| Abstract | Cilj ovoga istraživanja bio je uvesti analitičku metodu koja bi omogućila brzo, pouzdano i precizno određivanje koncentracije limunske kiseline u uzorcima komercijalno pribavljenih voćnih sokova. Za određivanje koncentracije limunske kiseline korištena je tekućinska kromatografija visoke djelotvornosti (HPLC) s UV/Vis detektorom. Istraživanje je provedeno na HPLC (Knauer, Berlin, Njemačka) koji se sastojao od izokratne pumpe, UV/Vis detektora, manualnog injektora i analitičke kolone obrnute faze dimenzija 125,0x4,6 mm s punilom veličine 5 μm (LiChrospher RP-18, Merck, Darmstadt, Njemačka). Mobilna faza bila je 50 mM otopina fosfatnog pufera pH 2,8. Protok mobilne faze bio je podešen na 0,5 mL/min što je stvaralo tlak od 16,7 MPa. Valna duljina detekcije na UV/Vis detektoru bila je podešena na 214 nm. Iz osnovne matične otopine limunske kiseline (2,0 g/L) razrjeđenjem s MilliQ vodom pripremljen je niz standardnih otopina limunske kiseline u koncentracijskom rasponu od 0,125 do 1,0 g/L. Pri opisanim uvjetima na HPLC-UV/Vis vrijeme zadržavanja limunske kiseline na koloni iznosilo je oko 3 minute, a ukupna analiza trajala je 10 minuta. Dobivena kalibracijska krivulja je bila pravac s jednadžbom y = 223,02x - 4,2652 i pripadajućim koeficijentom korelacije R2 = 0,9998. Ponovljivost izražena kao relativna standardna devijacija (RSD) iznosila je manje od 5% za sve ispitane standardne otopine. Granica dokazivanja bila je 0,02 g/L, a granica određivanja 0,06 g/L, što pokazuje da je metoda vrlo osjetljiva. Mjerenjem koncentracije limunske kiseline u komercijalno pribavljenim uzorcima voćnog soka (n = 5) dobivene su vrijednosti u rasponu od 1,6 g/L (uzorak 5) do 3,98 g/L (uzorak 2), pri čemu je u dva uzorka koncentracija limunske kiseline bila viša od dopuštene (3,0 g/L; Uredba komisije EU br. 1129/2011). Iz dobivenih rezultata može se zaključiti da je razvijena metoda brza, pouzdana i precizna te se može koristiti za određivanje koncentracije limunske kiseline u uzorcima voćnih sokova. S obzirom da je u dva uzorka koncentracija limunske kiseline prelazila dopuštenu koncentraciju važno je pratiti njenu koncentraciju u proizvodima namijenjenima konzumaciji ljudi. |
| Abstract (english) | The aim of this research was to introduce an analytical method that would allow fast, reliable and precise determination of citric acid concentration in samples of commercially obtained fruit juices. To determine the concentration of citric acid, high performance liquid chromatography (HPLC) with UV/Vis detector was used. This research was conducted on HPLC (Knauer, Berlin, Germany) consisted of isocratic pump, UV/Vis detector, manual injector and reverse phase analytical column (dimension: 125.0 x 4.6 mm, particle size: 5 μm; LiChrospher RP-18, Merck, Darmstadt, Germany). The mobile phase was 50 mM phosphate buffer with pH adjusted to 2.8. The mobile phase flow was 0.5 mL/min and the resulting pressure 16.7 MPa. Wavelength on the UV/Vis detector was set at 214 nm. From the citric acid stock solution (2.0 g/L) by diluting with MilliQ water a set of working standards solutions was prepared in a concentration range of 0.125 to 1.0 g/L. In described HPLC conditions, the citric acid retention time was approximately 3 minutes and the analysis lasted 10 minutes. Calibration curve was linear (equation: y = 223.02x – 4.2652; correlation factor: R2= 0.9998). Repeatability expressed as relative standard deviation (RSD) was less than 5% for all tested working standard solutions. The limit of detection was 0.02 g/L and the limit of quantification was 0.06 g/L, indicating that developed method is very sensitive. The concentration of citric acid in commercially provided fruit juice samples (n = 5) ranged from 1.6 g/L (sample 5) to 3.98 g/L (sample 2). In two samples the concentration of citric acid was higher than 3.0 g/L as recommended by Commission regulation EU No. 1129/2011. From the results it can be concluded that the developed method is fast, reliable and accurate and can be used to determine the concentration of citric acid in fruit juices. Since in two samples of commercially obtained fruit juices citric acid concentration exceed the recommended concentration, it is important to monitor its concentration in products intended for human consumption. |
