Physicochemical properties of dragon fruit peel pectin and citrus peel pectin: a comparison

Dragon fruit is a tropical fruit belongs to Cactae family of genus Hylocereus. Popular as buah naga in Malay, dragon fruit is not only sought after for its wonderful taste, but this fruit is also nutritionally and medicinally attractive for consumers. The dragon fruit flesh is usually eaten raw or it is processed into juices and the remaining peel will be discarded. To minimize the waste created from dragon fruit consumption, an idea of transforming the peel of dragon fruit into edible pectin has been proposed. Pectin can be found in food products as a gelling agent and it is used to maintain the quality of foods. Recent market demand for pectin is increasing annually and to meet the supply-demand, researchers are working towards obtaining pectin from available non-conventional sources. In this study, pectin was extracted from dragon fruit peel by hot acid extraction at pH 3.5 and 75 mins of extraction. Distilled water was used as a solvent and the acidic environment was achieved with dilute HCl. Degree of esterification (DE) analysis was performed by FTIR. The extracted pectin was categorized as high methoxyl pectin based on DE (52%). The endothermic properties of dragon fruit pectin analyzed by DSC indicating the stability of extracted pectin compared to citrus peel pectin. Pectin extracted was characterized in terms of moisture (14.03±1.925), ash content (8.73±1.218), as well as water and oilholding capacity (4.08±0.0.81) (2.18±0.096), swelling capacity (5.94±1.271) and texture profile in terms of gumminess (33.66±6.05), cohesiveness (1.06±0.08) and hardness (31.63±3.81). All characteristics were compared with citrus peel pectin.


Introduction
Red dragon fruit (Hylocereus polyrhizus) is an exotic, sweet and nutritionally wise fruit that is widely cultivated around Southeast Asia. In Malaysia, the annual production of dragon fruit is about 10, 961 tons (Hoe, 2017). Red dragon fruit is highly sought after because of their high nutritional content. Dragon fruit contains up to 9 g per 100 g of vitamin C which is the highest in the fruit among other vitamins (Perween et al., 2018). Protein, carbohydrate, fat, B1, B2, and B3 vitamins, thiamine, niacin, flavonoids, betacyanin are among any other nutrients that can be found in dragon fruit (Hashim, 2018). The dragon fruit is eaten as fresh fruit, peeling the skin away. They are also refined into juice, beverages and syrups. The peel, considered waste from the processing of dragon fruit, is about 22% of the whole fruit . Such waste is also abundant, according to literatures, with beneficial compounds like flavonoids and pectin (Hashim, 2018) (Ismail et al., 2012;Muhammad et al., 2014). Dragon fruit peel consists mainly of pectin (10.79%), betacyanin pigments (150.46/ 100g db) and a total dietary fibre of up to 69.3% (Chia and Chong, 2015). Commercial pectin has been most prevalently derived from apple pomace and citrus peel and the use of dragon fruit waste to produce value-added ingredients such as pectin has a huge potential.
The word 'pectin' came from Latin and was first developed by Henri Braconnot in 1825. Pectin is a standardized heteropolysaccharide composed of α-1, 4 bound D-galacturonic acid which is commonly found in the primary cell walls of terrestrial plants. The use of pectin as a gelling agent was developed in the food industry as early as in the last two centuries. (Sriamornsak, 2003). Pectin has been used in food products such as jellies, jams, confectionery products, frozen foods, baking products and drinks as a gelling agent, thickener, emulsion and stabiliser. Pectin demand is increasing from time to time and pectin from various sources have been investigated to cater the growing needs such as banana, mango, pumpkin, blueberry, sugar beet, grapefruit, and watermelon (Khan et al., 2014;Castillo-Israel et al., 2015;Girma and Worku, 2016;Huang et al., 2017;Petkowicz et al., 2017Koh et al., 2018Dona, 2019). The structure and characteristics of pectin can vary depending on its source and recovery techniques. Pectin attribute has influenced how it is used in the industry and the degree of esterification (DE) is an important feature in exploring the efficacy of pectin gelling agent (Zaid et al., 2019).
Earlier studies have reported findings of pectin from the peel of dragon fruit and citrus. Pectins are complex carbohydrate molecules that are used in numerous food applications as a gelling agent, thickener, stabiliser, and emulsifier. Dragon fruit (Hylocereus polyrhizus) is one of the tropical fruits that belong to the cactus family, Cactaceae. Since the peels of dragon fruit are often discarded as waste, it would be an advantage to convert it into a value-added product such as pectin. The objective of this study was to investigate the extraction of pectin from dragon fruit peels under different extraction time using hot water extraction method. The dragon fruit peels were extracted using distilled water at 80°C with different extraction time of 20, 40, 60 and 80 mins. The extracted pectin was characterised by its yield, moisture and ash content, degree of esterification and antioxidant activity. Determination of moisture and ash content was conducted using AOAC standard method. The determination of the degree of esterification of pectin was performed using Fourier Transform Infrared Spectroscopy (FTIR). DPPH assay was used to determine the antioxidant activity of the pectin extract. Based on the result, the yield of pectin decreases (20.34 to 16.20%) with the increase of extraction time, moisture contents were between 4 to 6% while ash contents were between 7 to 10%. Pectin from dragon fruit peels was determined as low methoxyl pectin and has a high percentage of antioxidant activity with a low value of inhibition concentration (IC50) (0.0063 to 0.0080 mg/ mL). 60 mins extraction sample exhibits the highest antioxidant activity (81.91% at 40 µg/mL), followed by 80 mins extraction (81.68% at 40 µg/mL), 40 mins extraction (81.38% at 40 µg/mL) and 20 min extraction (81.31% at 40 µg/mL) (Izalin et al., 2016;Rubaiyi et al., 2016;Abang Zaidel et al., 2017;Ahmed and Sikder, 2019). These fruits may vary in functionality and nutritional properties, depending on their variety. This study builds on the existing knowledge fruit pomaces' nutritional properties. This stresses the need to consider both the nutritional and functional characteristics of a fruit product before being integrated into a food product.

Sample preparation
Dragon fruits were obtained from Malaysian Agricultural Research and Development Institute (MARDI)'s supplier. The fruits were washed and the peel were cut into smaller sizes (≈2cm 2 ) and stored in a sealed polyethylene bag in a freezer for 48 hrs. The samples were then freeze-dried for 72 hrs using a freeze dryer (Martin Crist, Germany; Alpha 1-4 LD Plus with RZ 2.5 vacuum pump) and ground into fine powder and kept at 4 o C prior to analysis. Citrus peel pectin was obtained from KES Saintifik (Malaysia) and all reagents were from Sigma-Aldrich (USA).

Pectin Extraction
A total of 8.0 g of powdered sample was rehydrated with 250 mL of distilled water. The pH was adjusted to 3.5 using 1M hydrochloric acid with the aid of a pH meter equipped with a temperature compensation probe (METTLER TOLEDO). The extraction process was carried out at 70 o C using stirring hot-plate (500rpm) for 75 mins. The mixture was then centrifuged for 5 mins at 1000 rpm. Thereafter, a double volume of 95% ethanol was mixed to aid the overnight precipitation. The next day, pectin was isolated using a double layer of cheesecloth and washed three times with 70% alcohol. The final washing of extracted pectin was done using undiluted alcohol to remove the remaining impurities such as monosaccharide and disaccharide (Tang et al., 2011). Finally, pectin collected was freeze-dried to remove all water for 72 hrs. For further examination, the samples were cooled, weighed, ground and stored in small plastic bags. The yield of pectin was calculated as the ratio of the weight of dried pectin to the dried extraction process powder.

Degree of Esterification (DE) of pectin (FTIR analysis)
FTIR analysis was carried out using infrared spectrophotometer (Nicolet TM iS TM 10 FTIR Spectrometer-Thermo Fisher Scientific). The spectra were recorded in the absorbance range from 4000 to 400 cm -1 . Peak area relation of the non-esterified carboxyl groups (COO-) (1614.98 cm -1 ) and esterified carboxyl groups (C=O) (1737.77 cm -1 ) were used to determine the DE. DE was calculated by referring to this calculation: Where APECG is the peak area of esterified carboxyl FULL PAPER eISSN: 2550-2166 © 2020 The Authors. Published by Rynnye Lyan Resources groups, and APNECG is the peak area of non-esterified carboxyl groups.

DSC analysis of extracted pectin
Differential scanning calorimetry (DSC) was used to measure the change in heat flow by a sample after heating quantitative and qualitatively during the endothermic process. DSC was conducted based on the method described by Wang et al. (2014). Samples (5 g) were heated in a closed aluminum pan at 10 o C / min from -30 o C to 255 o C. The system was purged with nitrogen gas at a flow rate of 50 mL/min.

Moisture and ash content of extracted pectin
Moisture and ash content analyses of extracted pectin were performed according to the Association of Official Agricultural Chemists AOAC methods as explained by Jauhari et al. (2013). Moisture content was obtained by drying the sample in the oven at 105 o C for 4 hrs. For ash content, the sample was burnt in furnace muffle at 550 o C for 2 hrs. All contents were determined in percentage form and calculated based on the followings:

Water holding capacity (WHC), oil holding capacity (OHC) and swelling capacity (SC) of extracted pectin
WHC, OHC and SC of extracted pectin were measured using the method by Izalin et al. (2016). 100 mg of sample was weighed and added into 50 mL centrifuge tube containing 25 mL of distilled water. The mixture was incubated overnight at 4 o C, centrifuged for 20 mins at 13 o C (600 x g) and left at room temperature for 2 hrs. The dried sample was then weighed to obtain the weight of dry residue. WHC calculation is as below: A total of 100 mg of sample was added into a centrifuge tube containing 10 mL of corn oil to determine the oil-holding capacity (OHC). The content was stirred and centrifuged at 1500 x g for 30 mins. The supernatant was weighed and OHC was expressed as below: For swelling capacity, sample (100 mg) was weighed in a cone-shaped plastic tube. Distilled water (5 mL) was added into the tube and the mixture was left overnight.
The swelling capacity of the extracted sample was determined using the following equation:

Texture profile analysis of extracted pectin 2.7.1 Gel preparation
Pectin gel was prepared according to Rascón-Chu et al. (2009) by adding 3% pectin concentration and sucrose in aqueous solution. The environmental pH was adjusted to 2.7 with a few drops of 1M hydrochloric acid (HCl) and the solution was boiled for 30 mins. Gel formed was kept overnight at 4 o C. The gel was measured using TA-XT Express Texture Analyzer (Stable Micro Systems, England) using 0.5" cylinder probe (P/ 0.5) at 1.0 mm/s up to 15 mm depth. The gel was analyzed for hardness, gumminess and cohesiveness.

Statistical analysis
All collected data were expressed as mean ± standard deviation (SD). Independent t-test was conducted using XLSTAT-Pro (2014) Statistical Software (Addinsoft, France) at 95% confidence level is p< 0.05.

Degree of Esterification (DE) of extracted pectin
FTIR spectroscopy was used to determine the DE of extracted pectin. FTIR includes both direct and nondestructive processes and the procedure requires only a small sample quantity to be completed within a shorter time frame. The DE was estimated at 52%, indicating that the pectin of dragon fruit peel is high methoxyl pectin (HM pectin) (Marić et al., 2018). The previous study reported that the ability to form gel is increasing in HM pectin (Coelho et al., 2017). High methoxyl (HM) pectin is a pectin group with DE value more than 50% and low methoxyl (LM) pectin has a DE value of less than 50% (Mamet et al., 2017). The present finding seems to be consistent but slightly lower than reports of dragon fruit peel pectin DE range of 57 -63% (Tang et al., 2011;Adzahan et al., 2014;Tongkham et al., 2017). Possible causes for variation in DE values are the differences in growing environments, plant varieties and pectin extracting conditions (Tongkham et al., 2017).
In the region between 4000-400 cm -1 , the FTIR spectra identified the major chemical groups in pectin such as OH, COOH, and COOCH 3 and provided structural information on pectin molecule. Figure 1 display the structural information of pectin. The FTIR spectra of extracted pectin from the peel of dragon fruit are presented in Figure 2(a) and are compared with the spectra for citrus peel pectin Figure 2(b). It could be seen FULL PAPER that the peak at 3296.92 cm -1 indicates OH groups and the stretching band around 3400-3200 cm -1 is due to inter -and intra-molecular hydrogen bonds of the galacturonic acid polymer (Coelho et al., 2017;Hashim, 2018). The absorption bound at 2935.67 cm -1 was due to C-H stretching of CH 2 groups. The major functional group in pectin is located at the region in between 2000 and 1000 cm -1 of the FTIR spectra. Absorption bands appeared at peak 1737 cm -1 was due to stretching vibrations of nonionized C=O groups and the appearance of peak 1614.98 cm -1 is the stretching vibration of carboxyl ions (COO-). These data indicate that the extracted substance is pectin since the spectral region between 1800 and 1500 relates to carboxylic acids and carboxylic esters, which are the key functional groups responsible for the characteristics of pectin resembles the spectral region of citrus peel pectin (Coelho et al., 2017). Also, the fingerprint region for carbohydrate, which is located at wavenumber 800-1300 cm -1 can be used to identify the major chemical group of polysaccharide (Abang Zaidel et al., 2017).

DSC Analysis of dragon fruit peel pectin
DSC analysis was used to measure the enthalpy changes in the sample as a result of physical and chemical changes by a function of temperature and time. The temperature difference between -30 o C and 255 o C was used to study the thermodynamic properties of dragon fruit peel pectin. Figure 3a shows a single endothermic peak of dragon fruit peel pectin and Figure   3b shows a distinct endothermic peak of citrus peel pectin in DSC thermogram for comparison. The single endothermic peak at 120°C for dragon fruit peel pectin and at 150°C for citrus peel pectin was observed in DSC curves. Melting temperature (T m ) was determined from the maximum of the endothermic peak (Mhd Sarbon et al., 2013). T m (122.01 o C) and melting enthalpy ΔH m (385.40 mJ) of extracted pectin were higher than those in citrus peel pectin (107.11 o C and 77.81 mJ, respectively). The values indicated that extracted pectin had a better capacity to sustain water as it requires more energy to remove water completely as reflected by higher melting temperature and enthalpy (Wang et al., 2014). Higher water content and different conformation of dragon fruit peel pectin also shifted the peak of endothermic toward lower temperature (Wang et al., 2016). Table 1 shows mean and standard deviation (SD) of moisture and ash content in percentage. Prior studies have reported the importance of moisture and ash content to determine the quality and purity of the pectin (Castillo -Israel et al., 2015). The low moisture content of pectin (<10%) is necessary for storage and prevention of bacterial growth that can affect its quality (Girma and Worku, 2016). The moisture content of dragon fruit peel pectin is 14.03% which is significantly higher than that of citrus peel pectin. The literature stated lower moisture content of dragon fruit peel pectin (4.37-13.13%) (Ismail et al., 2012;Nurul and Asmah, 2014;Abang Zaidel et al., 2017). A possible explanation for this might be the soggy environment used to keep the sample or human error while conducting the test. Hence, the data could be improved in terms of repeating the experiments and monitoring the sample storage. Several studies using different sources of pectin also resulted in a different amount of moisture content (Castillo-Israel et al., 2015; FULL PAPER Figure Girma and Worku, 2016;Jaswir et al., 2016;Mohamed, 2016). Another possible explanation for this difference is due to the difference in the hygroscopic nature of pectin as different pectin has different degrees of esterification (DE) (Sharma et al., 2014). The moisture content of pectin should be below 10%. Higher reading will cause the humid environment and suitable for bacterial growth (Mohamadzadeh et al., 2010;Ismail et al., 2012;Mohamed, 2016) and reducing the shelf life of the food products (Jauhari et al., 2013).

Moisture and ash content of extracted pectin
Ash content is used to determine the purity of pectin. For good quality of pectin, the ash content must be below than 10% (Girma and Worku, 2016). In this study, the extracted pectin contains 8.73% of ash content which is not significantly different than the ash content found in commercial pectin (9.08%). The present finding seems to be consistent with other research which found the ash content below 10% (Bhat and Singh, 2014;RamLi et al., 2014). The previous study also suggested that the value of ash in dragon fruit peel pectin can form better gels than that from citrus peel pectin. Table 2 compares the result of WHC, OHC and SC obtained from extracted dragon fruit peel pectin and citrus peel pectin. The results were expressed in mean and standard deviations (S.D). The ability of a material to retain water and oil following the centrifugation is dependent on its WHC and OHC. Specifically, WHC signifies the ability of soluble dietary fiber to absorb and retain water to form a viscous solution. In addition to that, high WHC is commonly associated with greater pectin content Rubio-Senent et al., 2015). In this study, the WHC of dragon fruit peel pectin (4.08) is slight lower than the citrus peel pectin (4.61). This current data also shows lower reading than previous study of WHC of dragon fruit peel (5.5). This means that the ability of dragon fruit peel pectin to form gel is poorer than citrus pectin due to lower pectin content (Izalin et al., 2016). However, the value obtained was sufficient to prevent syneresis and to modify texture of foods (Dona, 2019). WHC in pectin is important to maintain the texture of high-fat foods (Mingyue, 2017), improve the sensory attributes and reduce calories of foods (Bouaziz, Rassaoui, and Besbes, 2014). Another significant functional property of some ingredients used in the prepared food is OHC. Ingredients with a high OHC  promote stabilization of high fat food products and emulsion, assisting the dispersion of two immiscible liquids (Rubio-Senent et al., 2015). In this study, OHC of dragon fruit peel (2.18) is higher compared to citrus peel pectin (2.083), signified higher pectin content (Rubio-Senent et al., 2015). The possible explanation for this dissimilarity is due to the difference in their chemical and physical structure and polysaccharides (Chia and Chong, 2015). Pectin with higher OHC is widely used as an emulsion in beverages (Dona, 2019) and in soups, baked goods and as an extender in meat (Izalin et al., 2016).

WHC, OHC and SC of extracted pectin
In this study, the mean score for SC of dragon fruit peel pectin is slightly lower than that of citrus peel pectin. This result may be explained by the fact that different sources of pectin will have different swelling properties, solution and also gel formation (Sriamornsak, 2011). It is important for pectin to have high SC as it will enhance hydrodynamic volume especially when dissolving in water. In the frozen food industry, pectin is required to maintain the juiciness of cooked fish and meat products and to lessen the drip loss with the help of good SC (Gomez-Guillen et al., 2011). In biological application, pectin has been used as a wound care material for having a good swelling capacity .

Texture profile analysis of dragon fruit peel pectin
The texture of gel processed from dragon fruit peel pectin was analyzed in terms of hardness, cohesiveness and gumminess and compared with commercial pectin gel. Figure 4(a) shows the image of gel made up from extracted pectin and figure 4(b) is the gel from citrus peel pectin. Sucrose was added into the pectin solution during gel processing. This is to dehydrate the pectin by competing for water and reduce its ability to stay in a dispersed state. Sugar also reinforce the junction zones between pectin molecules and strengthen the hydrophobic interactions between methyl groups (Niezgoda, 2015). Formation of gel occurred due to the bonding of hydrogen in between of free carboxyl groups on the pectin molecules and the hydroxyl groups of neighboring molecules (Pilgrim et al., 1991). In this study, the value of hardness and gumminess of dragon fruit peel pectin gel is higher than that of commercial pectin gel (Table 3). For cohesiveness, the mean value is similar in both gels and the result is harmony with previous researchers (Izalin et al., 2016). Further statistical analysis revealed that there were no significant difference between two different gels for all attributes. This result is in good agreement with the previous report which stated that the gelling quality of extracted pectin was comparable to that of commercial pectin (Nazaruddin et al., 2011). Therefore, it can be concluded that gel produced from dragon fruit peel has an excellent texture comparable to commercial pectin gel.

Conclusion
Dragon fruit peel is usually discarded after the consumption of the fruit's flesh. The copious of waste that is produced from this operation will contribute to environmental pollution. Since the peel provides many advantages, researchers found a way to turn it from waste to wealth, by extracting pectin from the peel. Dragon fruit peel is one of the potential sources to supply the market demand for pectin. Results obtained indicate that dragon fruit peel pectin is of the high methoxyl pectin type and this is important for further applications in the food industry. Melting properties of extracted pectin show no significant difference when compared with commercial pectin. The extracted sample is pure and original as supported by the ash analysis. However, further analysis should be given attention especially on the moisture content to ensure that the quality of pectin is established. Overall, other physicochemical analysis and texture profile analysis concluded that the extracted pectin have good potential to become another source of pectin. The quality of pectin is as good as pectin from citrus peel pectin.