A rapid and sensitive liquid chromatography-tandem mass spectrometry method for determination of olopatadine concentration in human plasma.

A sensitive and rapid method based on liquid chromatography- tandem mass spectrometry (MS-MS) was developed for the determination of olopatadine in human plasma. Sample preparations were carried out by protein precipitation with the addition of acetonitrile followed by liquid-liquid extraction with ethyl acetate/dichloromethane after internal standard (IS, amitriptyline) spiked. After evaporation to dryness, the resultant residue was reconstituted in mobile phase. Separation of olopatadine and IS from the interferences was achieved on a C(18) column followed by MS-MS detection. The analytes were monitored in the positive ionization mode with a TurboIonspray source. Multiple reaction monitoring using the transition of m/z 338 → 165 and m/z 278 → 91 was performed to quantify olopatadine and IS, respectively. The method had a total chromatographic run time of 3.5 min and linear calibration curves over the concentration range of 0.2-100 ng/mL. The lower limit of quantification was 0.2 ng/mL. For each QC concentration level the intra- and interday precisions were less than 11.4%, and relative errors ranged between -6.40% and 9.26%. The validated method was successfully applied to the quantification of olopatadine concentration in human plasma after administration of olopatadine at an oral dose of 5 mg in order to evaluate the pharmacokinetics.

Brain histamine H receptor occupancy of orally administered antihistamines measured by positron emission tomography with (11)C-doxepin in a placebo-controlled crossover study design in healthy subjects: a comparison of olopatadine and ketotifen.

AIMS: The strength of sedation due to antihistamines can be evaluated by using positron emission tomography (PET). The purpose of the present study is to measure histamine H(1) receptor (H(1)R) occupancy due to olopatadine, a new second-generation antihistamine and to compare it with that of ketotifen. METHODS: Eight healthy males (mean age 23.5 years-old) were studied following single oral administration of olopatadine 5 mg or ketotifen 1 mg using PET with (11)C-doxepin in a placebo-controlled crossover study design. Binding potential ratio and H(1)R occupancy were calculated and were compared between olopatadine and ketotifen in the medial prefrontal (MPFC), dorsolateral prefrontal (DLPFC), anterior cingulate (ACC), insular (IC), temporal (TC), parietal (PC), occipital cortices (OC). Plasma drug concentration was measured, and correlation of AUC to H(1)R occupancy was examined. RESULTS: H(1)R occupancy after olopatadine treatment was significantly lower than that after ketotifen treatment in the all cortical regions (P < 0.001). Mean H(1)R occupancies for olopatadine and ketotifen were, respectively: MPFC, 16.7 vs. 77.7; DLPFC, 14.1 vs. 85.9; ACC, 14.7 vs. 76.1; IC, 12.8 vs. 69.7; TC, 12.5 vs. 66.5; PC, 13.9 vs. 65.8; and OC, 19.5 vs. 60.6. Overall cortical mean H(1)R occupancy of olopatadine and ketotifen were 15% and 72%, respectively. H(1)R occupancy of both drugs correlated well with their respective drug plasma concentrations (P < 0.001). CONCLUSION: It is suggested that 5 mg oral olopatadine, with its low H(1)R occupancy and thus minimal sedation, could safely be used an antiallergic treatment for various allergic disorders. Abbreviations histamine H(1) receptor (H(1)R), histamine H(1) receptor occupancy (H(1)RO), dopamine D(2) receptor (D(2)R), positron emission tomography (PET), blood-brain barrier (BBB), binding potential ratio (BPR), distribution volume (DV).

[Effect of olopatadine hydrochloride, a novel antiallergic agent, on the QT interval in dogs].

Olopatadine hydrochloride (olopatadine), a novel antiallergic agent, is effective in the treatment of allergic rhinitis, chronic urticaria, eczema and dermatitis. It has been reported that terfenadine and astemizole cause side effects on the circulatory system such as QT prolongation followed by serious ventricular arrhythmias (torsades de pointes). To investigate the possibility of QT prolongation, we used both conscious normal dogs and hypokalemia-anesthetized dogs under two conditions: 1) olopatadine used alone and 2) olopatadine used in combination with itraconazole, the CYP3A4-inhibiting antifungal agent, in the present investigation. The group treated with terfenadine alone (30 mg/kg, p.o.) and the group treated with a combination of terfenadine (10 mg/kg, p.o.) and itraconazole (100 mg/kg, p.o.) had a significantly prolonged QT interval. On the other hand, the group treated with olopatadine alone (30 mg/kg, p.o.) and the group treated with a combination of olopatadine (30 mg/kg, p.o.) and itraconazole (100 mg/kg, p.o.) did not show any significant changes in QT interval. Moreover, olopatadine (1 and 5 mg/kg, i.v.) did not influence the QT interval in hypokalemia-anesthetized dogs. These results suggest that there is very little possibility of QT prolongation as a result of clinically used olopatadine.

Determination of olopatadine, a new antiallergic agent, and its metabolites in human plasma by high-performance liquid chromatography with electrospray ionization tandem mass spectrometry.

A rapid, sensitive and specific assay method has been developed to determine plasma concentrations of olopatadine hydrochloride (A) and its metabolites, M1 (B), M2 (C) and M3 (D), using high-performance liquid chromatography with electrospray ionization tandem mass spectrometry (LC-ESI-MS-MS). Olopatadine, its metabolites, and internal standard, KF11796 (E), were separated from plasma using solid-phase extraction (Bond Elut C18 cartridge). The eluate was dried, reconstituted and injected into the LC-ESI-MS-MS system. The calibration curves showed good linearity over the ranges 1-200 ng/ml for olopatadine and M3, and 2-100 ng/ml for M1 and M2, and the method was thoroughly validated and applied to the determination of olopatadine and its metabolites in plasma collected during Phase I clinical trials. Furthermore, the assay values were compared with those determined by the radioimmunoassay method, which has been routinely used to determine olopatadine in plasma.

Central 5-HT2A and D2 dopamine receptor occupancy after sublingual administration of ORG 5222 in healthy men.

It has been suggested that a combined blockade of 5-HT2A and D2-dopamine receptors improves efficacy and reduces the risk for extrapyramidal symptoms when treating schizophrenic patients with antipsychotic drugs. ORG 5222 is a new potential antipsychotic drug which has high affinity for 5-HT2A, D2-dopamine and alpha 1 adrenergic receptors in vitro. The objective of this study was to examine if ORG 5222 occupies 5-HT2A and D2-dopamine receptors in human subjects in vivo. Central receptor occupancy was measured by positron emission tomography (PET) in three healthy subjects after sublingual administration of 100 micrograms ORG 5222. [11C]N-methylspiperone ([11C] NMSP) was the radioligand used to measure 5-HT2A receptor binding in the neocortex and [11C]raclopride to measure D2-dopamine receptor binding in the striatum. The 5-HT2A occupancy was 15-30% and the D2-dopamine receptor occupancy was 12-23%. The study confirms that ORG 5222 binds to 5-HT2A and D2-dopamine receptors in human brain. Since receptor occupancy of ORG 5222 is rather low, doses higher than 100 micrograms are suggested in future clinical trials to evaluate the antipsychotic drug effect of ORG 5222.

Determination of AJ-3941, a possible agent for the treatment of cerebrovascular disorders, in plasma and brain by means of high-performance liquid chromatography with fluorescence detection.

A sensitive and selective high-performance liquid chromatographic method with fluorescence detection is described for the determination of AJ-3941 (I), a possible agent for the treatment of cerebrovascular disorders, in plasma and brain tissue. A simple hexane extraction was used for plasma, and for brain homogenate the hexane extract was further purified by solid-phase extraction. The determination limit was ca. 3 ng/ml for both plasma (0.5 ml) and 10% (w/v) brain homogenate (1 ml). The method was applied to the determination of I in plasma and brain samples of experimental animals.

Biotransformation of the partial beta-agonist doxaminol in dog.

The biotransformation of the positive inotropic compound doxaminol (N-methyl-N-(2-hydroxy-3-phenoxy-propyl)-11-(2-amino-ethyl)-6,11- dihydrodibenz[b,e]oxepine, neutral fumarate; BM 10.188) was examined in bastard shepherd dogs. Metabolic products, formed by oxidative cleavage of various side chain carbon atoms of the molecule, as well as conjugated complexes with glucuronic and sulfuric acid, were isolated from urine and plasma. As main metabolites 2-hydroxy-3-phenoxy-propionic acid and phenoxyacetic acid were formed. By means of 1HNMR and 13C-NMR spectroscopy and various mass spectroscopic methods, the chemical structures of the metabolites were elucidated.

Pharmacokinetics of the partial beta-agonist doxaminol in dog.

The pharmacokinetic behaviour of doxaminol (N-methyl-N-(2-hydroxy-3-phenoxy-propyl)-11-(2-amino-ethyl)-6, 11-dihydrodibenz[b,e]oxepine, neutral fumarate; BM 10.188) was examined in dogs using peroral and intravenous application of the 14C-labelled drug. The maximum plasma concentration was reached 1 h after application, indicating a relatively quick absorption of doxaminol. Decrease of total radioactivity after intravenous and peroral application is characterized by two phases, the elimination half-lives being 1.33 and 1.55 h, respectively, and 24.05 and 21.05 h, respectively. The biological availability of doxaminol was ca. 60%. The plasma levels of the unchanged drug showed that doxaminol was very rapidly eliminated and metabolized. Within the examined period of 96 h, the elimination of doxaminol and its metabolites via urine and faeces amounted to 76.5% after intravenous application, and 44.1% of the applied dose after peroral application. The major amount of radioactivity is eliminated via faeces (61.5% and 31.2% of dose, respectively) while the elimination through urine is found to be 15.0 and 12.9% of the dose, respectively.

[A correlation between the species differences in anti-inflammatory activity of AD-1590, a non-steroidal anti-inflammatory drug, and its plasma level].

Authors have reported that the oral potency ratio of AD-1590 to indomethacin varies with the animal models employed; the ratio is 4, 2.3 and 31 in the tests of acetic acid-induced vascular permeability (male mice), carrageenan hind paw edema (male rats) and UV-erythema (female guinea pigs), respectively. Thus, the relationship between the difference in the anti-inflammatory activity of AD-1590 among animal models and the species difference of the plasma AD-1590 level was investigated in experimental animals in order to ascertain the cause of the difference in the potency ratio. Inhibitory potency of AD-1590 on UV-erythema and increased vascular permeability induced by acetic acid in male rats was about 2.1 and 2.3 times, respectively, that of indomethacin. On the other hand, after a single oral administration of 5 mg/kg, the highest plasma AD-1590 level was seen in female guinea pigs (AUC9-8 hr = 63.1 micrograms.hr/ml); and followed by that in mice (male, 32.1; female, 36.1) greater than male dogs (11.5) greater than or equal to rats (male, 9.02; female, 12.5), male rabbits (9.17) greater than male monkeys (9.34 at 6 mg/kg). Hucker et al. have reported that the plasma level of indomethacin in rats is several times higher than that in guinea pigs, rabbits and monkeys. These results suggest that most of the species difference in the relative potency of AD-1590 to indomethacin in the anti-inflammatory activity results from the species difference in the plasma level of both drugs.

Quantitative analysis of oxepinac in human plasma, urine and saliva by gas chromatography--mass fragmentography.

A sensitive and specific method is described for the quantitative analysis of 6,11-dihydro-11-oxo-dibenz[b,e]oxepin-3-acetic acid (oxepinac) in human plasma, urine and saliva. Oxepinac and internal standard are extracted from acidified plasma, urine or saliva, converted to the corresponding n-propyl esters and analysed by gas chromatography--mass fragmentography using selected ion monitoring. The method is accurate and precise over the range 100 microgram/ml to 1.0 ng/ml. The method has been applied to the analysis of plasma, urine and saliva from healthy volunteers receiving therapeutic doses of oxepinac.

Species differences in the disposition and metabolism of 6,11-dihydro-11-oxodibenz[be]oxepin-2-acetic acid (isoxepac) in rat, rabbit, dog, rhesus monkey, and man.

The disposition and metabolism of 6,11-dihydro-11-oxodibenz[be]oxepin-2-acetic acid (isoxepac), a new nonsteroidal anti-inflammatory agent, has been studied in rat, rabbit, dog, rhesus monkey, and man. Animals were given single oral or parenteral doses of 5 or 50 mg/kg; man received approximately 3 mg/kg orally. Fecal excretion of radioactivity occurred in the rat (26--37%) and dog (33--49%), whereas in the other species elimination was mainly urinary (less than 83%). Biliary excretion accounted for 18--52% of the dose in the rat and dog. Enterohepatic circulation was demonstrated in both species. Plasma of all species was found to contain mainly unchanged isoxepac. The compound was rapidly eliminated from plasma of dog, rhesus monkey and man, but was more slowly eliminated in rat and rabbit. In the rabbit and dog the principal metabolites were the glycine and taurine conjugates of isoxepac, respectively, whereas in the rhesus monkey and man, isoxepac was excreted unchanged or as the glucuronide.

Quantitative analysis of 6,11-dihydro-11-oxo-dibenz[b,e] oxepin-2-acetic acid (isoxepac) in plasma and urine by gas--liquid chromatography.

A method is described for the quantitative analysis of 6,11-dihydro-11-oxo-dibenz[b,e]-oxepin-2-acetic acid (isoxepac) in plasma and urine. Isoxepac and internal standard was analysed by gas-liquid chromatography using a flame ionization detector. The method is accurate and precise over the range 0.1--30 microgram/ml. The method has been applied to the analysis of plasma and urine from both healthy volunteers and patients receiving therapeutic oral doses of isoxepac.

Acute, subacute and chronic toxicity of oxepinac.

The acute, subacute and chronic toxicity of 6,11-dihydro-11-oxodibenz[b,e]oxepin-3-acetic acid (oxepinac) was investigated in several animal species. The LD50 value was lower in rats than in rabbits, mice and dogs. The major cause of death was perforative ulcer in the gastrointestinal tract. Long-term study in rats revealed that oxepinac produced no hematological, blood chemical and pathological changes except for minor anemia and fatal ulcer formation occurring predominantly in females. Oxepinac proved to be less toxic than indometacin in chronic toxicity in rats.