Maximizing Yield of Phenolic Compounds Extracted from White Turmeric Through Extraction Process Design

Designing the extraction process and the condition is very important for the extraction of phenolic compounds from the solid phase to obtain the highest yield. This is necessary due to the sensitivity of phenolic compounds to factors such as heat, oxidation, and degradation during the extraction process. The main objective of this study is to optimize the extraction process for phenolic compounds from raw materials and determine the ideal extraction conditions that result in a high yield of these compounds. White turmeric was selected as the investigated raw material and ultrasound-assisted ethanolic extraction (UAE) was used as the extraction technique. Response Surface Methodology (RSM) with Box-Behnken was applied for designing the UAE process in terms of three parameters (solid-to-solvent ratio, extraction time, and extraction temperature). Within the designed levels of solid-to-solvent ratio (0.2 to 1.0 g/10 ml), extraction time (20 to 180 min), and extraction temperature (20 to 70 °C), it is noticed that the extraction yield of phenolic compounds varied depending on the levels of those extraction process conditions. The extraction yield of phenolic compounds was low at the solid-to-solvent ratio of 1.0 g/10ml, the temperature at 20 °C, and extraction time at 20 min and it was high at the solid-to-solvent ratio (1.0 to 1.8 g/10ml), temperature (45 to 70 °C), and extraction time (100 to 180 min). However, based on the statical data and confirmation experiment, it was possible to obtain the highest yield of phenolic compounds of 9.0 mg GAE/g DW, extracting from white turmeric at UAE condition of 1.1 g/10 ml for solid-to-solvent ratio, 100 min for extraction time, and 45 °C for temperature. The result of this study may be helpful for the intensification of ultrasound-assisted solvent extraction for phenolic compounds from agricultural products.


Introduction
Basically, to achieve the highest possible yield of total phenolic compounds from a solid sample, proper experimental design plays a crucial role in reducing the time and cost associated with traditional trial-and-error methods [1,2].Furthermore, it is important to optimize the operating factors, including extraction time, extraction temperature, solid-to-solvent ratio, and other relevant variables, by employing appropriate statistical methods such as the Box-Behnken Design (BBD) with Response Surface Methodology.The BBD serves as a statistical technique utilized to optimize responses that are affected by various independent variables within a specified range [3,4].
Moreover, Curcuma longa, generally known as turmeric, has been considered a medicinal plant and a lot of studies have been published about its nutritional applications and biological activities [5].Otherwise, white turmeric (C.zedoaria) rhizomes are usually used as traditional medicine in Asia countries to treat some diseases such as stomach diseases, toothache, and blood stagnation.Some previous reports claimed that C. zedoaria is rich in source essential oil, terpenoids, and curcuminoids [6].However, according to the available information, the optimization of total phenolic compounds for modelling and maximizing from white turmeric is still limited.
Therefore, this study aimed to optimize the extraction process of total phenolic compounds from Cambodian white turmeric rhizomes.This involved investigating the effects of operating factors such as extraction time, extraction temperature, and solid-to-solvent ratio using the Box-Behnken Design (BBD) methodology to achieve maximum yield.

Determination of moisture content and sample preparation
The white turmeric, specifically the Khmer cultivar, was obtained from Phnom Kulen in Siem Reap province, located in north-western Cambodia.Before extraction, the sample underwent a cleaning process to eliminate unwanted substances like dust and leaves.The moisture content (10.18±0.55%;w.b) of the sample was determined using the conventional oven method, with a drying time of 16 hours at a temperature of 50 ℃.Once dried, the white turmeric sample was ground, sieved, and subsequently used for the extraction of phenolic compounds.

Ultrasound-assisted extraction of phenolic compounds
The extraction of total phenolic compounds (TPC) from white turmeric was performed using ultrasound-assisted extraction (UAE) under the conditions specified in Table 1.After the extraction, TPC was quantified by Folin-ciocalteu assay with some modification from Sepahpour et al. [7].TPC result was expressed as a milligram gallic acid equivalent per gram of dried weight (GAE/g DW).The utilization of the Box-Behnken design (BBD) in conjunction with Response Surface Methodology (RSM) was employed to identify optimal extraction conditions for maximizing the yield of total phenolic compounds (TPC).The extraction time, extraction temperature, and solidto-solvent ratio were investigated at three different levels to determine the conditions that would result in the highest extraction yield of TPC [8].The mathematical equation defining the BBD is given in (1).
Y is the response; β0, βi, βii, and βij are the regression coefficient of the intercepts, linear, quadratic, and interaction terms, respectively; and Xi and Xj are the independent variables.

Statistical analysis of experimental data
The experimental data were determined by applying the repeated measures ANOVA and response regression technique at a 5 % significant level using JMP software version 14.0.

Optimization of phenolic compounds from white turmeric
This study focused on extracting total phenolic compounds (TPC) from white turmeric rhizomes by manipulating three operational factors: extraction time, extraction temperature, and solid-tosolvent ratio.The concentration of TPC in the extracts obtained from all experimental conditions is presented in Table 2. Within the designed range of solid-to-solvent ratio, extraction time, and temperature, the TPC in white turmeric extracts ranged from 4.75±0.01 to 11.13±1.86mg GAE/g DW.Furthermore, the independent variables exhibited their impact on the extraction yield of total phenolic compounds (TPC) from white turmeric.The statistical value of the student's t-test and the corresponding significance level are summarized in Table 3.
where X1 represents for solid-to-solvent ratio, X2 represents temperature, and X3 represents extraction time.
According to equation (2), the insignificant factors have been removed from the model, keeping only the significant independent variables.Moreover, the value set of extraction temperature (20 to 70 ℃) had no significant response, meaning that a lower temperature can be applied to extract phenolic compounds from white turmeric to save more energy consumption and avoid the degradation of bioactive compounds which are sensitive to high temperature.

Response surface optimization
Figure 1.shows the desirability plot of the maximum value of TPC extracted from white turmeric rhizomes.The surface plot revealed that the maximum yield of TPC was 9 mg GAE/g DW corresponding to 1.1 g/10 ml for solid-to-solvent ratio, 100 min for extraction time, and 45℃ for extraction temperature.The model generated by RMS with Box-Behnken Design will be significant for predicting the optimum extraction conditions to obtain the highest yield of phenolic compounds from Cambodian white turmeric.
all factors, except factors β2, β12, β13, β23, β11, and β22 can be deemed significant as their p-values are less than 0.05.These aforementioned factors can be regarded as insignificant based on their p-values.The equation for the quadratic model utilized to determine total phenolic compounds is expressed as follows: Y= 9.24 -2.15X1 + 0.83X3 -1.30X3 2 4. ConclusionThe investigated variables, namely extraction time (ranging from 20 to 180 minutes), extraction temperature (ranging from 20 to 70 ℃), and solid-to-solvent ratio (ranging from 0.2/10 to 1.8/10 g/ml), proved successful in maximizing the overall yield of phenolic compounds extracted from Cambodia white turmeric.The highest TPC value obtained was 9.0 mg GAE/g DW, achieved with a solid-to-solvent ratio of 1.1 mg/10 ml, an extraction temperature of 45 ℃, and an extraction time of 100 minutes.Additionally, the results indicated that the extraction yield was not significantly affected by temperature within the specified range.Future studies should consider white turmeric for extracting essential oils and curcuminoids for potential applications in pharmaceuticals and cosmetics.

Figure 1 .
Figure 1.Surface plots of the effect of extraction conditions on TPC

Table 1 .
Experimental design for UAE using RSM with Box-Behnken design 2.4.Response Surface Methodology with Box-Behnken Design

Table 2 .
Extraction yields of TPC from white turmeric extracts.

Table 3 .
Estimated regression coefficients for the model of TPC yields.