QUANTIFICATION OF PHENOL IN BLOOD AND URINE SAMPLES BY USING GC-MS METHOD

Ciljevi istraživanja: Razviti metodu određivanja fenola plinskom kromatografijom s masenim detektorom, ispitati optimalne uvjete rada GC-MS-a pri kojima je moguća kvantifikacija fenola te odrediti koncentraciju fenola u biološkom uzorku mokraće. Mjesto istraživanja: Kemijsko-toksikološki laboratorij Kliničkog odjela za sudsku medicinu Zavoda za patologiju, sudsku medicinu i citologiju, KBC Split. Materijali i metode: Pripremljena su četiri miksa standardnih otopina postupnim razrjeđivanjem fenola s kloroformom. Odabrane su tri skupine instrumentalnih parametara (metoda 1, 2 i 3) kako bi se utvrdilo kojom metodom se dobije najbolji odziv signala na kromatogramu. U tri biološka uzorka mokraće dodan je proizvoljno određen volumen MIX-a 2. Uzorci su za analizu obrađeni ekstrakcijom na čvrstoj fazi, SPE, i analizirani na GC-MS-u. Rezultati: Provedenom analizom jednake koncentracije standarda najbolji odziv signala na detektoru ostvaren je metodom 3 s parametrima analize: volumen injektiranja:1 μl, temperatura injektora:250 °C, protok plina nosioca:1,54 ml/min i ukupno trajanje temperaturnog programa:10 minuta (60 °C 1 minutu, 10 °C/min do 170 °C). Analizom uzoraka mokraće uspješno je detektiran i kvantificiran fenol u dva od tri uzorka. Dobivena koncentracija fenola u uzorku 1 bila je 593,74 ng/ml, a u uzorku 2 1080 ng/ml. Zaključak: Odabranim instrumentalnim parametrima moguće je detektirati i kvantificirati fenol iz bioloških uzoraka mokraće. SPE je vrlo dobra tehnika predobrade uzorka, ali su potrebne dodatne analize da bi se utvrdila količina nečistoća koje mogu interferirati u analizi. Osim toga, potrebno je provesti dodatne analize kako bi se potvrdila primjenjivost same metode na realne biološke uzorke mokraće.Objectives: Objectives of this study were to develop a method for determination of phenol with gas chromatography-mass spectrometry, determine optimal parameters of the instrument and to quantify the phenol in three different biological samples of urine by using the selected method. Settings: LABORATORY OF TOXICOLOGY, DEPARTMENT OF PATHOLOGY, MEDICINE AND CYTOLOGY, University Hospital of Split. Materials and methods: Four mixes of standard solutions were prepared by gradually diluting phenol with chloroform. Three different groups of instrumental parameters were selected (method 1, 2 and 3) to determine which method gives the best signal on the chromatogram. A voluntary defined volume of MIX 2 was injected in three urine samples. The samples were prepared for the analysis by using solid phase extraction and analyzed with GC-MS method. Results: By analysing three equal concentrations of standard solution using all three methods, it was noticed that method 3 gives the best signal on the detector. Parameters of analysis for method 3 were: injection volume:1 μl , injector temperature:250 °C, carrier gas flow (helium):1,54 ml/min, program time:10 minutes (60 °C for 1 minute and 10 °C per minute til 170 °C). Two out of three urine samples were successfully detected and quantified with phenol. Sample 1 had the phenol concentration of 593,74 ng/ml and sample 2 1080 ng/ml. Conclusion: It is possible to detect and quantify the phenol from the biological samples of urine by using selected method 3. SPE is a good technique of sample preparation, but additional analysis are neccessary to establish the quantity of interferences that drop behind and can affect the analysis.It is also neccessary to carry out additional analysis on real biological samples to be able to confirm these techniques of sample preparation and analysis