Multiplex Assay of Second-Line Anti-Tuberculosis Drugs in Dried Blood Spots Using Ultra-Performance Liquid Chromatography-Tandem Mass Spectrometry

Abstract

As dried blood spots (DBSs) have various advantages over conventional venous blood sampling, some assays for detection of one or two anti-tuberculosis (TB) drugs in DBSs have been developed. However, there are no assays currently available for the simultaneous measurement of three or more anti-TB drugs in DBSs. In this study, we developed and evaluated a multiplex method for detecting nine anti-TB drugs including streptomycin, kanamycin, clarithromycin, cycloserine, moxifloxacin, levofloxacin, para-aminosalicylic acid, prothionamide, and linezolid in DBSs by using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Seventy-nine patient samples of DBS were analyzed on the UPLC-MS/MS system. All drug concentrations were determined within 4 min, and assay performance was evaluated. All drugs were clearly separated without ion suppression. Within-run and between-run precisions were 1.7-13.0% and 5.7-17.0%, respectively, at concentrations representing low and high levels for the nine drugs. Lower limits of detection and quantification were 0.06-0.6 and 0.5-5.0 µg/mL, respectively. Linearity was acceptable at five level concentrations for each drug. Correlations between drug concentrations in plasma and DBSs by using Passing-Bablock regression and Pearson's rho (ρ, 0.798-0.989) were acceptable. In conclusion, we developed a multiplex assay to measure nine second-line anti-TB drugs in DBSs successfully. This assay provided convenient and rapid drug quantification and could have applications in drug monitoring during treatment.

Efficacy of Human Exposures of Gepotidacin (GSK2140944) against Escherichia coli in a Rat Pyelonephritis Model

“…This was also achieved for levofloxacin i.v. infusion q24h targeting the human exposure profile for a once-daily oral dose of 500 mg. For both gepotidacin (data on file) and levofloxacin (16, 17), concentrations in whole blood and plasma have been shown to be similar; therefore, additional adjustments for blood-plasma partitioning are not necessary. Concentrations of gepotidacin in kidney homogenates of rats dosed to the 1,500-mg oral human exposure target were ∼6-fold higher than in whole blood based on total drug C max and ∼7-fold higher based on total drug AUC 0–12 .…”

Determination of Kanamycin by High Performance Liquid Chromatography

“…Dried blood spots (DBSs) were more convenient than traditional venous blood sampling. In one anti-TB drug analysis, 0.1 mol/L HCl in mixed methonal solution was used for deproteinization of DBS samples [18].…”

“…The LOD and LOQ were 0.5 μg/mL and 2.5 μg/mL, respectively [16]. The LOD and LOQ were further expanded to 0.3 μg/mL and 5.0 μg/mL, respectively, and tested in dried blood spots (DBSs) samples in another study [18].…”

The Importance of Clinical Pharmacokinetic–Pharmacodynamic Studies in Unraveling the Determinants of Early and Late Tuberculosis Outcomes

“…For studies in TB endemic, resource poor countries bioanalysis often requires cold shipment to distant laboratories. Novel approaches to measuring drug exposure or activity in resource-limited settings include the use of dried blood spot methods to store and transport samples [ 25–27 ], urine colorimetry to detect low rifampicin exposure [ 28 ] or co-culturing patients’ Mtb isolate with their plasma on treatment in liquid culture to give an indication of the relative activity of the treatment regimen [ 29 ]. For some antituberculous drugs (e.g., amikacin [ 30 ], kanamycin [ 30 ], moxifloxacin [ 31 ] and linezolid [ 32 ] in MDR-TB) limited plasma sampling strategies have sought to determine which single time-point measurements best represent more complex PK indices.…”