Optimization of aqueous extraction of fructan from tubers of Eremurus spectabilis using box-behnken design

Fructans are an important product of the industry of prebiotics. In addition to their interesting nutritional and health benefit properties, fructans are also used in food formulations for their techno-functional properties such as fat substitute, bulk agent, water retention. Serish (Eremurus spectabilis) belongs to the family of Liliaceae and geographically distributed in the region of South Asia and Central Asia. Their roots accumulate high levels of fructans during their growth and are traditionally used to cure jaundice, liver disorders, stomach irritation, pimples and bone fractures and even as a glue for industrial application. Recently fructan extraction from numerous plants and fungus has drawn the attention of researchers. Also, several methods for fructan extraction have been developed such as hot-water extraction, precipitation by alcohol and ultrasound-assisted extraction. To the best of our knowledge, there are no reports on the fructan extraction from Serish. The present study is considered the first attempt aiming to determine the optimal conditions for water extraction of Serish fructans.
The Serish root powders were obtained from the local medical plant market, Mashhad, Iran. Moisture, ash, fat, protein, Carbohydrate and total dietary fiber were determined according to standard AOAC methods. All variables were examined in triplicate. Conventional extraction was carried out in a water bath. Total carbohydrate was assayed colorimetrically using the phenol–sulphuric acid method. The concentrations of soluble reducing sugars were measured using a 3,5-dinitrosalicylic acid (DNS) method. The fructan content was measured by the difference between total carbohydrate and reducing sugars. The percentage of fructan yield (%) was evaluated based on equation of Lingyun, Jianhua, Xiaodong & Yalin (2007). The purity was evaluated according to the method of Paseephol, Small & Sherkat (2007). As an index of degree of polymerization, the average chain length, was calculated according to Lingyun, Jianhua, Xiaodong & Yalin (2007). A Box-Behnken design was constructed using the software Design Expert Version 6.0.10 and was used for estimating the effect of independent variables on the extraction parameters. Three extraction variables considered for this research were x1 (extraction time), x2 (extraction temperature) and x3 (water: solid) for conventional extraction method. Lack of fit, coefficients of determination (R2), adj-R2, coefficient of variation (CV) and significant probabilities were calculated to check the model adequacy. Optimization was based on generation of the best results for fructan extraction. In order to determine the validity of the model, the experimental and predicted values were compared by paired t-test.
It has a marked amount of protein and fats. The ash value is relatively high, suggesting an important mineral content. The composition of total fiber suggests its possible use as a source of dietary fiber for enrichment of foods. In addition, results show that Serish root powder is a polysaccharide-rich material. The results indicated that extraction temperature has the most effect on the extraction yield. Increase of time, temperature and water to solid ratio led to significantly increase in extraction yield. Considering the significant quadratic term of extraction time, it becomes clear that yield rises as extraction time increase from 0 to 22.5 min but it decreases at higher times. The yield increase between 0 to 22.5 min might be due to the time requirement for contact of fructan to the release medium where the liquid penetrated into the Serish powder, dissolved the fructan and subsequently diffused out from the root. On the other hand, the yield decrease after 22.5 min may be ascribed to degradation of fructan to free sugar and enhancement of impurities extraction at higher times. When extraction time goes by certain threshold, the yield started to decrease. The yield showed a large tendency to increase when the extraction temperature was increased. This is maybe due to the enhancement of the mass transfer resulting from the increased solubility of fructan and the decreased viscosity of the solvent. Yield was increased by portion of water to solid. This might be attributed to the availability of liquid that increases the driving force of fructan out of the root. Based on the sum of squares, temperature had the most effect on degree of polymerization. The results indicated that increase of time, temperature and W/S ratio resulted in increase of the degree of polymerization which might be due to the enhancement of overall carbohydrates extraction. On the other hand, the degree of polymerization of extracted fructan was lower than 10. This is probably because the disruption of fructan branch to reducing sugars with increasing temperature that leads to chain length decrease. Thus, the produced oligosaccharides could be used as sweetener. W/S ratio had the most effect on purity. The results showed that increase of time, temperature and W/S ratio resulted in increase of the purity which might be due to the enhancement of overall carbohydrates extraction. With the increase in extraction time, the purity of extract increased gradually, but decreased after the purity reached a maximum at 22.5 min. This decrease may be ascribed to degradation of fructan to free sugar and enhancement of impurities extraction at higher times. Multiple response optimizations were performed to measure the optimum levels of independent variables to achieve the desired response goals. Extraction yield and purity were desired maximal.
The final results for this optimization was found to be extraction time of 28.38 min, extraction temperature of 88ºC and water to solid ratio of 50:1 v⁄w. Response surface methodology was an efficient statistical tool to model the influence of extraction conditions of fructan from Serish root powder on the extraction yield. These results also suggested that by modifying the proportion of these components, a large range of variations may be obtained. There was a good agreement between the experimental data and their predicted counterparts, showing the effectiveness of the proposed conditions and reliability of Box–Behnken analysis on fructan precipitation.