Liquid chromatographic determination of CPZEN-45, a novel anti-tubercular drug, in biological samples

Abstract

CPZEN-45 is a new drug candidate being considered for the treatment of tuberculosis (TB). The aim of this study was to develop and validate a reverse-phase high-performance liquid chromatographic (HPLC) method suitable to determine CPZEN-45 concentrations in biological samples. CPZEN-45 was extracted from biological fluids and tissues (plasma, lung and spleen from guinea pig) by sequential extraction with acetonitrile and quantified by a Waters HPLC Alliance System coupled with a ZORBAX Bonus-RP column, guard column and UV detection at 263 nm. The mobile phase was 20:80 acetonitrile:ultrapure-water with 0.05% TFA. The CPZEN-45 peak was eluted at 5.1 min with no interference from the inherent peaks of plasma, bronchoalveolar lavage (BAL), lung or spleen tissues. Recovery of CPZEN-45 from biological samples was >96% of the spiked amount. The limit of detection was 0.05 µg/ml and the limit of quantitation was 0.29 µg/ml which was more than 5 and 21 times lower than the reported minimal inhibitory concentration of CPZEN-45 (MIC = 1.56 µg/ml for Mycobacterium tuberculosis and 6.25 µg/ml for MDR-TB, respectively). Thus, HPLC method was deemed reliable, sensitive, reproducible and accurate for the determination of CPZEN-45 concentrations in plasma, BAL, lung and spleen tissues. Therefore, this method was used in subsequent studies in the guinea pig model to determine the disposition of CPZEN-45 after administration of solutions by the IV and SC routes.

Electrochemical determination of selected antihypertensive and antituberculosis drugs at a tyrosine-modified electrode

“…Tuberculosis (TB) is caused by Mycobacterium tuberculosis, and is one of the leading causes of adult deaths worldwide, although the number has been decreasing since 2006. 1,2 In order to prevent the development of tuberculosis, isoniazid (ISN), pyrazinamide (PZM) and ethambutol hydrochloride (EBH) have been widely prescribed for the treatment of tuberculosis, along with a xed dose of rifampicin. 3,4 Another leading cause of death in the early 19th century was congestive heart failure due to hypertension, 5 although the death rate was reduced in the 1950s and 1960s by the introduction of antihypertensive drugs.…”

The Pharmacokinetics of CPZEN-45, a Novel Anti-Tuberculosis Drug, in Guinea Pigs

“…After extraction from plasma, BALF, and tissues, CPZEN-45 concentrations were determined by a validated HPLC method [ 12 ]. In plasma samples, the limit of detection (LOD) and limit of quantification (LOQ) for this method were 0.05 μg/mL and 0.29 µg/mL, respectively, whereas in BAL and lung/spleen homogenates, the LOD and LOQ were 0.004 and 0.02 μg/mL, respectively [ 12 ].…”

“…A custom-made nose-only dosing chamber was designed to dose the animals by passive inhalation (Figure 3a,b) [17]. The design of this chamber, including the inhalation area size, the method of aerosol generation from CPZEN-45 powders, and the manner in which powders were introduced into the inhalation area were first optimized [17,20]. The CPZEN-45 dry powder aerosol was generated by passing air from a 10 mL syringe through a "dosator" that was developed to meter the powder dose.…”

“…In vivo, it showed therapeutic efficacy in mice infected with extensively resistant TB (XDR-TB) when injected subcutaneously (SC) [ 7 ]. However, CPZEN-45 has a low oral bioavailability due to its poor absorption from the gastrointestinal tract [ 12 ]. Administration of CPZEN-45 by the pulmonary route as an inhaled aerosolized powder would deliver the drug directly to the primary site of infection, achieving higher drug concentrations in the lungs and potentially reducing treatment time.…”

“…Efficacy studies were then performed in the guinea pig model of TB using the optimized CPZEN-45 formulation and a preliminary dose selected from a brief PK study [17]. Pulmonary administration of CPZEN-45 powder for inhalation resulted in a 1-log reduction in the bacterial burden in the lungs of animals treated with dry powder aerosols of CPZEN-45 or an SC injection of solution compared to untreated animals [18].…”