Physicochemical and rheological properties of modified rice amylose by dynamic high-pressure microfluidization

ABSTRACT

The effect of dynamic high-pressure microfluidization on physicochemical and rheological properties of rice amylose pastes were examined. The amylose dispersions were pressurized at 60, 100, 140, and 180 MPa. The microfluidization treated rice amylose showed elevated solubility, swelling power, and moisture absorbability. The retrogradability was significantly decreased with the pressure increase. Iodine blue value showed a significant increase at 60~140 MPa, while a slight decrease at 180 MPa. The rheological results indicated that native amylose was a non-Newtonian fluid and displayed pseudoplastic fluids characteristics. The amylose moved toward a Newtonian fluid behavior and its rheopexy decreased with the pressure increased.

KEYWORDS:

• Amylose
• Dynamic high-pressure microfluidization
• Rice
• Physicochemical properties
• Rheological properties

Introduction

Starch is an abundant and widespread polymer that is present in various plant tissues.^[1] Starches have been used in different industries, such as the paper, textile, and food industries, due to their physicochemical properties.^[2] The physicochemical properties of starch are determined by its source and the molecular structures of amylopectin and amylose, which are two major components of starch granules. Rice is one of the most important crops in the world and an important source of starch. However, the application of native rice starch granule is limited by its low solubility, low swelling power, low heat resistance, and uncontrolled paste consistency.^[3,4]

Starch is commonly modified by physical, chemical, or enzymatic methods to obtain the desired physicochemical properties. Given the concern for health issues, physical modification methods are preferred in the food industry. High-pressure technologies, such as high-hydrostatic pressure (HHP) treatment, have been applied in the preparation of modified starch.^[5] The alteration of the morphological characteristics and crystalline structure of starch by HHP has been reported.^[6] Yun et al.^[7] indicated that appropriate HHP processing could significantly improve the functionality of rice starch. Hu et al.^[8] suggested that the retrogradation rate of rice starch gelatinized by HHP is slower than that of starch gelatinized by heat.

Dynamic high-pressure microfluidization (DHPM) is an innovative high-pressure technology that is different from HHP. Combined strong forces are exerted on starch during DHPM treatment, including high-velocity impact, high-frequency vibration, instantaneous pressure drop, intense shear, and cavitation.^[9] In addition, DHPM can achieve ultra-high pressure (up to 200 MPa) and continuous operation. This treatment has been considered as a promising method in improving many food constituents,^[10,11] for example, improving the physicochemical properties of whey protein.^[12]

In this study, we investigated the effects of DHPM treatment on the physicochemical properties of rice amylose. Rice amylose was isolated and subjected to DHPM at pressures ranging from 60 to 180 MPa. The physicochemical properties, including the solubility, swelling power, moisture absorbability, iodine blue value, retrogradability, and rheological characteristics, were evaluated before and after DHPM treatment.

Materials and methods

Materials and reagents

Commercial rice starch was purchased from Dacheng Industrial Group Co. (Changchun, China). All of the chemicals (analytical grade) were purchased from Da Mao Reagent Co. (Tianjin, China).

Isolation and purification of rice amylose

Rice amylose was isolated and purified using the method of Takeda et al.^[13] with minor modifications. Rice starch (10 g) were filtrated with a small amount of anhydrous ethanol and dissolved in 250 mL of 0.5 mol·L⁻¹ NaOH. The suspension was placed in a boiling water bath and incubated for 30 min. After the starch solution was cooled, the undissolved substances were removed by centrifugation (Shanghai Anting Science and Technology Instrument Company, China) at 3000 rpm for 20 min. The pH of supernatant was then adjusted to neutral with 2 mol·L⁻¹ of HCl before addition of 200 mL of butanol–isopentanol (v/v, 3:1) into the mixture. The mixture was incubated in boiling water for 25 min and centrifuged at 3000 rpm for 20 min. The precipitate was then used as crude rice amylose. Subsequently, the crude rice amylose was dissolved in 120 mL of an n-butyl alcohol aqueous solution (the volume ratio of n-butyl alcohol and water was 1:9). The mixture was stirred in a boiling water bath until the dispersed solution was transparent. The mixture was then cooled in a refrigerator at 2–4°C for 24 h. The precipitate was obtained by centrifugation at 3000 rpm for 20 min. The above procedure was performed three times, and then the precipitate was filtrated with pure ethyl alcohol six times. The purified rice amylose was obtained after lyophilization. The extracted rice amylose content was estimated by iodine colorimetry based on the method of Hoover and Ratnaykae.^[14] The amylose content of the extracted starch sample was about 85%.

DHPM treatment of rice amylose

A rice amylose suspension (6%, w/v) was prepared in deionized water at 25°C with stirring. The mixture was treated with a micro jet homogenizer (M-700; Microfluidics Company, USA) at 60, 100, 140, and 180 MPa, respectively. After which, 500 mL of each DHPM-treated rice amylose suspension was collected, then these samples were lyophilized (ALPHA 1-2; Martin Christ Company, Germany).

Solubility and swelling power

The solubility and swelling power of DHPM-treated and untreated amylose were determined using the method of Choi et al.^[3] with minor modifications. The 2% (w/v) amylose suspension 250 mL was divided into five aliquots. Four parts were separately heated at 30, 50, 90, and 110°C in an oil bath for 30 min with continuous stirring at 75 rpm. The gelatinized samples were cooled to room temperature and centrifuged at 4800 rpm for 20 min. The resulting supernatant was further dried at 100°C and then weighed repeatedly until a constant weight was achieved. The solubility (90 and 110°C) and swelling power (30 and 50°C) were calculated by using the following formulas:

where W is the weight of dry amylose (g), A is the weight of the soluble amylose (g), and P is the weight of precipitate in centrifugal tube (g).

Moisture absorption

The moisture absorbability of DHPM-treated and untreated amylose was investigated using the method of He et al.^[15] with minor modifications. The samples (0.5 g) were accurately weighed and transferred into an incubator (Beijing Hualeda Science and Technology, Ltd., China). After incubation at 25°C for 50 h, the samples were weighed again. The tests were performed in triplicate, and the moisture absorbability (Ma) was calculated as follows:

where W₀ and W are the dry-weight values of amylose before and after incubation, respectively.

Iodine blue value

The blue value was measured as described by Shi et al.^[16] with some modifications. Rice amylose (0.1 mg) was dissolved in 1 mL iodine reagent (contain iodine 0.02 g, potassium iodide 0.04 g), then the solution was made up to 10 mL with distilled water. The blue value of samples at 680 nm were measured using ultraviolet (UV)-visible spectrophotometer (UV-T6, Beijing PERSEE company, China) according to the method of Yu,^[17] the blue value was calculated with the absorbance measured at 680 nm, according to the following equation:

where BV: blue value, Abs₆₈₀: absorbance at 680 nm, and C concentration of starch in the solution in 1 mg/100 mL.

Retrogradation

Retrogradability of DHPM-treated and untreated rice amylose was determined according to the method of Singh et al.^[18] with minor modifications. The amylose suspension (1%, w/v) was fully gelatinized by heating in boiling water for 20 min with continuous stirring at 75 rpm. The amylose paste was transferred to a measuring cylinder at approximately 25°C. The volumes of the supernatant and precipitate were measured, and the degree of retrogradation was expressed in terms of the ratio of supernatant to precipitate.

Rheological measurement

The rheological characteristics of DHPM-treated and untreated rice amylose were evaluated using the methods reported by Xiao et al.^[19] with some modifications. Amylose suspension (2%, w/v) was fully gelatinized by boiling in a water bath for 30 min, with agitation every 5 min. The amylose paste was then placed onto the testing platform of a dynamic rheometer (Viskograph-E; Brabender Company, Germany). The viscosity was measured using a controlled strain rheometer with a parallel plate mould (40 mm diameter and 1 mm gap). After trimming off the over-loaded portion of a sample around the plates, the sample was covered with a thin layer of silicon oil to prevent moisture loss. To measure thixotropic loop of amylose, rotor speed was uniformly increased from 5 to 50 r s⁻¹ and then uniformly slowed down to 5 r·s⁻¹. Correspondingly, the shear rate increased from 10 to 100 s⁻¹ and then decreased to 10 s⁻¹. The shear stress of the amylose paste was recorded during the measurement. The flow behaviors of samples were obtained at 25°C. The flow behavior index was described by fitting the experimental data (shear rate-shear stress) with the power law model by using the following equation:^[20]

where τ is shear stress, k is consistency coefficient (Pa), γ is shear rate (s⁻¹), and n is the flow behavior index. n = 1 for Newtonian liquids, n < 1 for shear-thinning samples, and n > 1 for shear-thickening samples.

Statistical analysis

All experiments were performed in triplicate, and data were reported as mean values ± standard deviations. The differences among varieties were analyzed by one-way analysis of variance (ANOVA) and Tukey’s multiple comparison tests. The statistical significance was defined at p < 0.05.

Results and discussion

Solubility and swelling power

The solubility and swelling power of native and DHPM-treated rice amylose are shown in Figs. 1 and 2. As shown in Fig. 1, DHPM treatment resulted in a significant increase in the solubility of rice amylose at 90 and 100°C. Meanwhile, similar results were obtained for the swelling power at 30 and 50°C as shown in Fig. 2. When amylose is heated in water, the crystalline structure is disrupted and water molecules become linked by hydrogen bonding to the exposed hydroxyl groups of amylose.^[21] DHPM treatment had a significant impact on the amorphous–crystalline transition and granule morphology of amylose.^[22] When pressure was applied to the amylose suspension, the water molecules entered the broke amylose structure and formed hydrogen bonds with amylose polymers. As a result, this caused an increase in the amylose swelling and solubility.^[21,23] Oh et al.^[24] reported that the degree of swelling for normal rice starch increased as the HHP treatment pressure increased to 500 MPa.

Figure 1. Effect of DHPM on the solubility of rice amylose.

Figure 2. Effect of DHPM on the expansion ratio of rice amylose.

In addition, the solubility and swelling power were significantly increased as the temperature increased from 90 to 110°C and 30 to 50°C, respectively. This trend was in agreement with the results of Guo et al.,^[20] who reported that the swelling power and solubility of lotus seed starch both increased as the temperature increased from 55 to 95°C. Yu et al.^[17] also reported that the solubility of rice starch was positively correlated to the temperature. Torre-Gutiérrez et al.^[25] reported that the solubility and swelling power of square banana starch and corn starch increased with the increase in temperature from 70 to 90°C.

Moisture absorption

Amylose is a polysaccharide that readily absorbs moisture because its macromolecules contain numerous hydroxyl groups.^[15] The moisture absorbability of rice amylose treated by DHPM is presented in Fig. 3. The native rice amylose exhibited the lowest moisture absorbability, with a value of 3.90%. When the pressure was 60 MPa, no obvious increase was observed in the moisture absorbability of rice amylose. When the pressure ranged from 100 to 180 MPa, DHPM treatment significantly increased the moisture absorbability of rice amylose, increasing from 4.80 to 6.09%. The absorption of water is related to its rate of diffusion into the amylose granules.^[26] The granular structure and blocks of gel-like regions of amylose are suggested to be destroyed by DHPM. Under high pressure, the softening of the amorphous region changed the granule shape.^[22] Moreover, the original smooth surface of amylose granules became rougher and showed pores and fissures after DHPM treatment. Some coarseness or damage had been generated on the surface, which may increase the surface area, expose free hydroxyl groups, and enhance water diffusion into the crystalline region, thereby contributing to an increase in the moisture absorbability of rice amylose.^[5,7,22] These results were agreement with those of Oh et al.,^[24] in which the crystalline regions were distorted by pressure and became more accessible to water.

Figure 3. Effect of DHPM on the moisture absorbability of rice amylose.

Iodine blue value

Iodine blue value of rice amylose after different pressure DHPM treatment were shown in Table 1. The iodine blue value was increased when DHPM pressure was applied, ranged from 0.863 to 1.113. When the pressure was 140 Mpa, the OD_680nm showed the highest value of 0.1391 among all pressure treatments. The DHPM processing may make the chain rupture by mechanical force, therefore, increasing the amount of amylose that can react with iodine. In order to prove this, the content of amylose was tested. The result was shown in Table 1. The native amylose content was 84.74%, with the increase of DHPM pressure, the content of amylose ranged from 92.41~95.67%. The iodine blue value decreased when DHPM pressure was raised to as high as 180 MPa. Most likely, the linear chain molecules bond was disrupted by high pressure DHPM to form some short straight chains, which cannot react with iodine. Yu et al.^[17] reported similar results, in which the maximum starch-iodine absorbance was positively related to amylose content.

Table 1. Content and iodine blue value of rice amylose after DHPM treatment under different pressures.

Sample	Iodine blue value	Amylose content (%)
Amylose	0.863 ± 0.003^a	84.74 ± 0.02^a
60 MPa	0.921 ± 0.007^b	92.41 ± 0.01^b
100 MPa	1.054 ± 0.001^d	95.67 ± 0.02^c
140 MPa	1.113 ± 0.005^e	93.21 ± 0.03^b
180 MPa	1.007 ± 0.008^c	93.10 ± 0.01^b

Mean values ± standard deviations of three determinations.

Sample means with different lower case letters in the same column are significantly different at p < 0.05.

Retrogradation

Gelatinized amylose recrystallizes during storage, thereby affecting the texture and shelf life of food products. This phenomenon is known as retrogradation. Better understanding of the retrogradation process in starch is essential for product development and quality control.^[6] As shown in Fig. 4, the retrogradability of amylose rapidly increased in the first 10 h and reached a plateau at 12 h. With the increase of DHPM pressure, the retrogradability of amylose decreased. This indicates that the stability of amylose increased. The results were in accordance with Guo et al.,^[20] who reported that ultra-high pressure treated lotus seed starch had a lower retrogradation tendency as compared to native starch. In addition, Doon et al.^[27] reported that less retrogradation was observed in pressurized starch. This result may be related to a smaller volume of freezable water when starch is subjected to ultra-high pressure. However, conflicting results have been reported by Kiyoshi et al.,^[28] in which retrogradability of potato starch was promoted by HHP treatment at low temperature. This result may be dependent on the botanical sources of starch.

Figure 4. Effect of DHPM on syneresis of rice amylose.

Rheological measurement

The apparent viscosity and shear stress of rice amylose paste by DHPM treatment is shown in Figs. 5 and 6, respectively. Regardless of the DHPM treatment, the apparent viscosity of amylose fell sharply at low shear rate, whereas this value gradually stabilized under high shear rate. Generally, the apparent viscosity and shear stress of amylose decreased after the rice amylose paste was homogenized at different pressures. The apparent viscosity of the suspension containing macromolecules is a function of the intermolecular force, which restricts molecular motion.^[29] After DHPM treatment, the strong mechanical forces caused the fracture and degradation of amylose, which has a longer chain. During DHPM treatment, the ruptured amylose macromolecule produced modified amylose with much lower molecular weight and much smaller molecular size. Consequently, the entanglement between macromolecules was possibly decreased, thereby resulting in the significantly lower apparent viscosity of the pastes. Similar results were observed by Fu et al.^[30] and Wang et al.,^[31] both groups reported that the apparent viscosity and shear stress of starch pastes obviously decreased within the shear rate range during high pressure treatment.

Figure 5. Shear thinning of rice amylose after DHPM treatment under different pressures.

Figure 6. Effect of DHPM on the rheological properties of rice amylose.

The rheological index (n), consistency coefficient (k), and the correlation coefficient (R²) of the straight-chain starch paste before and after DHPM processing treatment can be obtained by non-linear regression fitting (Table 2). The experimental data for starch to the power law model was found to have a good fit with a high correlation coefficient (R² = 0.9923–0.9994). With the increased dealing pressure, k decreased from 0.5811 ± 0.015 to 0.0476 ± 0.001, whereas n increased from 0.4281 ± 0.017 to 0.5696 ± 0.003, thereby demonstrating shear-thinning behavior; this change signifies the reduction of apparent viscosity with the increase in pressure.^[32] The results indicated that rice amylose samples showed non-Newtonian behavior and pseudoplastic behavior, with n being well below unity. With the increased pressure, the value of n followed a rising tendency, thereby showing that the amylose paste moved toward Newtonian fluid behavior after DHPM treatment.^[31]

Table 2. Parameters of rheological curves of rice amylose after DHPM treatment under different pressures.

Sample	k (Pa·s^−m)	n	R^2
Amylose	0.5811 ± 0.015^d	0.4281 ± 0.017^a	0.9957
60 MPa	0.1983 ± 0.004^c	0.4787 ± 0.002^b	0.9923
100 MPa	0.1018 ± 0.008^b	0.5180 ± 0.012^c	0.9987
140 MPa	0.0476 ± 0.001^a	0.5696 ± 0.003^e	0.9987
180 MPa	0.0592 ± 0.003^a	0.5419 ± 0.005^d	0.9994

Mean values ± standard deviations of three determinations.

Sample means with different lower case letters in the same column are significantly different at p < 0.05.

Thixotropic curves

The thixotropic curves of rice amylose before and after DHPM processing at 25°C are shown in Fig. 7. After the shearing loop, a lag and a lag circle was observed between the upward and downward curves of shear stress. The lag circle of the flow curves indicated that the damage on the amylose network as caused by shear stress. The size of the lag circle represents the size of thixotropy.^[33] From Fig. 7, the native rice amylose paste showed thixotropy, whereas the thixotropy of amylose was reduced after DHPM treatment. Similar results were observed by Wang et al.,^[31] who reported the area of the hysteresis loop became much smaller after HHP treatment, thereby indicating that the thixotropic behavior of the samples was weakened.

Figure 7. Thixotropic curve of rice amylose granules treated under different conditions: A: native, B: 60 MPa, C: 100 MPa, D: 140 MPa, and E: 180 MPa.

Conclusions

In this study, the effects of DHMP on the solubility, swelling power, moisture absorbability, iodine blue value, retrogradability, and rheological characteristics of rice amylose pastes were investigated. The applied pressures in the DHPM treatment were 60, 100, 140, and 180 MPa. The solubility, swelling power, and moisture absorbability of rice amylose pastes increased alongside homogenizing pressure of the DHPM treatment. After DHPM treatment, the iodine blue value increased because of increase in amylose. The retrogradability of paste decreased significantly, which indicated that DHPM treatment could increase amylose stability. The apparent viscosity values and shear stress of rice amylose pastes decreased after DPHM treatment. Results of the rheological measurements indicated that amylose exhibited non-Newtonian behavior and shear thinning behavior after DHPM treatment. Meanwhile, the thixotropy of amylose was slightly reduced after DHPM treatment. Therefore, the study on DHPM treatment of the rice amylose provides important information in developing modified starch and help broaden the use of DHPM in starch industry.

Funding

This work was supported financially by Key Project for Science and Technology Innovation of Jiangxi Province (20124ACB00600).

Additional information

Funding

This work was supported financially by Key Project for Science and Technology Innovation of Jiangxi Province (20124ACB00600).