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Pharmacological effect of a drug depends on its free fraction in plasma. By certain drugs, like immunosupresive mycophenolic acid, concentrations below therapeutic lead to rejection of the transplanted organ while concentrations above therapeutic lead to toxic effects, primarily on kidney and liver. For this reason constant monitoring of mycophenolic acid in plasma is necessary. Concerning frequency of sampling, availability and non-invasive way of sampling, saliva presents one of the best biological materials for analysis. On the other hand, numerous interfering endogen substances, impurities and high viscosity of saliva make this biological material difficult for purification. Mycophenolic acid has to be purified and concentrated from saliva to be consequently determined applying appropriate analytical method. In this paper solid phase extraction method for purification of saliva has been presented. The efficacy of solid phase extraction has been influenced by many different experimental parameters, so these parameters has to be strongly controlled, and protocol of extraction procedure has been proposed. In this way conditioning of polymeric sorbent has been done with methanol and purified water, samples have been applied together with 0.1 % formic acid, washing has been done with purified water and elution of mycophenolic acid has been performed with acetonitrile and 0.2% formic acid. Farmakološki efekti leka direktno zavise od koncentracije leka u plazmi, odnosno njegove slobodne frakcije. Kod određenih lekova, kao što je imunosupresiv mikofenolna kiselina, smanjenje koncentracije dovodi do izostanka terapijskog efekta, dok povećanje koncentracije može dovesti do toksičnih efekata. Ozbiljni neželjeni efekti mogu dovesti do fatalnih ishoda, bilo da se radi o odbacivanju transplantiranog organa ili ozbiljnom oštećenju bubrega i jetre. Iz tog razloga potrebno je sprovoditi stalno praćenje nivoa mikofenolne kiseline u krvi. Imajući u vidu učestalost uzorkovanja, dostupnost i neinvazivan način sakupljanja, saliva ima prednost nad plazmom zbog čega se u skorije vreme sve više koristi kao uzorak. Međutim, zbog brojnih interferirajućih endogenih supstanci, nečistoća i viskoznosti, salivu je neophodno prethodno obraditi za kvalitativno i kvantitativno praćenje mikofenolne kiseline UPLC metodom. U ovom radu je opisana upotreba čvrsto-tečne ekstrakcije kao metode izbora za pripremu uzoraka salive. Pošto na efikasnost ekstrakcije utiču različiti eksperimentalni uslovi koje je potrebno strogo kontrolisati, predložen je protokol koji podrazumeva: kondicioniranje polimernog kertridža za čvrsto-tečnu ekstrakciju metanolom i vodom, nanošenje uzorka zajedno sa 0,1 % mravljom kiselinom, ispiranje kertridža vodom i eluiranje mikofenolne kiseline smešom acetonitrila i 0,2% mravlje kiseline.

There are several techniques like time of flight secondary ion mass spectrometry (ToF SIMS) that require a special protocol for preparation of biological samples, in particular, those containing single cells due to high vacuum conditions that must be kept during the experiment. Frequently, preparation methodology involves liquid nitrogen freezing what is not always convenient. In our studies, we propose and validate a protocol for preparation of single cells. It consists of four steps: (i) paraformaldehyde fixation, (ii) salt removal, (iii) dehydrating, and (iv) sample drying under ambient conditions. The protocol was applied to samples with single melanoma cells i.e. WM115 and WM266-4 characterized by similar morphology. The surface and internal structures of cells were monitored using atomic force, scanning electron and fluorescent microscopes, used to follow any potential protocol-induced alterations. To validate the proposed methodology for sample preparation, ToF SIMS experiments were carried out using C 60 + cluster ion beam. The applied principal component analysis (PCA) revealed that chemical changes on cell surface of melanoma cells were large enough to differentiate between primary and secondary tumor sites. Subject category : Mass spectrometry.