White and orange fleshed-sweet potato starches modified by autoclave

The objective of this research, using a Composite Central Rotational Design, was to evaluate whether the moisture and exposure time to a temperature of 121 °C and pressure of 1.1 kgf. cm in an autoclave would be capable of modifying the morphological, thermal, functional and pasting properties of orange (OSP) and white (WSP)-fleshed sweet potato starches. The modification increased: the WAI at 60oC of WSP starch and of WSI at 60oC of OSP starch, the setback of WSP starch, the final viscosity of OSP starch and enthalpy for both, besides reduced the breakdown of OSP starch. The micrographs of modified starches showed cracks and on the surface of the granules, and pregelatinization of OSP starch, confirming that the moisture and exposure time affected their morphological, thermal, functional and pasting properties. Therefore, due the promising use the authors suggest future researches with WSP and OSP autoclaved starches to test technological applications.


Introduction
The sweet potato (Ipomoea batatas L.) starch is being widely used in the food industry (Zuo, Hébraud, Hemar, & Ashokkumar, 2012). The starches extracted from the colored flesh of the various sweet potato cultivars have similar physicochemical properties but different pasting and thermal properties (Guo, Liu, Lian, Li, & Wu, 2014;Lee & Lee, 2017).
Starch granules exhibit distinctive morphologies and technological properties that are affected by factors such as: color, root genotype, environmental conditions and the soil type where they were grown (Narpinder, Amritpal, Khetan, & Rajarathnam, 2013). Colored sweet potato starches extracted from sweet potatoes of different flesh colors, may have heterogeneous granules sizes and shapes and different crystallinity patterns. The viscoamylographic profile of native white and orange-fleshed sweet potato starches in previous studies showed good thermal stability, being indicated for the potential use in foods processed at high temperatures, such as sauces and snacks (Paixão & Silva et al., 2020). Deka and Sit (2016) stated that the use of native starches in food processing has limitations such as the formation of a weak, cohesive, and elastic paste when heated; and the formation of an unwanted gel when refrigerated or stored. Due to these limitations, the functional properties of these starches can be improved by modifications using physical, chemical, or enzymatic methods.
The physical modification of starch for application in food is one of the most viable ways, since it is associated with environmental preservation and sustainability and does not produce residues, to the contrary of chemical modifications. In addition, changes in the nutritional properties of starches modified by chemical means, which could affect human health, have been poorly studied (Zhu, 2015). Thus, the physical modifications of starch are more attractive since they are considered to be natural and safe for the consumer (Zuo et al., 2012).
Thus, in order to enhance the application of orange and white-fleshed sweet potato starch, the objective of this research was to evaluate whether the moisture and time of exposure to a temperature of 121 °C with a pressure of 1.1 kgf. cm 2 in an autoclave would be capable of modifying the morphological, thermal, functional and pasting properties of white (WSP) and orange (OSP)-fleshed sweet potato starches.

Methodology
The methodology used in this article was experimental, explanatory and quantitative, according to the classification described by Pereira, Shitsuka, Parreira, and Shitsuka (2018).

Material and starch extraction
Samples consisting of 20 Kg each of white (common variety) and orange (BRS Amélia variety) fleshed sweet potatoes (Ipomoea batatas L.) were obtained from a farm located at a latitude of 16° 58′ 11.46″, longitude of 49° 11′ 12.206″ and altitude of 830 m. The sweet potatoes were transported in polyethylene bags to the laboratory and immediately processed.
The starch was extracted as described by Bento et al. (2019) with modifications: the roots were washed, peeled, cut into 4 cm slices, immersed in sodium metabisulfite solution (5 g L -1 of water) for 15 min., and then ground with water in an industrial blender for 2 min. with a 2: 1 ratio (distilled water: sweet potato). The mass obtained was filtered through 35 and 400 mesh sieves, decanted for 4 h, and drained. The starch was resuspended in water, decanted again for 3 h, drained, dried in a forced air oven at 40 ° C for 48 h, reduced to a powder using a pestle and mortar, and finally sieved through a 60-mesh sieve.

Autoclave modification
Autoclave modification of OSP and WSP starches was performed according to a methodology proposed by Abraham (1993) with adaptations. 30 g of the starches were placed in autoclavable polyethylene bottles and adjusted to the desired moisture by adding distilled water. After kept 24 hours the moisture content of samples was verified by infrared balance 30; 37.09 and 40%). A vertical autoclave (Phoenix Scientific Equipment mod-AV50, Araraquara, Brazil), with a fixed temperature and pressure (121ºC and 1.1 kgf cm 2 , respectively) was used to carry out the changes in the OSP and WSP starches. After autoclaving, the modified starches were dried in a forced air oven at 40 ºC for 24 h. The lumps formed were reduced to powder with the aid of a mini-grinder and then sieved through a 60-mesh sieve.

Gel transparency or transmittance
The transparency of the gel was determined according to Zheng et al. (2013), whereby 0.1 g portions of the samples were added to 10 ml of water (1% suspension) in a test tube and heated in a water bath at 95ºC for 30 min. with agitation. They were then cooled for 15 min. to room temperature, and the transmittance determined in a spectrophotometer (Bel 107 Photonic, SP2000UV, Piracicaba, Brazil) at 620 nm, using water as the blank.

Water solubility and swelling power index
The water solubility and absorption indexes (WSI and WAI) were determined as described by Bento et al. (2019) at temperatures of 60 ºC, 70 ºC and 80 ºC. The result of WAI was expressed in g gel (g dry matter) -1 , and the WSI expressed in %.

Differential scanning calorimetry (DSC)
The thermal properties were determined in a Differential Scanning Calorimeter -DSC (TA Instruments, Q20, Newcastle, UK), as described by Bento et al. (2019). The gelatinization enthalpy (ΔH) and the onset (To), the peak (Tp) and the end of gelatinization (Tc) temperatures were obtained from the thermogram produced by the calorimeter.

Pasting properties
The pasting properties were evaluated in a Rapid Visco Analyzer (RVA) (Perten,131 RVA 4500,Huddinge,Sweden) according to the criterion adopted by Bento et al. (2020). The pasting properties evaluated were: peak viscosity, final viscosity, breakdown viscosity and the tendency to retrograde (setback), all expressed in cP and the pasting temperature in ºC.

Gel hardness
After completing the analysis of the pasting properties, the samples were kept under refrigeration overnight, and the gel strength at 25ºC subsequently determined in a texturometer (TA HD Plus Stable Micro Systems, Surrey, England), using a 20 mm cylindrical probe, 8 mm penetration depth, 10 g.f contact force and test, pre-test and post-test speeds of 0.5, 1.0 and 10.0 mm s -1 , respectively (Wani, Sogi, Wani, Gill, & Shivhare, 2010).

Scanning electron microscopy (SEM)
The granule morphology was analyzed in a scanning electron microscope (Jeol, JSM -6610, Tokyo, Japan) as described by Bento et al. (2019). The photographs were produced in triplicate, with magnifications of 1500x. The particle dimensions were determined using the IMAGE software (Schneider, Rasband, & Eliceiri, 2012).

Statistical analysis
All data were obtained in triplicate and evaluated by response surface methodology and multiple variance analysis, and models were adjusted and visualized by means of response surface graphs (Statsoft, Statistic 10.0, Tulsa, USA). Models were also validated in three-replicate assays using the central point of the first design. Table 1 shows the significant adjusted models (p <0.05) with non-significant lack of fit (p> 0.05) and the determination coefficients above 70% of WAI (at 60 and 70 ° C) and WSI (at 60, 70 and 80 ° C); breakdown, final viscosity and setback; gelatinization enthalpy obtained from the DSC; and the hardness and transparency of the gels, from OSP and WSP autoclaved starches. In the validation test, it was found that all the models were predictive (with an accuracy level greater than 95%) (error<5%) with the exception of the gelatinization enthalpy of OSP starch autoclaved (with compatibility of 90.47%).

Water solubility index (WSI) and water absorption index (WAI)
The adjusted significant multiple regression models (p <0.05) of theses parameters were: WAI at 70 ° C and WSI at 60 and 80 ° C (for OSP autoclaved starch), WAI at 60 and 70ºC and WSI at 70ºC (for WSP autoclaved starch) (Table 1).
For the OSP autoclaved starch WAI at 70 ° C, only the linear and quadratic effects of the moisture content were significant and positive (Table 1), and the highest values (12.0 g g -1 ) were found for moisture contents above 37% ( Figure 1A).   (Table 1). Higher WAI values for the WSP autoclaved starch at 60 ºC (14.8 g g -1 ) and at 70 ºC (15.5 g g -1 ) were observed with moisture contents above 40% ( Figures 1B and 1C).
The OSP autoclaved starch WSI at 60 ° C fluctuated significantly with the positive linear effects of reaction time and moisture content, the positive quadratic effect of moisture content and the positive interaction effect of the independent variables ( Table 1). The maximum water solubility (9.0%) was found after exposure times greater than 57 min. and moisture contents above 43% ( Figure 1D). The OSP autoclaved starch WSI at 80 ºC was affected by the positive linear and negative quadratic effects of the moisture content and was not influenced by time (Table 1). The highest water solubilities (5.0%) were determined at moisture contents above 30% ( Figure 1E). Also, for the WSP autoclaved starch WSI at 70 ºC, the linear and quadratic effects of the moisture content were positive (Table 1), reaching values greater than 6.5% with moisture contents above 40% ( Figure 1F).
Previous studies with native starches obtained for WAI of WSP e WSI of OSP at 60 ºC were lower (6.7 g g -1 and 1.0 %, respectively) (Paixão e Silva et al., 2020) than the indexes found in this research for the same autoclaved starches (14.8 g g - The autoclave treatment associated with a test temperature caused greater changes in the starch granules, since it favored water entering the structure (Jyothi et al., 2010). However, the changes are not only caused by the treatment, but also by the analysis temperature , as can be seen by the reduction in solubility at 80 °C for the OSP autoclaved starch ( Figures 2D and 2E). Also, the pasting temperature of OSP native starch observed previously (73.8 ± 0.5 ºC) (Paixão e Silva et al., 2020) was lower than analysis temperature.
The increase in solubility and swelling power is due to the breakdown of hydrogen bonds between the starch granule molecules, which provides greater magnitude of the interaction between the amorphous and crystalline domains of the starch (Bello-Pérez, Sánchez-Rivera, Núñez-Santiago, Rodríguez-Ambriz, & Román-Gutierrez, 2010). Thus, the increase in temperature weakens the intramolecular bonds in the starch, favoring the amylose leaching process, thus increasing solubility.
Weaker forces, such as those of Vander Walls, relax and cause the granules to absorb water and swell, and, as a result, the lower molecular weight amylose dissolves and is released from the granule into the surrounding medium, thus increasing the starch solubility . The low hydration capacity of the amylopectin-rich fraction is due to the water penetration restrictions, thus limiting the swelling and solubility capacity of these portions (Jamir & Kottapalli, 2017). Thus, the solubility and swelling power of the autoclaved starch granules are probably affected by the proportion of amylose and amylopectin in each starch, as well as by the arrangement between the amylose and amylopectin.

Thermal properties
The gelatinization temperatures determined from the DSC thermograms (Tonset, Tpeak and Tfinal) of the autoclaved starches were not significantly modified (Table 1), although the gelatinization enthalpy could be modelled (p <0.05).
The OSP autoclaved starch gelatinization enthalpy was reduced with increasing moisture content, while the exposure time and the quadratic effect of the moisture content increased the energy required for starch gelatinization (Table 1). The energy required for gelatinization was higher (22.0 J. g -1 ) in the time below 13 and above 62 min. and with moisture contents below 19% (Figure 2A). For WSP autoclaved starch, only an increase in moisture content reduced the energy required to gelatinize the granules (Table 1), and the energy required for gelatinization was higher (18.0 J. g -1 ) with moisture contents below 19% ( Figure 2B). These optimized values for the autoclaved starches are higher than the preliminary studies with natives WSP and OSP starches that found enthalpy of gelatinization of 13.5 and 11.24 J. g -1 , respectively (Paixão e Silva et al.,

Pasting properties
Regarding the pasting properties, only the models for the breakdown and final viscosities of OSP autoclaved starch were significant (p≤0.05); and that for setback for WSP autoclaved starch (Table 1).
The OSP autoclaved starch breakdown fluctuated significantly due to the negative linear and quadratic effects for the reaction time, positive linear and negative quadratic effects for the moisture content and for the time and moisture content interaction. There were also significant negative linear effects of the reaction time, quadratic moisture and the interaction between time and moisture content for the final viscosity of this starch (Table 1).
The OSP autoclaved starch models showed maximum points (final viscosity 3600 cP and breakdown 0 cP) with milder treatments: times shorter than 21 min and moisture contents of 26 to 40% ( Figures 2C and 2D). OSP native starch showed a value of 2334 ± 9 cP for the final viscosity and 1596 ± 18 cP for the breakdown (Paixão e Silva et al., 2020). The increase in the final viscosity of autoclaved sweet potato starches is in agreement with the results found for water absorption, which increased due to the autoclave treatment, which makes the gel more viscous. X.  using an autoclave at 121ºC for 20 minutes in ginger starch also observed an increase in the final viscosity and a reduction in the breakdown of the paste. This can be due the autoclaving breaks down the granular structure of the starch producing solubilized structures and non-solubilized fractions of amylose and/or amylopectin in the form of gelatinized starch (Agama-Acevedo, Pacheco-Vargas, Bello-Pérez, & Alvarez-Ramirez, 2018;Chung, Liu, & Hoover, 2009).

Gel hardness and transparency
The hardness of the OSP autoclaved starch gel was significantly affected by the linear component of time and by the interaction of time and moisture content, which contributed to a reduction in strength of the modified starch gel. However, the quadratic component of time increased the strength of the gel, as shown in Figure 3A. The maximum gel hardness values of this starch (6.5 N) were obtained in less than 7 min. For the hardness of the WSP autoclaved starch gel, only the linear time component contributed to a reduction in strength of the modified starch gel, and its quadratic component increased the gel strength, as shown in Figure 3B. The maximum hardness values (10.0 N) were also determined in less than 7 min. Thus, the maximum gel hardness of the WSP autoclaved starch (10 N) was greater than that of the OSP autoclaved starch (6.5 N). This can be explained because the gelatinization temperature of the WSP starch (76.55 ºC) was higher than that of the OSP starch  The transparency of OSP autoclaved starch was positively influenced by the moisture content used in the modification and negatively influenced by the starch exposure time (Table 1). The highest value (1.2%) was determined with the highest moisture (above 40 %) content and the shortest exposure time (less than 13 min.) ( Figure 3C).
The transparency of this native starch was 4.6 % (Paixão & Silva et al., 2020) higher than the autoclaved one, indicating that the method reduces the transparency of the gel due to degradation of the starch chains .
The paste translucency also decreased for autoclaved cassava starch by Abraham (1993).

SEM
The granules of native sweet potato starches have a polygonal, spherical and semi-spherical shape, with varying sizes, smooth surfaces, without holes or cracks (Paixão e Silva et al., 2020;Wang et al., 2020). The micrographs showed that the shapes and sizes were similar to natives one, however the autoclave process promoted the appearance of scratches, cracks, and depressions, also seen by Babu and Parimalavalli (2013); H.  with modification of sweet potato and ginger starches. The treatments with 30% and 20 % of moisture at 37,5 min. were the ones that showed the greatest change int the surface of granules (Figures 4 A, 4B, 4C and 4D). Additionally, it was possible to observe the presence of pre-gelatinized granules in the OSP autoclaved starch ( Figures   4C and 4D), confirming that the autoclave promotes the break of granular structure, as mentioned in RVA results. Deka and Sit (2016) in their studies using an autoclave to modify taro starch reported that a loss of physical integrity this might be attributed to the partial gelatinization of the starch during the heat moisture treatments as evidenced by swelling, breakup, and/or aggregation/fusion of the granules.

Conclusion
The physical modification by an autoclave proved to be advantageous for modify the analyzed starches. The maximum values of the response surfaces (of the models that were significant) showed an increase of WAI at 60ºC of WSP starch and of WSI at 60ºC of OSP starch; an increase of final viscosity of OSP starch; an increase of setback of WSP starch; an increase of gelatinization enthalpy for both starches; and a reduction of breakdown of OSP starch. Besides that, micrographs showed degradation and pre-gelatinization of the granules.
Therefore, due the promising use the authors suggest future researches with WSP and OSP autoclaved starches to test the technological applications in foods.