Use of High Performance Liquid Chromatography (HPLC) for the Analysis of Amino Acid of Sulawesi and Local Clone Cocoa Bean Fermentation

Fermentation is a very vital stage of processing mechanism to ensure the chocolate products have good taste. This study was conducted to obtain an optimal result of chocolate fermentation by determining the concentration and types of amino acid of Sulawesi and local clone cacao beans using HPLC (High Performance Liquid Chromatography) method. A Randomized Completely Design was used in this study. Two types of cocoa clones, Sulawesi clone and Local clone, were divided into 5 groups of treatment: without fermentation (control), fermentation for 3 days, 4 days, 5 days and 6 days. The analysis of amino acids was conducted using HPLC separation method based on the procedure at Marino et al. (2010), Nollet (1996). The measurement of amino acid performed in two phases, liquid hydrolysis, and derivatization proceeded by chromatographic analysis. Condition of HPLC was measured at 37 °C. Mobile phase contains of 60% acetonitril - AccqTag Eluent A, gradient system and the flow rate was 1.0 ml per minute. Fluorescence detector has 250 nm excitation and 395 nm emission. Injecting volume was 5 uL. The results of this study show that cocoa beans of Sulawesi clone with 6-days fermentation has higher products of aspartic acid, glutamic acid, hydrophobic amino acids (Alanine, leucine, proline, valine, isoleucine) and amino acids such as serine, glysine, histidine, treonine and lysine, while local clones of cocoa beans with 3-days fermentation produce more amino acids such as aspartic, glutamic, hydrophobic (isoleucine, leucine, valine) and amino acids such as histidine, threonine, glysine, serine and lysine.