Title Optimizacija postupka pripreme uzorka za kromatografsku analizu N-nitrozamina Title (english) Optimization of the sample preparation procedure for chromatographic analysis of N-nitrosamine Author Martina Vuković Mentor Sandra Babić (mentor)Committee member Sandra Babić (predsjednik povjerenstva)Committee member Irena Škorić (član povjerenstva)Committee member Kristina Tolić Čop (član povjerenstva)Granter University of Zagreb Defense date and country 2023-07-07, Croatia Scientific / art field, NATURAL SCIENCES Abstract N-nitrozamini su spojevi opće kemijske strukture R1N(-R2)-N=O, a nastaju reakcijama oksidacije sekundarnih ili tercijarnih amina uz pomoć nitrozacijskog sredstva. Pokazujući mutageno i kancerogeno djelovanje na različite organe i tkiva kod ljudi, posljednjih godina postaju predmetom brojnih analitičkih ispitivanja. Upravo provođenje analitičkih ispitivanja predstavlja ključ za sprječavanje nastanka N-nitrozamina i njihova uklanjanja iz vode. U ovome radu ispitivano je sedam N-nitrozamina: N, N-dibutilnitrozni amid (NDBA), N, N-dietilnitrozni amid (NDEA), N, N-dimetilnitrozni amid (NDMA), N, N-dipropilnitrozni amid (NDPA), 4-nitrozomorfolin (NMOR), 1-nitrozopiperidin (NPIP) te 1-nitrozopirolidin (NPYR). Kako bi se zadovoljio cilj rada u vidu optimizacije postupka pripreme uzorka za kromatografsku analizu N-nitrozamina primijenjen je Box-Behnkenov dizajn eksperimenata s tri numeričke varijable na tri razine u kombinaciji s metodom odzivnih površina (RSM). Uzorci vode pripremljeni su ekstrakcijom čvrstom fazom (SPE) uz upotrebu kokosova ugljena kao sorbensa. Nakon provedbe numeričke optimizacije kao optimalni uvjeti ekstrakcije čvrstom fazom primijenjeni su volumen uzorka od 300 ml, volumen eluensa od 12 ml te vrijeme sušenja sorbensa od 10 min. Provedena su dva eksperimenta pri optimalnim uvjetima, pri čemu je jedan uključivao uparavanje otapala iz uzorka na rotacijskom uparivaču, a drugi uparavanje otapala iz uzorka u struji dušika. Prisutnost N-nitrozamina u dobivenim ekstraktima u oba slučaja potvrđena je tekućinskom kromatografijom visoke djelotvornosti uz detektor s nizom dioda (HPLC-DAD) gdje je primjenom gradijentnog eluiranja uz protok od 0,5 ml/min provedena uspješna separacija svih sedam N-nitrozamina na kromatografskoj koloni Kinetex C18 uz korištenje MilliQ vode i acetonitrila kao pokretnih faza. Uspoređujući dobivene odzive odnosno vrijednosti učinkovitosti ekstrakcije moguće je zaključiti kako uparavanje u struji dušika biva učinkovitije za NDMA, NDEA, NDPA i NDBA u odnosu na uparavanje na rotacijskom uparivaču. Gledano s ekonomskog aspekta optimizacijom SPE postupka osigurava se ušteda reagensa i potrošnog materijala, ali i utrošenog vremena samoga znanstvenika.
Abstract (english) N-nitrosamines are compounds with the general chemical structure R1N(-R2)-N=O that are formed by oxidation reactions of secondary or tertiary amines with the help of a nitrosating agent. Showing mutagenic and carcinogenic effects on various organs and tissues in humans they have become the subject of numerous analytical tests in recent years. Analytical testing is essential to prevent the formation of N-nitrosamines and their removal from water. In this study, seven N-nitrosamines were analysed: N, N-dibutylnitrous amide (NDBA), N, N-diethylnitrous amide (NDEA), N, N-dimethylnitrous amide (NDMA), N, N-dipropylnitrous amide (NDPA), 4-nitrosomorpholine (NMOR), 1-nitrosopiperidine (NPIP) and 1-nitrosopyrrolidine (NPYR). In order to meet the goal of the study in the form of optimization of the sample preparation procedure for the chromatographic analysis of N-nitrosamines, the Box-Behnken design of experiments with three numerical variables at three levels in combination with the response surface methodology (RSM) was applied. Water samples were prepared by solid phase extraction (SPE) using coconut charcoal as a sorbent. After numerical optimization, a sample volume of 300 ml, an eluent volume of 12 ml and a sorbent drying time of 10 min were used as the optimal conditions for solid phase extraction. Two experiments were performed at optimal conditions, one involving the evaporation of the solvent from the sample on a rotary evaporator, and the other the evaporation of the solvent from the sample in a stream of nitrogen. The presence of N-nitrosamines in the obtained extracts in both cases was confirmed by high-performance liquid chromatography with diode array detector (HPLC-DAD) by applying gradient elution at a flow rate of 0,5 ml/min. A successful separation of all seven N-nitrosamines was carried out on the Kinetex C18 chromatographic column using MilliQ water and acetonitrile as mobile phases. By comparing the obtained responses and extraction efficiency values, it is possible to conclude that evaporation in a nitrogen stream is more efficient for NDMA, NDEA, NDPA and NDBA compared to evaporation on a rotary evaporator. From an economic point of view, the optimization of the SPE procedure ensures the saving of reagents and consumables, as well as the time spent by the scientist.
Keywords
N-nitrozamini
optimizacija
Box-Behnkenov dizajn
metoda odzivnih površina
ekstrakcija čvrstom fazom
tekućinska kromatografija visoke djelotvornosti
Keywords (english)
N-nitrosamines
optimization
Box-Behnken design
response surface methodology
solid phase extraction
high-performance liquid chromatography
Language croatian URN:NBN urn:nbn:hr:149:041194 Study programme Title: Materials Science and Engineering - Graduate study Type of resource Text File origin Born digital Access conditions Embargoed access Terms of use Created on 2023-07-26 13:36:56
