day repeatability coefficients of variation were 3-5% for dapsone (0.301-20.0 mg/L, n = 5) and 3-5% for hydroxylaminodapsone (0.0948-6.32 mg/L, n = 5), whereas the between-day repeatability coefficients of variation were 3-8% (0.301-20.0 mg/L, n = 5) for dapsone and 4-10% for hydroxylaminodapsone (0.0948-6.32 mg/L, n = 5). The mean recoveries -were 92-107% (0.301-20.0 mg/L, n = 2), 80-82% (0.0948-6.32 mg/L, n = 2) and 88% (0.0200 mg/mL, n = 5), for dapsone, hydroxylaminodapsone and diazoxide, respectively. The average correlation coefficient of the calibration curve was 0.99988 (n = 5) for dapsone at a concentration range of 0.301-20.0 mg/L, whereas the average correlation coefficient of the hydroxylaminodapsone calibration curve was 0.99981 (n = 5) at a concentration range of 0.0948-6.32 mg/L. The limits of detection were 0.00200 and 0.0470 mg/L for dapsone and hydroxylaminodapsone, respectively. The method is suitable for drug level monitoring and for pharmacokinetic studies.

Thalidomide dose proportionality assessment following single doses to healthy subjects.

Thalidomide is approved in the United States for treating erythema nodosum leprosum, a complication of leprosy. The present study determined the single-dose oral pharmacokinetics and dose proportionality from 50 to 400 mg of Celgene's commercial Thalomid thalidomide formulation in an open-label, single-dose, three-way crossover study. Fifteen healthy subjects were given 50, 200, and 400 mg of thalidomide on three occasions, and blood samples were collected over 48 hours. Pharmacokinetic parameters were determined using noncompartmental methods, and dose proportionality was assessed by linear regression of dose-normalized Cmax and AUC0-infinity. No serious or unexpected adverse events occurred. The most common adverse events were dizziness, somnolence, headache, and nausea. One patient was discontinued because of pharyngitis. There was a significant deviation from proportionality for Cmax with increases being less than proportional than changes in dose. AUC0-infinity increased proportionally with dose, suggesting that the overall amount of thalidomide absorbed, as well as its clearance, is independent of dose over the range used. V/F was found to increase with dose. This was most likely due to the terminal rate constant, which is used to calculate V/F, actually representing the absorption process rather than elimination (i.e., flip-flop phenomenon). The terminal rate constant (absorption rate constant) for the highest dose was 50% less than for the other two lower doses. The less than proportional increases in Cmax were most likely due to thalidomide's low aqueous solubility. Thalidomide shows reasonable dose proportionality with respect to AUC from 50 to 400 mg.

Single-dose oral pharmacokinetics of three formulations of thalidomide in healthy male volunteers.

Thalidomide was recently approved in the United States for the treatment of erythema nodosum leprosum, a complication of leprosy. The present study determined the bioequivalence and pharmacokinetics of Celgene's commercial and clinical trial thalidomide formulations and the Brazilian Tortuga formulation in an open-label, single-dose, three-way crossover design. Seventeen healthy subjects were given 200 mg of thalidomide on three occasions, and blood samples were collected over 48 hours. Pharmacokinetic parameters were determined using compartmental methods for the two Celgene formulations and using noncompartmental methods for all three formulations. All subjects reported adverse events, none of which was serious or unexpected. Celgene formulations were bioequivalent when comparing Cmax, tmax, and AUC. There was significant variability in plasma levels from the Tortuga formulation, giving a mean profile that was distinctly different from the two Celgene formulations with a lower Cmax value and a longer terminal phase. The lower Cmax was probably due to slower absorption. The terminal rate constant for the Tortuga formulation was significantly less, giving rise to a terminal half-life of 15 hours compared to about 5 to 6 hours for the Celgene formulations. Confidence intervals for Cmax between the Tortuga and the Celgene formulations were outside the 80% to 125% range, indicating a lack of bioequivalence. Extent of absorption, as measured by AUC0-infinity, was approximately equal for all three formulations. Terminal half-life for Tortuga was two to three times longer compared to the Celgene formulations and is clear evidence for absorption rate limitations. The two Celgene formulations showed similar pharmacokinetic parameters with profiles that were best described by a one-compartment model with first-order absorption and elimination. The authors conclude that Celgene's clinical trial and commercial thalidomide formulations are similar to each other and distinctly different from the Tortuga formulation and that all three formulations exhibited absorption rate-limited elimination.

HPLC determination of clofazimine in tissues and serum of mice after intravenous administration of nanocrystalline or liposomal formulations.

A simple HPLC method is described for the determination of clofazimine in mouse tissues and in serum. The main application of the method was the determination of the drug in mouse tissues after i.v. administration of nanocrystalline suspensions or liposomal encapsulated clofazimine. Tissues were extracted with a 10-fold (w/v) volume of an extraction solution consisting of methanol/glacial acetic acid 9:1 (v/v). Serum proteins were precipitated with a 2-fold volume of acetonitrile. Isocratic chromatography was performed using an anion exchange column (Nucleosil 100-5 SA, Macherey & Nagel) for separation. The mobile phase was a mixture of acetonitrile and 0.1 mol/l aqueous phosphoric acid (75:25, v/v), adjusted to pH 2.9 with sodium hydroxide solution. Absorption of the eluate was monitored at 495 nm. The assay was precise, simple to perform and fast. Recovery from tissues was > or = 98%, from nanoparticles > or = 98%, and from liposomes > or = 96%. No interference was observed in extracts from mouse liver, spleen, lungs and human serum.

Combined dapsone and clofazimine intoxication.

We report clinical findings and pharmacokinetic data regarding a combined dapsone and clofazimine intoxication in a man, who ingested 50 tablets of dapsone (100 mg) 20 capsules of clofazimine (100 mg) and two tablets of rifampicin (600 mg). Oral administration of activated charcoal (50 grams) and sodium sulphate (20 grams) after gastric lavage resulted in an elimination half-life in plasma of 11.1 and 10.8 h for dapsone and its main metabolite, monoacetyldapsone, respectively. A rapid initial decrease of the plasma concentration of clofazimine was observed after gastric lavage and administration of activated charcoal and sodium sulphate. 15 h after this treatment, clofazimine plasma levels remained relatively constant. Dapsone-induced methaemoglobinaemia (48% at admission) was treated successfully with methylene blue.

Determination of serum and tissue levels of phenazines including clofazimine.

A rapid and sensitive HPLC method is described for the analysis of synthetic phenazines, including clofazimine, from a variety of biological samples. Phenazines were extracted from serum, tissue and fat using a mixture of dichloromethane and sodium hydroxide. The drugs were then quantified on a reversed-phase C18 column using a mobile phase consisting of 594 ml of water, 400 ml of tetrahydrofuran, 6 ml of concentrated acetic acid and 0.471 g of hexanesulfonic acid. In this mobile phase, each phenazine tested had its own retention time. This allowed one phenazine to be used as an internal standard for the analysis of other phenazines. The method was validated for clofazimine [3-(4-chloroanilino)-10-(4-chlorophenyl)-2,10-dihydro-2-(isopro pylimino) phenazine] and B4090 [7-chloro-3-(4-chloranilino)-10-(4-chlorophenyl)-2, 10-dihydro-2-(2,2,6,6-tetramethylpiperid-4-ylimino)phenazine ] (VI) and shown to be accurate and precise across a broad concentration range from 0.01 to 50 micrograms/g (microgram/ml). Extraction was 100% for each agent across this range. This system was used to measure clofazimine and VI levels following their administration to rats. The pharmacokinetic profile of VI was different to that of clofazimine, with high tissue concentrations but lower fat levels.

A case of acute dapsone poisoning: toxicological data and review of the literature.

A case of nonfatal, acute poisoning following the ingestion of an undetermined amount of dapsone (DDS) in a 49-year-old woman is presented. The clinical features were dyspnea and deep cyanosis. Methemoglobinemia was 39.0% on admission. DDS was identified and quantitated in blood samples taken on the third day (sample A) and fifth day (sample B) in the hospital using a high-performance liquid chromatographic technique with diode-array detection. DDS concentrations were 26.99 and 8.40 micrograms/mL in samples A and B, respectively. Results are discussed in the light of an extensive review of the literature (1950-1993) available on DDS poisonings.

High-performance liquid chromatographic determination of rifapentine in serum using column switching.

A high-performance liquid chromatographic method with column switching has been developed for the determination of rifapentine in serum. The serum samples were injected onto a precolumn packed with Corasil RP C18 (37-50 microns) after simple dilution with an internal standard in a 1% ascorbic acid solution. Polar serum components were washed out using 0.05 M phosphate buffer. After valve switching, the concentrated drugs were eluted in the back-flush mode and separated by a mu Bondapak C18 column with acetonitrile-tetrahydrofuran-0.05 M phosphate buffer (pH 7.0) (42:5:53, v/v/v) as the mobile phase. The method showed excellent precision with good sensitivity and speed, and a detection limit of 0.1 microgram/ml. The total analysis time was less than 25 min and the mean coefficients of variation for intra- and inter-assay were less than 4.8%. The method has been successfully applied to serum samples from dogs after the oral administration of rifapentine.

Determination of three main antileprosy drugs and their main metabolites in serum by high-performance liquid chromatography.

The simultaneous analysis of main antileprosy drugs such as 4,4'-diaminodiphenyl sulfone (DDS), clofazimine, rifampicin and their main metabolites in serum was examined by high-performance liquid chromatography using a muBondapak C18 column. When the drugs dissoluted from serum were developed by tetrahydrofuran-0.5% acetic acid (40:60), clofazimine and rifampicins could be analyzed separately. Apart from the mutual separation of water-soluble conjugates of DDS, the individual analysis of DDS, its main liposoluble metabolite and a few related sulfone compounds is possible when the drugs are first developed by acetonitrile-water (20:80). By the use of tetrahydrofuran-water (50:50) containing PIC B-5, the rapid measurement of clofazimine isolated from the other compounds is also possible.