Optimization of ultrasound assisted extraction of pectin from black mulberry (Morus nigra.L) pomace

Pectin is a complex heteropolysaccharide presented naturally in the middle lamella and primary cell walls of plant tissues. Pectin is mostly composed of a linear backbone of α-(1, 4) linked D-galacturonic acid residues which are partially esterified with methyl alcohol or acetic acid at the carboxylic acid. Pectin is widely used as a gelling agent in food systems such as jams and jellies, fruit juices, confectionaries, bakery fillings and as a stabilizer in acidified milk drinks. Black mulberry (Morus nigra.L), originating from Iran, is a juicy fruit with dark red color and slightly acidic flavor. Black mulberry has gained an important position in fruit markets and food industry due to its distinctive flavor and phytonutrients in recent years. Black mulberries can be consumed as fresh or processed into several products such as juice, marmalades, liquors, natural dyes or even be used as frozen fruit in ice cream production. However, due to short harvesting season and susceptibility to spoilage, black mulberries are mostly processed into juice besides its fresh consumption. Press cake residue or pomace is the main by-product which is generated in large amounts from the commercial juice extraction processes from berry fruits which contain several bioactive compounds and valuable ingredients including antioxidants, phenolics, pigments, pectin and so on. These berry pomaces are considered suitable for pectin recovery although their pectin content is somewhat lower in comparison to rich known sources of pectin. Recently, ultrasound assisted extraction technique, as a novel method of extraction, has gained remarkable attention due to its some advantageous effects including shorter extraction time, reduced solvent and energy consumption and higher yield and better quality of extracted compounds as compared to conventional acid extraction method. Therefore, the present work was aimed to evaluate and optimize the ultrasound assisted extraction parameters (time of extraction, ultrasound power and liquid/solid ratio) to maximize the extraction yield, degree of esterification and galacuronic acid content of pectin from black mulberry pomace using three level three factor face centered central composite design.
Fresh and fully ripe black mulberry fruits were purchased from a local fruit market in Karaj (Iran) and subsequently pressed to separate pomace from juice. The obtained pomace was then vaccuum dried (until 7.8±1% moisture content), ground to obtain a homogenous powder, sieved, filled into polyethylene containers, and stored at refrigerator. Pectin extraction from this powdered pomace was carried out using an ultrasonic bath at operating frequency of 35 kHz under different parameters including extraction time (20-60 min), ultrasonic intensity (40-100%) and liquid-solid ratio (15:1 to 30:1 mL/g). The initial pH value of the solution was adjusted to 2.0 by adding 1.0 N HCl solution. At the end of extraction, extracts were quickly cooled, centrifuged, filtered using Buchner funnel, concentrated by 5 fold, coagulated with 96% ethanol (ER=1.5) and left for 8 hours in room temperature. The precipitated pectin was separated by Buchner funnel, rinsed twice with 96% ethanol, dried under vacuum (45°C) and finally powdered. The degree of esterification of pectin was determined by titrimetric method according to Bochek et al. (2001). The galacturonic acid content was quantified by the colorimetric method with m-hydroxydiphenyl reagent using a spectrophotometer at 520 nm. The intrinsic viscosity [ƞ] of pectin was determined using a capillary tube viscometer and accordingly the viscosity average molecular weight [Mv] was calculated from the Anger-Berth equation. Finally, the frequency sweep test was performed using a controlled stress rheometer to determine the values of storage modulus (G′) and loss modulus (G″). The experimental design and statistical analysis were performed using response surface methodology (RSM).
According to the obtained results, the extraction yield, degree of esterification and galacturonic acid content of pectin varied in the range of 4.73-7.68%, 42.42-58.33 and 18.70-31.45%, respectively. The extraction yield of pectin was linearly affected by the extraction conditions and its value significantly increased with increasing all the studied independent variables (p