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. 2026 Oct 7;39:104597. doi: 10.1016/j.fochx.2026.104597

Construction of electrostatic binding V-type starches using cationic starch and lauric acid: Insight into enhanced digestive resistance

Zhenfei Ding a,c,1, Wenjing Li b,1, Chao Xu b, Yuanhang Wang c, Xu Chen b,⁎
PMCID: PMC13677270  PMID: 42898054

Abstract

This study developed a V-type starch with enhanced structural stability and digestive resistance through an electrostatic interaction between a cationic starch (CS) and lauric acid (LA). X-ray diffraction analysis revealed that the non-etherified sample complexed with LA (ETA-0%-LA) exhibits a typical B-type crystalline structure. When the ETA content was increased from 5% to 30%, the V-type crystallinity of the CS–LA complexes increased from 9.22% to 24.60%. The degree of substitution (DS) exhibited a clear dose-dependent response to the etherifying agent, with ETA-30% yielding the highest DS (0.075%). Dynamic viscoelastic analysis further indicates that the electrostatic interaction between CS and LA substantially strengthened the gel network structure of CS–LA complexes. In vitro digestion tests confirmed that the moderate cationization markedly increasing resistant starch content and decreasing rapidly digestible starch content. These findings indicate that electrostatic interaction plays a key role in modulating the structural stability and digestive resistance of V-type starch.

Keywords: Cationic starch, Lauric acid, Electrostatic interaction, V-type starch, Digestive resistance

1. Introduction

In general, the linear component of starch, which mainly comprises amylose and some long side chains without amylopectin branching can interact with hydrophobic molecules and form inclusion complexes (ICs) in a V-type crystalline structure (Di Marco et al., 2022; Putseys et al., 2010). In the presence of suitable ligands, amylose undergoes conformational changes that result in a single left-handed V-type crystalline structure with an inner hydrophobic helical cavity. Various hydrophobic small molecules, organic solvents, and fatty acids can be accommodated in this cavity (Wang et al., 2020; Yu et al., 2026). Fatty acids have been encapsulated into amylose complexes to stabilize the starch guest compounds in environments unfavorable to their encapsulation and facilitate their controlled release (Birt et al., 2013). Because of their versatility and low cost, starch–guest assemblies have attracted major attention as an excellent vehicle in functional foods (Tan & Kong, 2020).

Ethanol-induced V-type granular starch (EVGS) formation is generally used to modify cold-water swelling starch. EVGS is commonly prepared through ethanol-aqueous heat treatment and ethanol–alkaline treatment (Lai et al., 2024; Li et al., 2022; Li et al., 2024). EVGS is soluble in cold water and produces higher-viscosity solutions compared with native starch (Li et al., 2025). EVGS can be used as an encapsulation material for certain guest molecules at room temperature, with suitability for encapsulation depending on the formation of a V-type single-helix cavity structure (Gao et al., 2022). A major limitation of EVGSs lies in their structural instability in aqueous systems, which restricts their application in drug encapsulation and sustained-release processes.

Cationic starch (CS), a chemically modified starch derivative functionalized with quaternary ammonium or tertiary amine groups, exhibits unique properties derived from its positively charged surface (Chatterjee et al., 2023; Wang et al., 2023). The primary synthesis methods include using 2,3-epoxypropyl trimethylammonium chloride (EPTAC) as the etherifying agent and reacting EVGS with starch hydroxyl groups through nucleophilic substitution under alkaline conditions (Suflet et al., 2015). This modification enables CS to form three-dimensional networks through either physical or chemical cross-linking, establishing it as an effective drug carrier system for controlled release (Chatterjee et al., 2023). Lauric acid (LA) carries a negative charge in alkaline environments, which enables robust electrostatic interactions with the positively charged surface of CS (Grządka, 2015; Merta & Stenius, 1999; Zhang et al., 2012). A composite hydrogel integrates the intrinsic positive charges of CS with the negative charges of LA, an interaction that confers a marked advantage in stabilizing the V-type single-helix cavity structure of the resulting complex. Although oral consumption of cationic starch is currently regulated due to its potential cytotoxicity, the direct toxicity data for EPTAC-modified cationic starches are limited. Due to their abundant availability, low cost, easy chemical modification, biodegradability and biocompatibility, extensive research is now being exploited in drug delivery, tissue engineering and biomedicine (Carmona et al., 2026; Chatterjee et al., 2023).

Instead of relying on the traditional ethanol-induced method, which is prone to disruption in aqueous environments, this study utilized native high-amylose maize starch as the starting material for cationic modification. We hypothesized that introducing positive charges onto the starch backbone would drive robust electrostatic interactions with anionic lauric acid during gelatinization, spontaneously forming a stable V-type structure that resists aqueous dissociation. To comprehensively characterize synthesized CS and CS–LA complexes, a multiscale analytical approach incorporating scanning electron microscopy (SEM), particle size distribution analysis, zeta potential measurements, Fourier-transform infrared (FTIR) spectroscopy, 1H-nuclear magnetic resonance (1H NMR) imaging, and X-ray diffraction (XRD) was employed. Rotational rheometry was used to characterize the shear recovery behavior of CS and the gelation characteristics of CS–LA complexes. In vitro digestion assays were performed to determine the digestibility of the CS and CS–LA complexes. This study developed an electrostatic binding V-type starch through an interaction between CS and LA; this complex exhibited high stability and digestive resistance.

2. Materials and methods

2.1. Materials

High-amylose maize starch (HAMS, purity: 98.52%, amylose content: 60.74%, crude fat: 0.36%, crude protein: 0.14%) was sourced from Henan Hengrui Starch Technology Co., Ltd. (Henan, China). EPTAC (molecular weight: 151.63) and lauric acid (LA) were purchased from Shanghai Macklin Biochemical Co., Ltd. (Shanghai, China). α-amylase from hog pancreas (10,080, 290 U/mL) and amyloglucosidase (CAS 9032-08-0, 100 U/mL) were purchased from Shanghai Macklin Biochemical Co., Ltd. (Shanghai, China). All auxiliary reagents used in this study were of analytical grade.

2.2. Preparation of the cationic starch

Cationic starch samples were synthesized under varying reaction conditions using high-amylose maize starch (3 g) dispersed in 30 mL of deionized water within a high-temperature reactor. The reaction system was maintained at 55 °C during starch heating. NaOH solutions (1.0 mol/L) were introduced, followed by 90 min of stirring at 55 °C. Subsequently, EPTAC was added at concentrations of 0, 5, 10, 15, 20, and 30 wt% (based on starch dry weight), and the etherification reaction proceeded for 6 h at 55 °C under continuous stirring. The resulting products were freeze-dried and ground through a 100-mesh sieve to obtain cationic starch powders. Samples prepared with 0 wt%, 5 wt%, 10 wt%, 15 wt%, 20 wt%, and 30 wt% EPTAC were labeled ETA-0%, ETA-5%, ETA-10%, ETA-15%, ETA-20%, and ETA-30%, respectively. Cationic starch (2.0 g, dry weight) was dispersed in 20 mL deionized water to prepare a 10% (w/v) suspension, which was heated at 55 °C for 30 min under stirring, then adjusted to pH 8.0 with 0.1 M NaOH solution. LA (15 wt%, dry weight) was firstly dissolved in ethanol, heated at 65 °C for 30 min, then added dropwise into the starch suspension. The mixture was held at 65 °C for 2 h with stirred constantly to obtain CS-LA complex. Whereafter, the sample was cooled at 25 °C and washed with anhydrous ethanol to remove uncomplexed LA. Then freeze-dried CS-LA complex was passed through a 200-mesh sieve to obtain CS-LA complex powders. The nitrogen content of CS, determined by the Kjeldahl method, was used to calculate its degree of substitution (DS):

DS%=162×N14×100−151.63×N×100%

where N represents the nitrogen content in the sample.

2.3. Scanning electron microscope (SEM) of cationic starch

The CS powder prepared in Section 2.2 was uniformly applied onto a sample platform pre-coated with double-sided conductive adhesive tape. A vacuum evaporator was employed to spray a thin layer of platinum on the surface of the powder for 30 s. Subsequently, scanning electron microscopy (Flex1000, Hitachi, Japan) was performed to characterize the sample morphology at an accelerating voltage of 3.0 kV, with an imaging magnification of 3000 × .

2.4. Zeta potential of cationic starch

A 0.1% (w/v) starch suspension was prepared by dispersing the sample from Section 2.2 in deionized water, followed by vortex oscillation to ensure homogeneous dispersion, and the PH of all suspensions was adjusted to and maintained at 7.0 prior to the measurements. The zeta potential of the suspension was subsequently measured using a Zetasizer Nano (ZSU3100, Malvern Instruments Ltd., Britain) at 25 °C.

2.5. 1H-nuclear magnetic resonance (1H NMR) of cationic starch

The cationic starch synthesized in Section 2.2 was dissolved in deuterated dimethyl sulfoxide (DMSO‑d6) for 1H NMR analysis. 15 mg of the starch sample was dispersed in 0.6 mL of DMSO‑d6. To ensure complete dissolution, the mixture was heated at 60 °C for 1 h and subsequently ultrasonicated for 30 min. After cooling, the cationic starch sample was collected and transferred into an NMR tube. The 1H NMR spectra were acquired using a nuclear magnetic resonance spectrometer (Ascend™, Bruker Biospin GmbH, Germany) at a magnetic field of 600 MHz. Data processing and spectral analysis were performed using MestReNova software.

2.6. X-ray diffraction (XRD) of CS and CS-LA complexes

The samples were sieved through a 200-mesh sieve to ensure particle size homogenization, followed by XRD analysis was subjected. The crystal structure of the samples was characterized using a X-ray diffractometer (SmartLab SE, Rigaku Corporation, Japan) with Cu Kα radiation (λ = 1.5418 Å). The diffraction patterns were collected over a 2θ range of 5°–50° at a scanning rate of 5°/min, and the data analysis were analyzed using MDI Jade 6.0 software. The calculation formula of relative crystallinity is as follows (Liu et al., 2024):

Relative crystallinity%=IcIc+Ia×100%

where Ic represents the area of the crystalline region, and Ia denotes the area of the amorphous region.

2.7. Fourier transform infrared spectroscopy (FTIR) of cationic starch

Thoroughly mix 1 mg of powdered sample from Section 2.2 with 100 mg of potassium bromide (KBr) to prepare a homogeneous blend, followed by compression into a thin pellet using a hydraulic press. The FTIR spectrum of the sample was acquired using a Fourier infrared spectrometer (IS50, Thermo Nicolet Corp., USA) over a wavenumber range of 4000–400 cm−1 at a spectral resolution of 4 cm−1. Data processing, including baseline correction and peak identification, was conducted using Omnic 9.2 software.

2.8. Differential scanning calorimetry (DSC) of CS-LA complexes

The mixed powder, prepared as described in Section 2.7 was subjected for differential scanning calorimetry (DSC) analysis. Specifically, 2 mg of this powder was placed in a crucible, mixed with deionized water at three times its volume, and then sealed and equilibrated overnight. Subsequently, the crucible was heated from 30 °C to 110 °C at a rate of 10 °C/min using a DSC 8000 calorimeter (Perkin Elmer Co. Ltd., Massachusetts, USA). The onset temperature (To), peak temperature (Tp), conclusion temperature (Tc), and enthalpy change (ΔH) were recorded.

2.9. Thermogravimetric analysis (TGA) of CS-LA complexes

Thermal stability of the samples was assessed by thermogravimetric analysis using a Discovery TGA55 instrument (TA Instruments, USA). 2.0 mg of sample was precisely weighed and loaded into a solid crucible. The measurement was conducted under a nitrogen with a flow rate of 60 mL/min, while the temperature was increased from 30 °C to 600 °C at a heating rate of 10 °C/min in ramp mode. The temperature corresponding to the maximum rate of weight loss was determined using the built-in TRIOS software of the instrument.

2.10. Rheological properties of CS-LA gels

The cationic starch samples were prepared in accordance with the preparation protocol specified in Section 2.2. Briefly, 3.0 g of the cationic starch sample was dispersed into 30 mL of deionized water under continuous stirring at 25 °C. Subsequently, the resulting starch suspension was heated in a 100 °C water bath for 30 min with continuous magnetic stirring to ensure homogeneous mixing. After CS–LA gels were synthesized and cooled to 25 °C, and the sample was subjected to oscillatory frequency tests with a rotational rheometer (DHR-1, TA Instruments, USA) equipped with a 40 mm diameter parallel plate geometry. The frequency range was set from 0.1 to 100 rad/s at a fixed strain of 1% (within the linear viscoelastic region), and the storage modulus (G′) and loss modulus (G″) were recorded. In addition, the steady shear tests were conducted at flow scanning mode over 0.1 to 100 s−1 and 100 to 0.1 s−1. The viscosity and stress were recorded using the instrument's proprietary software. Furthermore, gels that had been soaked in NaCl solution were tested under the aforementioned conditions to examine the influence of electrostatic shielding on the rheological properties of the system.

2.11. In vitro digestion of CS-LA complexes

The in vitro digestibility of the CS-LA complexes were assessed using a modified version of the method described by Liang et al. (2023). Briefly, 200 mg of CS-LA complex was mixed with 15 mL of acetic acid buffer (PH 5.2), followed by the addition of 10 mL of enzyme solution containing α-amylase and amyloglucosidase were purchased. The mixture was incubated at 37 °C. At 0, 20 and 120 min, aliquots of 0.25 mL were withdrawn and mixed with 4.75 mL of anhydrous ethanol to stop the enzymatic reaction. Glucose content was determined using a glucose assay kit (GOPOD). The contents of rapidly digestible starch (RDS), slowly digestible starch (SDS), and resistant starch (RS) were calculated according to Hao et al. (2023).

2.12. Statistical analysis

All experiments were conducted in triplicate, and the results are expressed as mean ± standard deviation. All statistical analyses were conducted using IBM SPSS Statistics version 25 (IBM, Armonk, NY, USA). One-way analysis of variance was used to assess significant differences between group means. Differences were considered statistically significant at p < 0.05.

3. Results and discussion

3.1. SEM images of cationic starch

Fig. 1 presents SEM images of CS synthesized with varying etherifying agent concentrations. The morphological characteristics of the control starch (ETA-0%), subjected to alkaline and heat treatment without the etherifying agent (Fig. 1A) closely resembled those of natural high-amylose maize starch, exhibiting heterogeneous particle morphologies, including spherical, irregular, and slender morphologies, with smooth surfaces. When the content of the etherifying agent was increased to 5% and 10% (Fig. 1B and C), the starch granules exhibited slight swelling, accompanied by the formation of distinct concavities in the hilum regions. This morphological evolution may be attributable to concentration-dependent alkaline effects. Higher concentrations of the etherifying agent disrupted the native hydrogen bond network within the starch granules, preferentially damaging the structurally vulnerable hilum region, which consists of an incomplete crystalline area and loosely packed amylopectin (Chi et al., 2022). When the content of the etherifying agent was further increased, the degree of morphological damage to the samples also increased. When the content reached 30%, pronounced collapse of the hilum region was observed (Fig. 1F). This morphological change can be ascribed to the etherifying agent causing greater damage to the starch granules at higher concentrations (Zheng et al., 2015).

Fig. 1.

Fig. 1

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Scanning electron microscopy images of cationic starches synthesized under various etherifying agent contents (A–F: 0%–30%).

3.2. Zeta potential and degree of substitution of cationic starch

Fig. 2A depicts the average particle size of different samples of CS and reveals no significant between-sample differences, indicating that the particle size of starch did not change during cationization. In addition, the zeta potential of CS synthesized under various conditions was measured (Fig. 2B). Before the etherifying agent was added, the zeta potential of the starch was −17.63 mV. When 5% etherifying agent was added, the starch particles exhibited a zeta potential of 18.22 mV, indicating that the surface of the high-amylose maize starch carried positive charges after modification. When the concentration of the etherifying agent was further increased to 30%, the zeta potential of the sample increased to 25.36 mV. This increase in zeta potential may be attributable to the higher concentration of the etherifying agent in the mixture, which facilitated the generation of additional positive charges on the starch surface and thereby increased its electrical potential. These zeta potential results are highly consistent with the trend observed in the DS, indicating a strong correlation between the surface charge density of starch and the extent of cationic substitution (Kaur et al., 2012). In summary, the high zeta potentials of the synthesized CS demonstrate its substantial potential for application in systems requiring electrostatic interactions with negatively charged substances.

Fig. 2.

Fig. 2

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Average size (A) and zeta potential values (B) and 1H-nuclear magnetic resonance (C) and degree of substitution (D) of cationic starches synthesized under various etherifying agent contents.

Fig. 2C presents the 1H NMR spectra of CS synthesized under varying conditions. A new characteristic peak at 3.20 ppm was observed in all CS samples relative to the ETA-0% group, corresponding to the methyl protons of the quaternary ammonium group introduced by EPTAC (Chang et al., 2014). This distinct spectral feature confirmed the successful introduction of cationic groups into the starch matrix. To quantitatively evaluate cationization efficiency, the DS was calculated (Fig. 2D). The results indicated that the DS exhibited a clear dose-dependent response to the etherifying agent, with the 30 wt% concentration (ETA-30%) yielding the highest DS (0.075%). This dose-dependent response likely occurred because the higher concentration of the etherifying agent increased the frequency of molecular collisions between the etherifying agent and starch, thereby promoting a substitution reaction (Chang et al., 2024). This phenomenon aligns with previous finding that the electrostatic interaction between CS and negatively charged substances is primarily governed by surface charge characteristics, with zeta potential serving as a crucial determinant (Anthony & Sims, 2013).

3.3. FTIR and XRD analysis of cationic starch

The FTIR spectra of CS synthesized under various conditions are depicted in Fig. 3A. The characteristic absorption bands observed at 2930 cm−1 (C—H stretching vibration), 1160 cm−1 (C—O—C asymmetric stretching), and the characteristic absorption band at 1020 cm−1 corresponds to the C—O stretching vibrations of the anhydroglucose ring in the starch backbone (Warren et al., 2016). After starch was modified into CS, the C—O—C asymmetric stretching and C Created by potrace 1.16, written by Peter Selinger 2001-2019 C stretching vibration peaks of the samples exhibited obvious red shifts from 1163 cm−1 (ETA-0% group) to 1156 cm−1 (ETA-30% group). By contrast, the C—H stretching vibration peak exhibited a blue shift from 2925 cm−1 (ETA-0% group) to 2931 cm−1 (ETA-30% group). These shifts in the stretching vibrations of the characteristic peaks suggested substantial modifications to the starch molecular groups after cationization. The introduction of quaternary ammonium groups from EPTAC altered the electron density distribution around the hydroxyl and ether linkages of the starch, thereby affecting their vibrational characteristics. Specifically, the red shifts in C—O—C stretching vibrations may be attributable to electrostatic repulsion between adjacent cationic groups, which weakened the C—O bonds; the blue shift in C—H stretching likely resulted from enhanced hydrogen bonding between hydroxyl groups and the grafted cationic moieties (Balsamo et al., 2011; Suflet et al., 2015). Fig. 3B presents the deconvolution spectra of CS synthesized under varying conditions. Compared with high-amylose maize starch without an etherifying agent, the ratio of 1047/1022 cm−1 in all samples decreased after cationization, indicating that the cationization process reduced the short-range orderliness of the starch (Chang et al., 2014).

Fig. 3.

Fig. 3

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Fourier transform infrared spectroscopy (A) and Fourier self-deconvolution spectra (B) of cationic starches synthesized under various etherifying agent contents. X-ray diffraction pattern (C) and relative crystallinity (D) of complexes formed between lauric acid and cationic starch synthesized under various etherifying agent contents.

Fig. 3C illustrates the XRD patterns of CS samples synthesized with varying etherifying agent concentrations. The ETA-0%–LA sample demonstrated characteristic B-type crystalline diffraction peaks at 17.09°, 19.76°, and 22.79° (Yang et al., 2013). This sample exhibited diffraction peak intensities nearly identical to those of native high-amylose starch, suggesting that this specific pre-treatment condition retained the inherent B-type crystal characteristics of the high-amylose maize starch. By contrast, the ETA-5%–LA sample indicated that the B-type diffraction peaks almost completely disappeared, accompanied by the emergence of weak V-type diffraction peaks at 12.86° and 19.79°. This pattern change may be attributable to the disruption of the original crystalline framework by the addition of the etherifying agent, which promoted the complexation of leached amylose molecules with LA, thereby facilitating the formation of new V-type crystals (Yang et al., 2025). Notably, the V-type crystallinity remained statistically unchanged at low etherifying agent dosages (5% and 10%), but increased significantly from 9.22% to 24.60% as the dosage was further elevated to 30% (Fig. 3D). This may have occurred because with increasing etherifying agent concentrations, the surface positive charge of the prepared CS increased, enabling it to more effectively attract the negatively charged LA, thus facilitating the formation of a V-type helical structure between the amylose and LA (Wang et al., 2020).

3.4. DSC and TGA analysis of cationic starch-lauric acid complexes

The thermal properties of the CS–LA complexes prepared with varying concentrations of the etherifying agent were investigated using DSC; the corresponding thermograms are presented in Fig. 4A. The ETA-0%–LA sample, which did not undergo cationic modification, exhibited no obvious endothermic peak in the range of 30°–120 °C. This poor thermal response suggests that the unmodified starch surface failed to interact with the negatively charged LA because it was itself negatively charged, resulting in negligible complex formation with LA. By contrast, the ETA-5%–LA and ETA-10%–LA samples exhibited a slight endothermic peak relative to the control, indicating the formation of a few V-type complexes. These V-type complexes may have formed because the CS prepared with a low concentration of the etherifying agent possessed a relatively low positive surface charge and thus exhibited weaker attraction for LA. Consequently, the proportion of the V-type complex formed was relatively low. When the concentration of the etherifying agent was further increased to 30%, the enthalpy value of the samples increased to 1.45 J/g (Fig. 4B), indicating that the proportion of the V-type structure in the CS–LA complexes prepared under this condition was the highest among all samples prepared with the etherifying agent. In addition, when the DS increased, the maximum weight loss rate of the sample measured by the thermogravimetric analyzer increased from 281.32 °C to 298.53 °C (Fig. 4C and D). This trend may be attributable to the progressively increasing positive surface charge of CS at higher etherifying agent concentrations, which enhanced the degree of electrostatic affinity toward the negatively charged LA. The resulting stronger interaction facilitated greater adsorption and more stable complex formation between amylose and LA, consistent with the XRD results (Qin et al., 2019). Together, these findings indicate that surface charge density plays a crucial role in modulating the complexation efficiency of CS with anionic lipid molecules.

Fig. 4.

Fig. 4

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Differential scanning calorimetry curve (A) and enthalpy change (B) and thermogravimetric analysis curves (C), and derivative thermogravimetric curves (D) of complexes formed between lauric acid and cationic starch synthesized under various etherifying agent contents.

3.5. Rheological properties of cationic starch-lauric acid complexes

To investigate the interaction between CS and LA and the effect of this interaction on the viscoelasticity of the system, this study systematically examined the dynamic oscillation and steady-state shear rheological behaviors of three samples (ETA-20%–LA, ETA-20%, and ETA-20%–LA–NaCl) (Fig. 5). Dynamic viscoelastic analysis (Fig. 5A and B) revealed that throughout the angular frequency scanning range (0.1–100 rad/s), the storage modulus (G') and loss modulus (G") of ETA-20%–LA were significantly higher than those of the other two groups, and G' was always greater than G" (G'/G" > 1), reflecting the typical characteristics of a weak gel. By contrast, the G' and G" values of the ETA-20% sample were substantially reduced, and the difference between G' and G" decreased with increasing frequency, indicating that CS alone was insufficient to construct a strong network structure. According to the literature, intermolecular chains primarily rely on limited hydrogen bonds or van der Waals forces to maintain a weak association (Lin et al., 2021). Notably, the G' and G" values of the ETA-20%–LA–NaCl sample were at their lowest throughout the frequency range, and they tend to overlap. This result indicated that the introduction of NaCl considerably disrupted the structure of the system. Steady-state shear rheological analysis (Fig. 5C) confirmed that the ETA-20%–LA sample exhibited the highest apparent viscosity and a marked shear-thinning behavior as the shear rate increased, a result attributable to the dissociation and rearrangement of the network structure under external force, conforming to the characteristics of a typical non-Newtonian pseudoplastic fluid. Moreover, the viscosity curve of the ETA-20% sample was centered, whereas the viscosity of the ETA-20%–LA–NaCl sample declined sharply to nearly the level of a Newtonian fluid and did not substantially change with the shear rate throughout the shear rate range, further confirming the collapse of its internal ordered structure (Chou et al., 2019). The macroscopic morphology data (Fig. 5D) were highly consistent with the rheological data: ETA-20%–LA formed a uniform, continuous, and cohesive gel-like system, whereas ETA-20%–LA–NaCl was a highly fluid and thinly dispersed solution with a morphology similar to that of the control group. Overall, the electrostatic interaction between CS and LA considerably strengthened the gel network structure of the system.

Fig. 5.

Fig. 5

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Rheological properties (storage modulus and angular frequency (A), loss modulus and angular frequency (B), and viscosity and shear rate (C)) and digital images of cationic starch and complexes formed between lauric acid and cationic starch.

3.6. Starch digestibility of cationic starch-lauric acid complexes

This study evaluated the in vitro digestibility of the CS–LA complexes prepared with varying etherifying agent concentrations. Fig. 6 presents the concentrations of RDS, slowly digestible starch, and RS. The unmodified complex (ETA-0%–LA) exhibited the highest RDS content (58.85%) and the lowest RS content (22.81%), indicating relatively poor resistance to enzymatic hydrolysis. This result may be attributable to the absence of cationic modification, which limited the electrostatic attraction between the starch and LA, thereby mitigating the formation of stable V-type inclusion complexes that typically hinder enzyme access. With the introduction of low concentrations of the etherifying agent (ETA-5%–LA and ETA-10%–LA), a slight reduction in RDS content (56.18.78% and 52.15%, respectively) and a slight change in RS content (21.78 and 25.75%, respectively) were observed relative to the control. This trend suggested that mild cationization altered the surface charge of starch, promoting superior interaction with LA and modestly enhancing the formation of enzyme-resistant structures. Notably, when the concentration of the etherifying agent dosage was increased to 15% and 20% (ETA-15%–LA and ETA-20%–LA), a substantial shift in the digestibility profile occurred. The RS content increased to 25.82% and 29.24%, respectively, whereas the RDS content decreased to 49.30% and 44.74%, respectively. This pronounced change indicated that intermediate levels of cationization substantially enhanced the formation of V-type helical complexes between amylose and LA, which considerably enhanced enzymatic resistance. Overall, the strong electrostatic interactions between the positively charged CS and the negatively charged LA likely promoted a more compact and ordered network structure that effectively shielded the starch granules from enzymatic attack (Qin et al., 2019). At the highest etherifying agent concentration (ETA-30%–LA), the RS content was 31.80% and the RDS content was 41.94%, indicating a balanced distribution between rapidly and slowly digestible fractions. Compared to ETA-20%-LA, the RS content of ETA-30%-LA decreased slightly despite having the highest V-type crystallinity. This is likely because the excessive degree of cationization imparted extreme hydrophilicity to the starch network (Kaur et al., 2012). The resulting massive swelling in the aqueous digestion buffer increased matrix porosity, thereby facilitating the diffusion and access of digestive enzymes to the amorphous regions (Dhital et al., 2017). Overall, these results demonstrate that EVGS was readily destroyed in the aqueous system. After modification, the digestibility of the CS–LA complexes increased and became highly dependent on the degree of cationization. Furthermore, moderate cationization substantially increased the RS content and reinforced structural stability, an effect attributable to electrostatic-interaction-driven network formation (Fig. 7). Specifically, the RS content increased in a dose-dependent manner with increasing cationization degree, accompanied by progressive enhancements in key structural parameters, including melting enthalpy and crystalline region integrity, suggesting a direct positive correlation between charge density and structural robustness.

Fig. 6.

Fig. 6

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Digestible components of complexes formed between lauric acid and cationic starch synthesized under various etherifying agent contents.

Fig. 7.

Fig. 7

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Schematic illustration of electrostatic interaction enhanced the stability of V-type complexes formed between lauric acid and cationic starch.

4. Conclusion

This study successfully developed a highly stable and highly digestive-resistant V-type starch through an electrostatic interaction between CS and LA. The zeta potential, 1H NMR analysis, and DS results confirmed the successful introduction of cationic groups into the starch matrix. When the concentration of the etherifying agent was increased, the V-type crystallinity of the prepared CS–LA complexes substantially increased. Analysis of the complexes' in vitro digestive properties verified that the digestibility of the prepared complexes strongly depended on the degree of cationization. Notably, moderate cationization markedly increased the RS content and decreased the RDS content in the prepared complexes. Rheological analysis further confirmed that the electrostatic interaction between CS and LA substantially increased the structural stability of the prepared complexes. In summary, these results suggest that electrostatic interaction can be used to establish a novel and versatile platform for modulating the structural stability and digestive resistance of V-type starch. A limitation of the current study is the reliance on EPTAC-modified starch, which has strict regulatory limits for food application. Future research should focus on utilizing natural, food-grade polycations (such as chitosan) to achieve similar electrostatic-driven V-type complexation, and conduct comprehensive in vivo safety and metabolic assessments.

CRediT authorship contribution statement

Zhenfei Ding: Writing – review & editing, Writing – original draft, Methodology, Funding acquisition, Formal analysis, Data curation. Wenjing Li: Validation, Methodology. Chao Xu: Formal analysis. Yuanhang Wang: Validation, Methodology. Xu Chen: Writing – review & editing, Funding acquisition, Formal analysis, Conceptualization.

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgments

This project was kindly funded by the Research Foundation of Education Bureau of Anhui Province, China (Grant No.2023AH051914), the compilation project of scientific research plan of Anhui Provincial Education Department (Grant No.2024AH010020), and the Anhui Agricultural University Foundation for Stability and Introduction of Talent (rc352301).

Data availability

Data will be made available on request.

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