Pharmacokinetics of oral fludrocortisone in septic shock.

AIM: The combination of hydrocortisone and fludrocortisone improved outcomes in septic shock. However, the specific role of fludrocortisone remains controversial and its pharmacokinetics (PK) has never been investigated in septic shock. This study aimed at characterizing the PK of fludrocortisone in septic shock. METHODS: This was a single-centre ancillary PK study of a large multinational trial of crystalloids versus colloids for acute hypovolemia in intensive care unit (ICU) patients. In 21 adults with septic shock, fludrocortisone plasma concentrations were measured by liquid chromatography-mass spectrometry tandem analysis, before and repeatedly until 18 h after an oral dose of 50 µg. PK parameters were estimated using a nonlinear mixed-effects modelling. RESULTS: Undetectable plasma concentrations were observed in 7 out of 21 patients. In the remaining 14 patients, plasma fludrocortisone concentrations were best described by a one-compartmental model with first-order absorption, a lag time (Tlag ) before the absorption phase, and first-order elimination. Severity of illness, as quantified by Simplified Acute Physiology Score II, significantly increased Tlag and apparent clearance. There was a large inter-individual variability in PK parameters. The population estimates of PK parameters (inter-individual variability) were: Tlag 0.65 h (98%), apparent clearance 40 l h-1 (49%) and apparent volume of distribution 78 l (75%). Plasma half-life was estimated at 1.35 h (95% CI, 0.84-2.03) and area under the curve of plasma concentrations was estimated at 1.25 µg h l-1 (95% CI, 1.09-1.46). CONCLUSIONS: A single oral dose of fludrocortisone yielded undetectable plasma concentrations in one-third of adults with septic shock. Fludrocortisone PK showed a short plasma elimination half-life and a large inter-individual variability.

A highly sensitive method for the quantification of fludrocortisone in human plasma using ultra-high-performance liquid chromatography tandem mass spectrometry and its pharmacokinetic application.

A simple and high sensitive ultra-high-performance liquid chromatography tandem mass spectrometry method for the determination of fludrocortisone in human plasma was developed and validated as per guidelines. The analyte and internal standard (IS), fludrocortisone-d5 , were extracted from human plasma via liquid-liquid extraction using tert-butyl methyl ether. The chromatographic separation was achieved on a Chromolith RP18e column using a mixture of acetonitrile and 2 mm ammonium formate (70:30, v/v) as the mobile phase at a flow rate of 0.7 mL/min. Quantitation was performed on a triple quadrupole mass spectrometer employing electrospray ionization technique, operating in multiple reaction monitoring and positive ion mode. The precursors to product ion transitions monitored for fludrocortisone and IS were m/z 381.2 → 343.2 and 386.2 → 348.4, respectively. The assay was validated with linear range of 40-3000 pg/mL. The intra- and inter-day precisions (relative standard deviation) were within 0.49-7.13 and 0.83-5.87%, respectively. The proposed method was successfully applied to pharmacokinetic studies in humans.

Human plasma quantification of fludrocortisone using liquid chromatography coupled with atmospheric pressure chemical ionization mass spectrometry after low-dosage administration.

BACKGROUND: Fludrocortisone acetate is given at very low dosage (50 µg) to patients suffering from septic shock with controversial clinical results. However, it is not clear if absorption is effective in these patients. METHODS: An analytical method based upon liquid chromatography coupled to triple quadrupole spectrometry detection with atmospheric pressure chemical ionization interface has been developed for the identification and quantification of fludrocortisone, the active molecule circulating in human plasma. A solid phase extraction of plasma was used after addition of fludrocortisone-D2 as internal standard. Compounds were separated on a C18 column with a gradient of methanol-formate buffer. The ion transitions used to monitor analytes were m/z 381→239 and m/z 381→181 for fludrocortisone and m/z 383→239 and m/z 383→181 for fludrocortisone-D2. RESULTS: Retention times were 4.0 min for both compounds. Calibration curves were linear for fludrocortisone in the 0.1-25 ng/ml range. The limits of detection and quantification were 0.05 ng/ml and 0.1 ng/ml, respectively. The intra- and inter-assay precisions were lower than 10.9% and the recovery was 101.8%. A slight matrix effect by about 10% was observed. Application of the method to a patient in septic shock treated with one 50-µg dose of fludrocortisone acetate has shown a maximal plasma concentration of 0.36 ng/ml obtained after 2h. CONCLUSION: This method allows fludrocortisone pharmacokinetic/pharmacodynamic studies when given at low dosage in an intensive care unit in case of adrenal insufficiency during a septic shock.

Liquid chromatography-mass spectrometry with on-line solid-phase extraction by a restricted-access C18 precolumn for direct plasma and urine injection.

In this paper, the on-line coupling of solid-phase extraction, based on a restricted-access support with liquid chromatography-mass spectrometry (LC-MS), for the analysis of biological samples is described. The system was tested with cortisol and prednisolone for plasma analysis and arachidonic acid for urine analysis. A precolumn packed with a 25-micron C18 alkyl-diol support is used for direct plasma or urine injection. Using column-switching techniques, the analytes enriched on the precolumn are eluted to the analytical column without transfer loss. An on-line heart-cut technique was employed and only the analyte-containing fraction eluting from the LC column is directed to the MS to protect the LC-MS interface and ion-source from contamination. The whole system is operated in a parallel mode, that is, sample pre-treatment and LC-MS analysis are performed simultaneously to provide the shortest possible analysis time. The only off-line sample pre-treatment step required was centrifugation to remove particulate matter. With the fully automated system, total analysis times of 5 and 9.5 min were achieved for cortisol in serum and arachidonic acid in urine, respectively. Cortisol and related compounds were quantitatively recovered from plasma with a detection limit for prednisolone (direct injection of 100 microliters on restricted-access precolumn) of 2 ng/ml.

A sensitive radioimmunoassay for fludrocortisone in human plasma.

Fludrocortisone has been a mainstay of therapy for orthostatic hypotension for many years. Clinical experience suggests that there exists a substantial interindividual variation in responsiveness to the drug. To assess this, we have developed an assay that permits measurement of the low concentrations of fludrocortisone found in human plasma. Fludrocortisone was detected by radioimmunoassay. A polyclonal rabbit antibody, raised against dexamethasone which cross-reacts strongly with fludrocortisone, was reacted with either standard or unknown samples in the presence of [125I]fludrocortisone-3-TyrNH2 (synthesized by coupling tyrosine amide to fludrocortisone-3-oxime and iodinating with chloramine T oxidation). The ED10, ED50, and ED80 were 0.34, 5.0, and 30 ng/mL of plasma, respectively. The cross reactivity with other 9-fluorinated steroids was found as follows: dexamethasone, 340%; betamethasone, 230%; and triamicinolone, 8%. To preclude an erroneous result, subjects who were pregnant or receiving any steroid medication were excluded from the study. The percent cross-reactivity with the main naturally occurring steroids was as follows: 11-desoxycortisol 3.2%, cortisol 1.1%, DOC 0.3%, pregnenolone 0.1%, corticosterone 0.06%, progesterone 0.05%, and aldosterone < 0.05%. The only compound with potential for interference, because of its high level in the circulation in the early morning, was cortisol.(ABSTRACT TRUNCATED AT 250 WORDS)

Quantitative determination of 21-hydroxy-deflazacort in human plasma using gradient semi-microbore liquid chromatography.

A sensitive and selective liquid chromatographic procedure to quantitate the deflazacort metabolite 21-hydroxy-deflazacort (DF-21OH) in human plasma was developed and validated. DF-21OH and fludrocortisone acetate (internal standard, IS) were isolated from human plasma (2 mL) by solid-phase extraction onto C-18 cartridges. Potential interferences were selectively removed and analytes were eluted with ethyl acetate. Following evaporation, the residue was reconstituted for HPLC analysis. Separation was achieved by gradient elution using a 5 microns YMC Basic column (2.0 x 100 mm) with mobile phases consisting of 20% methanol and 50% acetonitrile in 50 mM phosphate buffer (pH 3) at a temperature of 50 degrees C. Flow rate was maintained at 0.3 mL/min., and analytes were quantified spectrophotometrically at 246 nm. The assay was validated over the range 1.0 to 500 ng/mL DF-21OH. Calibration curves were prepared using a weighted (1/concentration) nonlinear quadratic regression algorithm. Peak-height ratios were proportional to the amount of DF-21OH added to plasma. Assay precision (%RSD) ranged from 4.2 to 11%, with a corresponding assay accuracy (% relative error) of +/- 2.8%. Absolute recovery of DF-21OH from plasma was 78-86% over the concentration range. The minimum quantitation limit was 1.0 ng/mL.