Method optimization for quantification of nicotine in tobacco by gas chromatography-mass spectrometry

Duhan, Nicotiana tabacum, porodice Solanaceae, jedan je od najčešće konzumiranih sredstava ugode i ovisnosti, najčešće pušenjem. Razlog njegove ovisnosti je nikotin, kao jedan od glavnih sastojaka samoga duhana. Ovisno o vrstama duhana kao što su Virginia, Burley, Maryland ili Oriental, ovisit će i koncetracija nikotina u duhanu. Nikotin djeluje na nikotinske acetilkolinske receptore (nAChR) stvarajući ovisnost, ali i brojne fiziološke te patofiziološke učinke. Često ilegalno trgovanje duhanom, kao i njegova uzročnost brojnih požara, uz brojne druge razloge, dovode do potrebe optimizacije metode kvantitativnog određivanja nikotina u duhanu. Tako je metoda pripreme uzoraka duhana za kvantitativno određivanje nikotina u duhanu optimizirana Box-Behnkenovim dizajnom temeljenim na metodi odzivne površine. Korištenjem tri varijable u spomenutom dizajnu, udjela metanola u otapalu za ekstrakciju, vremena soniciranja te broja ekstrakcija, dobivene su optimalne vrijednosti navedenih parametara za pripremu uzoraka duhana. Kvantitativno određivanje nikotina odrađeno je pomoću plinske kromatografije sa spektrometrijom masa (GC-MS) koja je validirana provjerom brojnih validacijskih parametara te zadovoljila postavljene kriterije prihvatljivosti. Razvijena metoda pokazala se prikladnom za navedenu svrhu kvantitativnog određivanja nikotina u duhanu te će se moći koristiti u Centru za forenzična ispitivanja, istraživanja i vještačenja „Ivan Vučetić'', MUP-a RH.Tobacco, Nicotiana tabacum, Solanaceae, is one of the most widely used stimulants and addictive substances, consumed mainly by smoking. The reason for its addictive nature lies in nicotine, one of the main components of tobacco. Depending on the type of tobacco, such as Virginia, Burley, Maryland or Oriental, the nicotine concentration in tobacco also varies. Nicotine is a supstrate of nicotine acetylcholine receptors (nAChR), which are addictive and cause many other physiological and patophysiological effects. Illegal tobacco and its causality in numerous fire incidents, among many other reasons, have led to the need to optimize the method of quantitative analysis of nicotine in tobacco. The method of preparing tobacco samples for quantitative analysis of nicotine in tobacco was optimized by Box-Behnken design based on response surface modeling (RSM). In this design, three parameters were used, namely the percentage of methanol in the extraction solution, the duration of sonication, and the number of extractions for which optimal results were obtained, which were then used to prepare the tobacco samples. The quantitative analysis of nicotine in tobacco samples was performed using Gas Chromatography-Mass Spectrometry which was validated by checking numerous validation parameters that met the established acceptance criteria. The method was thus found to be suitable for the quantitative analysis of nicotine in tobacco and will be used at the Ivan Vučetić Forensic Science Center in Zagreb, Croatia