Optimization of the sample preparation for analysis of polyphenolic compounds in the grape skins by high performance liquid chromatography

Cilj ovoga rada bio je optimirati uvjete priprave uzoraka kao i metode za ekstrakciju polifenola kožica grožđa. Ispitan je utjecaj različitog načina sušenja kožica bobica grožđa, sušenje u pećnici, na zraku pri sobnoj temperaturi te liofilizacija, na maseni udio pojedinih fenola te je utvrđeno da je liofilizacija najučinkovitija metoda. Optimiran je postupak zamrzavanja i odmrzavanja uzoraka bobica grožđa pri čemu su zamrzavane bobice sa i bez jastučića. Kožica je odvojena od ostatka bobice sa još zamrznutih bobica te nakon odmrzavanja u mikrovalnoj pećnici, hladnjaku i na sobnoj temperaturi. Najveći maseni udjeli pojedinih fenola očuvani su kod uzoraka zamrznutih sa jastučićem, a sa kojih je kožica odvojena s još zamrznutih bobica. Provedeno je optimiranje pet metoda (ekstrakcija čvrsto-tekuće, ultrazvukom potpomognuta ekstrakcija, mikrovalovima potpomognuta ekstrakcija, enzimima potpomognuta ekstrakcija te raspršenje matrice uzorka kroz čvrstu fazu) za ekstrakciju flavonoida iz kožica bobica grožđa te je usporedbom masenih udjela flavonoida u ekstraktima dobivenim tim metodama utvrđeno da je ultrazvukom potpomognuta ekstrakcija najučinkovitija metoda.The aim of the present study was the optimization of the sample preparations of grape skins and the extraction methods for polyphenols, as well. The investigation was done into the effect of the different drying methods of grape skins (oven drying, room temperature drying and freeze-drying) on the content of phenolic compounds. The most effective drying method was found to be freeze-drying. The process of freezing and thawing grape samples was undertaken. The grape berry samples were frozen with and without a receptacle. The skin of the frozen berries was separated from the rest of the berries after thawing in a microwave oven, a refrigerator and on room temperature. The largest content of individual phenols was preserved in samples frozen with a receptacle, where the skin was separated from the still frozen berries. The optimization was done for the five methods (solid-liquid extraction, ultrasound assisted extraction, microwave assisted extraction, enzyme assisted extraction and matrix solid phase dispersion) of flavonoid extraction from the grape berry skins. The comparison of flavonoid content in the extracts obtained by the five methods showed that the ultrasound assisted extraction was the most effective method