Bioanalytical Method for the Determination of Hydroxyproline in Mouse Kidney by High-Performance Liquid Chromatography with Tandem Mass Spectrometric Detection.

An analytical method was developed and validated for the measurement of hydroxyproline (Hyp) levels in mouse kidney by high-performance liquid chromatography with tandem mass spectrometric detection (LC/MS/MS) using an analytical column specially designed for the LC/MS/MS analysis for intact amino acids. Tissue hydrolyzed with hydrochloric acid could be directly injected into the LC/MS/MS, as well as separated and detected using the deuterium labelled Hyp as an internal standard. The calibration curve showed good linearity from 5 to 500 nmol/mg of tissue; the precision and accuracy, including within- and between-run, were less than 6% and within 100 ± 6%, respectively.

Absorption and safety of alendronate, a nitrogen-containing bisphosphonate, after intrapulmonary administration in rats.

Alendronate, a nitrogen-containing bisphosphonate, has been used as a first-choice drug for the treatment of hypercalcemia and osteoporosis. In the present study, we examined the absorption and safety of alendronate after intrapulmonary administration in rats. The bioavailability (BA) of alendronate after intrapulmonary administration was 47% at a dose of 5 mg/kg, while the BA after oral administration was only 2.9% at a dose of 50 mg/kg in rats. Plasma calcium level, an index of the pharmacological effect of alendronate, was effectively reduced after intrapulmonary administration of alendronate. Furthermore, alendronate continuously reduced the increase in plasma calcium levels for 9 days after a single intrapulmonary administration in rats with 1α-hydroxyvitamin-D(3)-induced hypercalcemia. Intrapulmonary administration of alendronate also effectively suppressed the decrease in bone mass in a rat model of osteoporosis. Alendronate significantly increased the activity of lactate dehydrogenase (LDH) in bronchoalveolar lavage fluid (BALF), indicating that pulmonary mucosal damage was induced by intrapulmonary administration of alendronate. However, co-administration of superoxide dismutase (SOD) with alendronate completely suppressed the alendronate-induced increase in LDH activity in BALF, while maintaining sufficient pulmonary absorption and therapeutic effects of alendronate in rats with 1α-hydroxyvitamin-D(3)-induced hypercalcemia. These findings indicated that the lung is a promising, noninvasive alternative route for the delivery of alendronate in the treatment of bone diseases.