Solid Phase Extraction Purification of Saliva Samples for Antipsychotic Drug Quantitation

Abstract

Saliva is far less popular as a diagnostic material than blood. This has resulted in a lack of procedures for the sampling and handling of saliva, e.g., effective ways to purify endogenous compounds from saliva to enable a simultaneous determination of xenobiotics such as neuroleptics. Therefore, the aim of this study was to develop an analytical procedure to purify saliva samples so that it is then possible to simultaneously determine five neuroleptics (aripiprazole, clozapine, olanzapine, quetiapine and risperidone) and the antiepileptic drug carbamazepine, and their respective metabolites (dehydroaripiprazole, N-desmethylclozapine, N-demethylolanzapine, norquetiapine, 9-OH-risperidone and carbamazepine-10,11-epoxide). A study of three types of solid-phase extraction (SPE) columns showed that the purest eluates were obtained using columns containing ion exchange sorbent. The sorbents were first washed with water then with a mixture of water and methanol (1:1), and the adsorbed residue was eluted with a 5% ammonia solution in methanol. Saliva samples for SPE were diluted with 2% formic acid and a mixture of methanol and water (1:1). This procedure was developed to purify a saliva sample spiked with a mixture of neuroleptics and carbamazepine, and their respective metabolites. A chromatographic analysis confirmed the isolation of all compounds, indicating that this procedure can be used in further development and validation for a method designed to monitor the levels of neuroleptic drugs in saliva and to monitor their uptake by patients.

Deproteinization as a Rapid Method of Saliva Purification for the Determination of Carbamazepine and Carbamazepine-10,11 Epoxide

“…The methods for the determination of anti-epileptic drugs in blood have often been modified to determine carbamazepine in saliva [2,3,6,8,9,17,18]. Most often, saliva samples were 1 mL [14,18,22,23,25] or more [3,6,8], and were rarely less than 1 mL [15,21,24,27]. The method developed in this study allows for the determination of carbamazepine and its active metabolite, in as little as 200 µL of saliva.…”

“…This allows for the use of a small sample volume (200 µL). Preliminary studies showed that the use of Strata X-C columns, which were rinsed with water and a mixture of water and methanol (1:1), and saliva samples diluted with 2% formic acid solution and a mixture of water and methanol (1:1), is a good procedure for purifying saliva samples [22]. Applied to the analysis of neuroleptics and carbamazepine, it allowed for good saliva purification and an analysis of patient samples.…”

“…Saliva sampling for the analysis of the carbamazepine content is most often taken following stimulating its secretion by chewing Parafilm [3,9,14] or eating citric acid [15][16][17][18]. Saliva samples can also be collected in a tube without prior stimulation [8][9][10][11][12][13][14][15][16][17][18][19][20] or on cotton swabs specially designed for this purpose, which are placed in the mouth to soak up the saliva [21][22][23]. After removing the swab from the mouth, it is placed in a tube and is centrifuged.…”

“…The most common method of carbamazepine isolation from saliva samples is liquid-liquid extraction [8,14,16,[24][25][26]. Saliva samples are also purified using a modified liquid-liquid extraction, cloude-point extraction [26], solid phase extraction [22,23], and saliva deproteinization [15,20].…”

“…Carbamazepine in saliva can be quantified by an enzymatic method (FPIA-fluorescence polarization immunoassay) [7,19], as well as by separation techniques, gas chromatography [2,16,27], and liquid chromatography coupled with UV detection [3,8,9,14,15,17,18,[20][21][22][23][24][25][26].…”