1-Adamantylamine a simple urine marker for screening for third generation adamantyl-type synthetic cannabinoids by ultra-performance liquid chromatography tandem mass spectrometry.

Background Synthetic cannabinoids (NOIDS) are novel psychotropic drugs (NPS) currently freely sold in the United Kingdom as 'research chemicals'. Detection of NOIDS use is not available in current routine methods. Here we describe a marker which helps determine which patients have used these substances. Methods In a test case, ultra-performance liquid chromatography mass spectrometry (UPLC-Tof) was used to screen the legal high Herbal Haze II, the contents of hand-rolled cigarettes and five patient samples for NOIDS and their metabolites. Results Analysis of legal high Herbal Haze II and cigarettes identified the third generation adamantyl-type NOIDS N-(1-adamantyl)-1-pentyl-1H-indazole-3-carboxamide (AKB-48), 5F-AKB-48 and N-adamantyl-1-fluoropentylindole-3-carboxamide (STS-135). Out of 18 potential metabolites, 1-adamantylamine (C10H17N) was detected in all five urine samples. This adamantyl-type NOID marker was incorporated into our routine LC-MS/MS urine screen. Out of 14,436 random urine samples screened over eight months, 296 (2.05%) tested positive for the adamantyl-type NOID marker. Conclusion We have discovered a urine marker for identifying patients smoking legal high products containing the third generation adamantyl-type NOIDS such as AKB-48 and its fluoropentyl analogue 5F-AKB-48, which are among the most popular NOIDS currently available in legal high products sold in UK. This marker can be incorporated into routine LC-MS/MS drug screening alongside classic drugs of abuse. Positive detection rates for this new legal high marker are greater than for established classic drugs that are routinely screened such as amphetamine. This work highlights the need for a flexible toxicology screening service capable of adapting to changes in drug use such as the growing popularity of legal highs/NPS.

The effect of intravenous sulfobutylether7 -ß-cyclodextrin on the pharmacokinetics of a series of adamantane-containing compounds.

Intravenously administered (i.v.) drug-cyclodextrin (CD) inclusion complexes are generally expected to dissociate rapidly and completely, such that the i.v. pharmacokinetic profile of a drug is unchanged in the presence of CD. The altered pharmacokinetics of a synthetic ozonide in rats has been attributed to an unusually high-binding affinity (2.3 × 10(6) M(-1) ) between the drug and sulfobutylether7 -ß-cyclodextrin (SBE7 -ß-CD) with further studies suggesting a significant binding contribution from the adamantane ring. This work investigated the binding affinity of three adamantane derivatives [amantadine (AMA), memantine (MEM) and rimantadine (RIM)] to SBE7 -ß-CD and the impact of complexation on their i.v. pharmacokinetics. In vitro studies defined the plasma protein binding, as well as the impact of SBE7 -ß-CD on erythrocyte partitioning of each compound. SBE7 -ß-CD binding constants for the compounds were within the typical range for drug-like molecules (10(2) -10(4) M(-1) ). The pharmacokinetics of AMA and MEM were unchanged; however, significant alteration of RIM plasma and urinary pharmacokinetics was observed when formulated with CD. In vitro studies suggested two factors contributing to the altered pharmacokinetics: (1) low plasma protein binding of RIM, and (2) decreased erythrocyte partitioning in the presence of high SBE7 -ß-CD concentrations. This work demonstrated the potential for typical drug-cyclodextrin interactions to alter drug plasma pharmacokinetics.

Determination of amantadine in biological fluids using simultaneous derivatization and dispersive liquid-liquid microextraction followed by gas chromatography-flame ionization detection.

A one-step derivatization and microextraction technique for the determination of amantadine in the human plasma and urine samples is presented. An appropriate mixture of methanol (disperser solvent), 1,2-dibromoethane (extraction solvent), and butylchloroformate (derivatization agent) is rapidly injected into samples. After centrifuging, the sedimented phase is analyzed by gas chromatography-flame ionization detection (GC-FID). The kind of extraction and disperser solvents and their volumes, amount of derivatization agent and reaction/extraction time which are effective in derivatization/dispersive liquid-liquid microextraction (DLLME) procedure are optimized. Under the optimal conditions, the enrichment factor (EF) of the target analyte was obtained to be 408 and 420, and limit of detection (LOD) 4.2 and 2.7ngmL(-1), in plasma and urine respectively. The linear range is 14-5000 and 8.7-5000ng/mL for plasma and urine, respectively (squared correlation coefficient≥0.990). The relative recoveries obtained for the spiked plasma and urine samples are between 72% and 93%. Moreover, the inter- and intra-day precisions are acceptable at all spiked concentrations (relative standard deviation <7%). Finally the method was successfully applied to determine amantadine in biological samples.

1H NMR detection of small molecules in human urine with a deep cavitand synthetic receptor.

A water-soluble deep cavitand recognized alkylammonium salts, including the drug amantadine hydrochloride, in spiked samples of human urine. The signals of the guests are detected by (1)H NMR upfield of 0 ppm and so occur in a spectroscopic window that is outside of the normal region and distinct from the signals of the biofluid components.

High-performance liquid chromatographic determination of memantine in human urine following solid-phase extraction and precolumn derivatization.

A validated HPLC-UV method is presented for the quantification of urinary memantine hydrochloride, a novel medication approved to treat moderate and advanced cases of Alzheimer's disease. The drug and amantadine hydrochloride, the internal standard, were extracted from human urine using SPE. The extract was then buffered and derivatized at room temperature using o-phthalaldehyde in the presence of N-acetyl-L-cyteine. Chromatographic separation of the formed derivatives was achieved on a C18 column using methanol-water mobile phase adjusted to pH 7 and pumped isocratically at 1 mL/min. The UV detector was set at 340 nm. The chromatographic run time did not exceed 10 min. The LOD and LOQ were 8 and 20 ng/mL, respectively. The RSDs for intraday and interday precisions did not exceed 5.5%. The method was used to monitor memantine hydrochloride in human urine in order to determine an appropriate sampling interval for future noninvasive therapeutic drug monitoring. The assay could also be applied to the determination of amantadine. The described assay showed that a postdosing time interval of 25-75 h seems adequate for sampling and monitoring memantine in urine.

Analysis of amantadine in biological fluids using hollow fiber-based liquid-liquid-liquid microextraction followed by corona discharge ion mobility spectrometry.

A method based on liquid-liquid-liquid microextraction combined with corona discharge ion mobility spectrometry was developed for the analysis of amantadine in human urine and plasma samples. Amantadine was extracted from alkaline aqueous sample as donor phase through a thin phase of organic solvent (n-dodecane) filling the pores of the hollow fiber wall and then back extracted into the organic acceptor phase (methanol) located in the lumen of the hollow fiber. All variables affecting the extraction of analyte including acceptor organic solvent type, concentration of NaOH in donor phase, ionic strength of the sample and extraction time were studied. The linear range was 20-1000 and 5-250 ng/mL for plasma and urine, respectively (r(2)≥0.990). The limits of detection were calculated to be 7.2 and 1.6 ng/mL for plasma and urine, respectively. The relative standard deviation was lower than 8.2% for both urine and plasma samples. The enrichment factors were between 45 and 54. The method was successfully applied for the analysis of amantadine in urine and plasma samples.

A comparative study of pharmacokinetics, urinary excretion and tissue distribution of platinum in rats following a single-dose oral administration of two platinum(IV) complexes LA-12 (OC-6-43)-bis(acetato)(1-adamantylamine)amminedichloroplatinum(IV) and satraplatin (OC-6-43)-bis(acetato)amminedichloro(cyclohexylamine)platinum(IV).

PURPOSE: This study compared the pharmacokinetics, tissue distribution, and urinary excretion of platinum in rats after single oral doses of LA-12 and satraplatin. METHODS: Both platinum derivatives were administered to male Wistar rats as suspensions in methylcellulose at four equimolar doses within the range of 37.5-300 mg LA-12/kg body weight. Blood sampling was performed until 72 h, and plasma and plasma ultrafiltrate were separated. Moreover, urine was collected until 72 h, and kidney and liver tissue samples were obtained at several times after administration. Platinum was measured by atomic absorption spectrometry. The pharmacokinetics of platinum was analyzed by population modelling and post hoc Bayesian estimation as well as using non-compartmental pharmacokinetic analysis of the mean concentration-time curves. RESULTS: Platinum was detected in all plasma and ultrafiltrate samples 15 min after oral administration of both compounds and peaked between 3-4 h and 1-3 h, respectively. Similar for LA-12 and satraplatin, the C (max) and AUC values of plasma and ultrafiltrate platinum increased less than in proportion to dose. The mean C (max) and AUC values of plasma platinum observed after administration of LA-12 were from 0.84 to 2.5 mg/l and from 20.2 to 75.9 mg h/l. For ultrafiltrate platinum, the corresponding ranges were 0.16-0.78 mg/l and 0.63-1.8 mg h/l, respectively. The AUC of plasma platinum was higher after satraplatin (P < 0.001). However, administration of LA-12 resulted in significantly higher AUC values of ultrafiltrate platinum after the doses of 150 mg and 300 mg/kg (P < 0.01), respectively, and the C (max) values were significantly higher starting from the dose of 75 mg/kg LA-12 and upward (P < 0.01). Cumulative 72-h urinary recovery of platinum dose was below 5% for both compounds, and it decreased with the dose of satraplatin (P < 0.01), while a numerical decrease was observed after administration of LA-12 that did not reach statistical significance (P = 0.41). The renal clearance of free platinum was similar regardless of the dose and compound administered. Platinum concentrations in the liver homogenate exceeded those in the kidney. Distribution of platinum to tissues was higher after LA-12 compared to satraplatin. The difference in kidney platinum increased with dose and was twofold after 350 mg/kg LA-12. Liver platinum was twofold higher after LA-12 across all four doses. CONCLUSIONS: In conclusion, this first comparative pharmacokinetic study with LA-12 and satraplatin shows that characteristics of platinum exposure evaluated in the plasma, plasma ultrafiltrate and kidney and liver tissues increase less than in proportion to dose following a single-dose administration of 37.5-300 mg/kg to Wistar rats. These findings together with the dose-related elevation in the pharmacokinetic characteristics V/F and CL/F of platinum and ultrafiltrate platinum as well as a drop in platinum urinary recovery are consistent with a dose-related decrease in the extent of oral bioavailability most likely due to saturable intestinal absorption.

Fluorimetric liquid chromatographic analysis of amantadine in urine and pharmaceutical formulation.

A simple and sensitive liquid chromatographic method is described for the analysis of amantadine and memantine. The method is based on the derivatization of amantadine and memantine extracted from alkalified samples with (2-naphthoxy)acetyl chloride at mild conditions. The resulting derivatives were analyzed by isocratic HPLC with a fluorimetric detector (lambdaex, 227 nm; lambdaem, 348 nm). The linear range for the determination of amantadine or memantine spiked in urine (1.0 ml) was 1.0-10.0 nmol with a detection limit of about 0.2 nmol (S/N = 3; injected sample 20 microl). Only amantadine preparations are available on our local market, and application of the method to the analysis of amantadine in formulation and in the urine of a dosed subject was demonstrated and proved feasible. Quantitation of AT in tablets or capsules is capable in the linear range of 2.0-50.0 microM. Toluene was used as the solvent for extracting amantadine or memantine in samples and the resulting toluene extract was directly subjected to subsequent derivatization without solvent replacement leading to a simpler analytical procedure.

In vivo analysis of amantadine renal clearance in the uninephrectomized rat: functional significance of in vitro bicarbonate-dependent amantadine renal tubule transport.

Amantadine transport into renal proximal and distal tubules is bicarbonate dependent. In the present study, we addressed the effects of bicarbonate on renal clearance and urinary excretion of amantadine. Renal clearance of kynurenic acid was also studied to determine whether bicarbonate effects are specific for organic base transport by the kidney. After a moderate diuresis was established, animals received i.v. [(3)H]amantadine or [(3)H]kynurenic acid followed by an acute dose of sodium bicarbonate or physiological saline. Urine and blood samples were analyzed for [(3)H]amantadine or [(3)H]kynurenic acid, blood gases, and pH. Amantadine and kynurenic acid were excreted by the kidneys, and both compounds underwent renal tubular secretion. Amantadine metabolism occurred, and one metabolite was detected in the urine. In the bicarbonate-treated rats, the total amount of amantadine excreted in the urine was decreased, whereas the amount of metabolite recovered was similar in both groups. Bicarbonate treatment caused a sustained increase in blood bicarbonate levels, a mild increase in blood pH, and a decrease in amantadine renal clearance and in the amantadine/creatinine clearance ratio. Only a transient decrease in the renal clearance of kynurenic acid and the kynurenic acid/creatinine clearance ratio was observed. This study demonstrates that short-term changes in bicarbonate concentration may have significant effects on renal organic cation elimination. Coupled with our previous in vitro demonstration of bicarbonate-dependent organic cation transport, the present study suggests that bicarbonate inhibition of renal tubule organic cation secretion may explain the previous observation that bicarbonate dosing decreases amantadine excretion by the kidney.

Amantadine acetylation may be effected by acetyltransferases other than NAT1 or NAT2.

Amantadine is a drug with a primary amino group, and consequently a likely candidate for metabolism by acetylation. This study assessed the possibility that a person's polymorphic (NAT2) acetylator phenotype could be used to predict the extent of amantadine acetylation. Thirty-eight normal, healthy volunteers were NAT2 acetylator phenotyped with sulfapyridine. Of the six fastest (75-86%) and six slowest (34-40%) sulfapyridine acetylators, two and three, respectively, had acetylamantadine present (18-338 microg) in the 8-h urine collection. There was no correlation between NAT2 acetylator phenotype and amantadine acetylation (p<0.5), and no difference in the total urine amantadine excreted over 8 h between acetylators and nonacetylators (28.3+/-9.7 vs. 30.4+/-9.6 mg, respectively, mean +/- SD). Acetylamantadine represented 0.1-1.5% (median 0.5%) of urinary drug content over 8 h. Our data confirm that amantadine is acetylated in humans and demonstrate for the first time that the extent is not correlated with NAT2 acetylator phenotype. Parallel in vitro enzyme studies indicate the possibility that neither NATI nor NAT2 is responsible for acetylation of amantadine.

Chronic tobacco smoking and gender as variables affecting amantadine disposition in healthy subjects.

Amantadine HCl (3 mg kg-1) was administered orally to 20 young healthy adults. Its apparent volume of distribution (V2/F) was higher in smokers than nonsmokers, 6.05 +/- 0.86 vs 4.87 +/- 0.85 l kg-1; (mean +/- s.d., 10/group, P < 0.011), and no gender-associated effect was observed. Renal clearance did not vary with time-interval, but urinary recovery at 48 h was higher in men than in women (60.2 +/- 7.5% vs 47.0 +/- 15.0%, P < 0.032). Males had higher renal clearances than females when normalised for body mass index (BMI, 0.492 +/- 0.284 vs 0.248 +/- 0.137 l-1 BMI h-1, (10/group, P < 0.032)). On combining data from a previous study, the weight normalised renal clearance was also higher in men than in women, 0.160 +/- 0.075 vs 0.102 +/- 0.053 l kg-1 h-1 (19/group, P < 0.01). Chronic tobacco smoking did not alter the plasma or renal amantadine clearance. We conclude that gender and tobacco smoking are independent variables effecting amantadine disposition.

Polymeric reagents for derivatizations in micellar electrokinetic chromatography.

A polymer immobilized o-nitrobenzophenone reagent was prepared for analysis of amine drugs in micellar electrokinetic chromatography (MEKC). A model compound, propylamine, was used to characterize the reagent's performance in MEKC. Derivatizations were performed on the CE instrument with reagent in the sample vial. The yielded derivative was directly sampled from the reaction mixture, and directly injected onto the MEKC system. The derivatization reagent was also applied to the derivatization of n-alkyl amine mixtures and amino acids. The method was validated for adamantanamine in urine and in plasma by single-blind spike analysis. Precisions and accuracies for all samples were less than 6.0% for urine samples and 10% for plasma samples. The procedure was a direct injection technique requiring minimal sample preparation for the analysis of drugs in biofluids.

Direct determination of adamantanamine in plasma and urine with automated solid phase derivatization.

A simple, highly sensitive and selective method is described for adamantanamine determination in plasma and urine by high-performance liquid chromatography with fluorescence detection. The method involved a simultaneous extraction and derivatization of biological fluids with a 9-fluoreneacetate (9-FA) solid-phase derivatization reagent. This approach eliminated tedious sample preparation steps and provided automatic derivatization with selective and efficient sample clean-up for direct injection of biological fluids. Derivatized adamantanamine was separated under conventional reversed-phase conditions and determined by fluorescence detection. The optimization and validation of the derivatization method with the 9-FA solid-phase reagent is described.

[Determination of amantadine by pre-labeling with 3-(7'-methoxycoumarin-3'-carbonyl)-benzoxazoline-2-thione and high-performance liquid chromatography with fluorescence detection].

3-(7'-Methoxycoumarin-3'-carbonyl)-benzoxazoline-2-thione (MCBT) was synthesized as a precolumn fluorescent labeling reagent for amantadine for use in high-performance liquid chromatography (HPLC). Amantadine was derivatized quantitatively into a fluorescent compound through the amino group by treatment with MCBT at room temperature for 5 min in aqueous 95% acetonitrile solution in the presence of 4-dimethylaminopyridine. The derivative was subjected to HPLC on TSK gel ODS-80TM (250 x 4.6 mm i.d.) with methanol-water (10:1) as the mobile phase and monitored with an excitation wavelength of 355 nm and an emission wavelength of 405 nm. The limit of detection was 2.0 micrograms/ml in the urine. This method was satisfactory with respect to simplicity and precision to quantify amantadine spiked in the urine.

Amantadine neurotoxicity in a pediatric patient with renal insufficiency.

Amantadine hydrochloride, a dopamine agonist with antiviral and antiparkinsonism properties, is used for the prevention and treatment of influenza A respiratory infections in high-risk populations. The occurrence of amantadine-induced hallucinations and tremors is described in a young, renal transplant patient with declining renal function. Following discontinuation of amantadine, plasma amantadine concentrations were correlated with central nervous system toxicity. In view of the usage of amantadine in renal transplant recipients and the elderly, clinicians must be alert to the possibility of amantadine-induced neurotoxicity in patients with changing renal function.

High-performance liquid chromatographic determination of amantadine in urine after micelle-mediated pre-column derivatization with 1-fluoro-2,4-dinitrobenzene.

Cationic micelles have been used for the derivatization of the anti-Parkinson drug amantadine with the chromophore 1-fluoro-2,4-dinitrobenzene in urine. In the presence of 90 mM cetyltrimethylammonium bromide (CTAB), the conversion of amantadine into its derivative is complete within 4 min at 60 degrees C and pH 11. Such a short reaction time allows a fully automated pre-column derivatization of amantadine in an on-line combination with reversed-phase high-performance liquid chromatography. This cannot be attained when using purely aqueous derivatization mixtures because then the reaction takes some 20 min at the same temperature. Without the use of an internal standard, the repeatability of the automated determination at the 0.5 microgram ml-1 level is ca. 6%, whilst the detection limit is 75 ng ml-1 (S/N = 3). The present study clearly demonstrates that micellar systems can be beneficially used for the on-line precolumn derivatization of amines in urine.

Pharmacokinetics of amantadine hydrochloride in subjects with normal and impaired renal function.

Amantadine is useful for the prevention and treatment of influenza A and for the treatment of Parkinson's disease and drug-induced extrapyramidal disorders. We have compared the pharmacokinetics of amantadine in patients with impaired or negligible renal function to that in normal subjects. The half-life of elimination in subjects with normal renal function was 11.8 +/- 2.1 hours (range, 9.7 to 14.5 h). Eight patients with various degrees of renal insufficiency (creatinine clearance from 43.1 to 5.9 mL/min . 1.73 m2) had half-lives of elimination from 18.5 h to 33.8 days. We also studied 10 patients on thrice-weekly hemodialysis. Assuming complete bioavailability of the drug, less than 5% of the dose was removed by each 4-hour hemodialysis. The mean half-life of elimination during chronic hemodialysis was 8.3 days (range, 7.0 to 10.3). We present guidelines for use of amantadine in patients with impaired renal function, including those on maintenance hemodialysis.

Gas chromatographic determination of amantadine hydrochloride (Symmetrel) in human plasma and urine.

A method for the determination of amantadine hydrochloride at concentrations down to 10 ng/ml in human plasma and urine is described. After addition of a known amount of amphetamine sulphate as internal standard to 1 ml of plasma or urine, amantadine is extracted at basic pH in toluene. Both compounds are derivatized with trichloroacetyl chloride. The derivatives are determined by gas chromatography using a 63Ni electron-capture detector. The technique was applied in a study of the elimination of amantadine after oral administration to man; plasma concentrations are reported.