Determination of tigecycline in human skin using a novel validated LC-MS/MS method.

BACKGROUND: To develop and validate a sensitive and novel bioanalytical method for measuring tigecycline concentrations in human skin using LC-MS/MS. RESULTS: The method utilizes addition of a stabilizing agent to the human skin or surrogate (human liver or rat skin), homogenization of human skin in a strong acidic-methanol extraction solvent, centrifugation of the skin suspension, filtration of the skin suspension supernatant, separation by LC (Polaris™ C18-A 50 × 2.0 mm), and detection of tigecycline by MS/MS. Linearity was 50-20,000 ng/g, using a sample size of 100 mg. The intra-and inter-day accuracy and precision of the assay met acceptance criteria. CONCLUSION: This method has been successfully applied to 17 incurred human skin samples from volunteers with surgical infections who received intravenous doses of tigecycline (100 mg initial loading dose and 50 mg every 12 h for at least 2 days). Tigecycline concentrations in these samples ranged from 185 to 2853 ng/g.

A sensitive human bone assay for quantitation of tigecycline using LC/MS/MS.

Tigecycline (Tygacil,Wyeth) is a first-in-class, broad spectrum antibiotic with activity against multiple-resistant organisms. In order to address the unexpectedly low tigecycline concentrations in human bone samples analyzed using a LC/MS/MS method developed elsewhere, we have developed and validated a new and sensitive human bone assay for the quantitation of tigecycline using LC/MS/MS. The new method utilizes the addition of a stabilizing agent to the human bone sample, homogenization of human bone in a strong acidic-methanol extraction solvent, centrifugation of the bone suspension, separation by liquid chromatography, and detection of tigecycline by mass spectrometry. Linearity was demonstrated over the concentration range from 50 to 20,000 ng/g using a 0.1g human bone sample. The intra- and inter-day accuracy of the assay was within 100+/-15%, and the corresponding precision (CV) was <15%. The stability of tigecycline was evaluated and shown to be acceptable under the assay conditions. The extraction recovery of tigecycline with the current method was 79% when using radio-labeled rat bone samples as a substitute for human bone samples. Twenty-four human bone samples collected previously from volunteers without infections who had elective orthopedic surgery after receiving a single dose of tigecycline were re-analyzed using the current validated method. Tigecycline concentrations in these samples ranged from 238 to 794 ng/g with a mean value 9 times higher than the mean concentration previously reported. The data demonstrated that the current method has significantly higher extraction efficiency than the previously reported method.

A novel antibiotic bone assay by liquid chromatography/tandem mass spectrometry for quantitation of tigecycline in rat bone.

Tigecycline (Tygacil) is a first-in-class, broad spectrum antibiotic with activity against antibiotic-resistant organisms. In rats and humans, tigecycline readily distributes to bone tissue but its accuracy of quantitation via liquid chromatography/mass spectrometry (LC/MS/MS) is hindered by a low extraction recovery when using a conventional plasma extraction method. To overcome this issue, we have identified an effective extraction solvent for quantitation of tigecycline in rat bone. The current LC/MS/MS bone assay is novel, simple, and sensitive, and has a wide linear range of 50-10,000 ng/g. The assay requires homogenization of the rat bone in a strong acidic-methanol extraction solvent, centrifugation of the bone suspension, separation of the supernatant with liquid chromatography, and detection of tigecycline with tandem mass spectrometry. The incurred pooled rat bone samples obtained from rats given 3mg/kg/day of [(14)C]-tigecycline and non-radio-labeled tigecycline were analyzed with the current method. The absolute extraction recovery of the bone assay for tigecycline was 77.1%. The intra-day accuracy ranged from 91.7 to 106% with precision (CV) of 1.9-10.7%, and inter-day accuracy ranged from 96.1 to 100% with a precision of 6.3-8.7%. In addition, biological activity was demonstrated for the tigecycline extracted from incurred rat bone. This bone assay provides an important analytical tool for the determination of drug concentrations (especially, antimicrobials) in rodent bone tissues and has served as the foundation of development and validation of a similar bone assay for tigecycline in human bone tissues.