Effect of Industrial-Scale Microfluidizer Treatment on the Physicochemical Properties and Quality of Whole-Component Dehulled Foxtail Millet Slurry

Abstract

The effects of an industrial-scale microfluidizer (ISM) on the physicochemical properties and quality of whole-component dehulled foxtail millet slurry were investigated under varying processing pressures (0, 60, 90, and 120 MPa). ISM treatment significantly enhanced the apparent stability of the whole-component dehulled foxtail millet slurry, with ISM–120 exhibiting the best apparent stability. The results of dispersion characteristics, serum cloudiness, and zeta potential measurements indicated that ISM processing enhanced the physical stability of the slurry. As processing pressure increased, the particle size of whole-component dehulled foxtail millet slurry first decreased sharply and then showed a slight increase. Compared to the untreated slurry, the D was reduced by approximately 81.32%, 81.72%, and 78.44% after treatment at 60, 90, and 120 MPa, respectively. Concurrently, the apparent viscosity of the slurry rises with increasing processing pressure, with ISM–120 displaying the highest apparent viscosity. Furthermore, CLSM analysis revealed that ISM–90 and ISM–120 exhibited overall more uniform and stable structures. The content of damaged starch correspondingly increased with higher processing pressures, further corroborating the findings from particle size and scanning electron microscopy observations. Simultaneously, the soluble solids content also increased with rising ISM processing pressure. However, increasing ISM processing pressure progressively reduced the L*, a*, b*, and C* values of the slurry, while the ΔE and h values progressively increased. Compared to the untreated slurry, the ΔE value increased by approximately 1.92%, 3.85%, and 6.41% after treatment at 60, 90, and 120 MPa, respectively. These changes resulted in a deterioration of the color quality of the whole-component dehulled foxtail millet slurry.

1. Introduction

Foxtail millet (Setaria italica (L.) P. Beau.) grain consists of the husk bran, germ, and endosperm. Dehulled foxtail millet, obtained by removing only the husk, is a whole-grain product. In contrast, polished millet—the most commonly consumed form—is a refined product resulting from the further removal of the bran and germ, leaving primarily the endosperm. Compared to polished foxtail millet, the dehulled counterpart retains higher levels of nutrients such as fat, protein, and dietary fiber, as well as a richer profile of phytochemicals including polyphenolic compounds and carotenoids [1,2]. These phytochemicals can act as natural modulators of inflammatory markers like adipokines and cytokines [3]. Furthermore, whole-grain diets can reduce the risk of chronic diseases [4]. Therefore, developing innovative whole-grain foods from dehulled foxtail millet is pursued to reduce processing waste, enhance nutritional value, and meet consumer demand for health-promoting foods.

Grain-based beverages are an ideal vehicle to deliver these nutritional benefits and have seen rapidly growing market interest as sustainable, plant-based alternatives [5]. However, processing dehulled foxtail millet into palatable and stable beverages poses distinct technological challenges. Its high fiber and hardness render conventional size-reduction technologies (e.g., colloid mills and homogenizers) inefficient and energy-intensive [6], often failing to effectively disrupt the bran layer. Consequently, the resulting beverages frequently suffer from a coarse texture, rapid sedimentation, and poor stability—key factors limiting consumer acceptance. Polishing the grain to ease processing sacrifices its nutritional premium and value-added potential [7]. Critically, the lack of novel, efficient processing technologies specifically designed for this challenging whole-grain matrix remains a major bottleneck, which limits the development and commercialization of high-value dehulled foxtail millet products.

Dynamic high-pressure microfluidization (DHPM) is a novel non-thermal technology that utilizes intense shear, cavitation, and impact forces to modify food microstructure and has shown promise in dairy and emulsion systems [8]. However, its application to fibrous plant materials is often limited in food processing due to its small flow channels (which are prone to blockage) and low processing capacity [9]. The industrial-scale microfluidizer (ISM) is an engineered development of DHPM designed to overcome these limitations. The ISM primarily relies on triplex high-pressure pumps and microfluidic chamber nozzles to achieve an exceptional ultrafine grinding of materials. It features an optimized microchannel design compared to traditional models, enabling effective processing of high-fiber food resources while preventing blockages, and offers a substantially higher throughput suitable for industrial-scale processing [10]. Intense mechanical action within the ISM is expected to degrade macromolecular polymers and alter their physicochemical properties.

Previous investigations have demonstrated the efficacy of ISM in modifying food macromolecules and enhancing the properties of whole-component slurries. For instance, He et al. [11] reported that ISM treatment at 120 MPa increased pea protein solubility from 16.99% to 64.28%, underscoring its potential for developing pea protein-based foods. In another study, He et al. [12] demonstrated that ISM treatment effectively modified potato starch, leading to structural disorganization and altered thermal, pasting, and rheological properties. Li et al. [13] found that whole-bean soymilk treated with ISM at 60–120 MPa exhibited better stability during 21 days of storage at 4 °C compared to untreated controls. Guo et al. [14] utilized ISM to prepare whole-component corn slurry, observing improvements in both physical stability and nutritional value with increasing pressure. Furthermore, Dai et al. [10] showed that ISM effectively enhanced the physical stability and volatile flavor profile of whole peanut milk, with the sample processed at 120 MPa remaining stable without delamination for 67 h at 4 °C. Collectively, these findings highlight the potential of ISM to address common stability issues in plant-based systems. However, its application has not been extended to dehulled foxtail millet—a matrix with unique compositional and structural challenges, as outlined above. Additionally, previous studies have primarily focused on beverages made from high-protein legumes or on modifying single or enriched components of cereals, whereas the subject of this study, dehulled foxtail millet, represents a complex mixed system. Investigating ISM for this specific grain is therefore essential to reveal the effects of ISM in multi-component coexisting systems and unlock its potential for creating stable, whole-grain beverages.

Therefore, this study utilized dehulled foxtail millet as the raw material. The millets were pre-ground using a wet grinder and a medium-pressure jet mill, and the resulting slurry was processed using an ISM at pressures of 0, 60, 90, and 120 MPa. The aim was to systematically investigate the effects of ISM pressure on key physicochemical properties (e.g., particle size, rheology, stability) and quality attributes of the whole-component slurry. This work is expected to elucidate the processing property relationships induced by ISM and evaluate its potential as an innovative solution for the industrial production of high-quality, value-added dehulled foxtail millet beverages.

2. Materials and Methods

2.1. Materials

Dehulled foxtail millet was obtained from Zhangjiakou Xuntian Food Co., Ltd. (Zhangjiakou, China). Fluorescein isothiocyanate (FITC), dimethyl sulfoxide (DMSO), and Rhodamine B were purchased from Shanghai Yuanye Bio–Technology Co., Ltd. (Shanghai, China). Nile Red was purchased from Shanghai Macklin Biochemical Technology Co., Ltd. (Shanghai, China). All other reagents and chemicals were of analytical grade.

2.2. Preparation of Whole-Component Dehulled Foxtail Millet Slurry

The dehulled foxtail millet was first mixed with distilled water at a 1:9 (w/v) ratio and pre-ground once using a wet grinder (FG–1018, Beijing Collaborative Innovation Food Technology Co., Ltd., Beijing, China). The frequencies of the wet grinder were set at 40 Hz, and 27.5 and 25 Hz for the screw feeder and water pump, respectively, with discharge temperatures of 27.5 °C. Subsequently, the slurry was transferred to a medium-pressure jet mill (FL–3030, Beijing Collaborative Innovation Food Technology Co., Ltd., Beijing, China), which operated at a frequency of 41 Hz, with a discharge temperature of 35 °C. The resultant slurry was further processed using an industrial-scale microfluidizer (FJ–3037S, Beijing Collaborative Innovation Food Technology Co., Ltd., Beijing, China) at pressures of 0, 60, 90, and 120 MPa. The slurry processing frequencies at 60, 90, and 120 MPa were 23.5, 30, and 36 Hz, with discharge temperatures of 41, 51, and 59 °C, respectively. Samples were processed continuously in the wet mill, medium-pressure jet mill, and ISM with minimal residence time. The final whole-component slurries were designated as ISM–0, ISM–60, ISM–90, and ISM–120, corresponding to the microfluidization pressure applied. All slurries used for observing apparent stability were preserved with 0.05% (w/w) Proclin to inhibit microbial growth.

2.3. Apparent Stability

Equal volumes of the prepared slurries were dispensed into transparent glass bottles and stored at 4 °C. The apparent stability of the samples was assessed and imaged at 0 h, 2 h, 8 h, and 24 h, respectively.

2.4. Dispersion Characteristics

The dispersion characteristics of the samples were determined using a concentrated system dispersion stability analyzer (LUMiSizer^®, LUMiSizer GmbH, Berlin, Germany), according to the method described by Primoz et al. [15] with minor modifications. After homogenization, the sample was transferred into the sample tube until the calibration line was reached. Scanning was performed at 2500 rpm and 25 °C at 10 s intervals for a total of 255 scans. The scan results were automatically recorded by the instrument, followed by calculation of the sample instability index and light transmittance finger pattern.

2.5. Serum Cloudiness

The serum cloudiness of the samples was determined according to the method of Lv et al. [16] with slight modifications. Briefly, the aliquot of each sample was centrifuged at 1600× g for 15 min. The absorbance of the resultant supernatant was then measured at 660 nm, with deionized water serving as the blank.

2.6. Zeta Potential

The zeta potential of the samples was determined using a Zetasizer Nano ZS90 (Malvern Instruments Ltd., Worcestershire, UK), following the method of Mocanu et al. [17] with minor modifications. Prior to analysis, each sample was diluted 20 times with distilled water. Measurements were conducted in disposable folded capillary cells at 25 °C, with an equilibration time of 120 s.

2.7. Particle Size

The particle size distribution of the samples was determined using a laser diffraction particle size analyzer (Mastersizer 3000, Malvern Instruments Co., Ltd., Worcestershire, UK), based on the method of Arapi et al. [18] with minor modifications. The parameters were set as follows: particle refractive index 1.52, particle absorption coefficient 0.100, and deionized water as the dispersant (refractive index of 1.330). The width of the particle size distribution was expressed as the span value, calculated using the following formula:

$$Span={\displaystyle \frac{D\left(90\right)- D\left(10\right)}{D\left(50\right)}}$$

(1)

where D(10), D(50), and D(90) denote the particle sizes at 10%, 50%, and 90% of the cumulative volume distribution, respectively.

2.8. Rheological Properties

The rheological properties of the samples were analyzed using a rheometer (AR 2000, TA Instruments, Newcastle, DE, USA), following a procedure adapted from Malafronte et al. [19]. A 16 mL aliquot of each sample was loaded into the concentric cylinder. Apparent viscosity was recorded as a function of shear rate from 0.1 s⁻¹ to 100 s⁻¹ at 25 °C. The rheological behavior of the sample was analyzed using the Ostwald–de Weale (Power law) model:

$$\tau = {k\gamma }^n$$

(2)

where τ is the shear stress (Pa), k is the consistency coefficient (Pa·sⁿ), γ is the shear rate (s⁻¹), and n is the flow behavior index (n = 1 for Newtonian fluids and n < 1 for non-Newtonian fluids).

2.9. Scanning Electron Microscopy (SEM) Analysis

Freeze-dried powder samples were sprinkled on conductive double-sided adhesive tape attached to circular specimen stubs and then sputter-coated with a thin gold film [20]. The microstructure was observed using an SEM (JSM–IT 700HR, Japan Electronics Co., Ltd., Tokyo, Japan) at an accelerating voltage of 5.0 kV and a magnification of ×1000.

2.10. Confocal Laser Scanning Microscope (CLSM) Analysis

The microstructure of the slurries was analyzed using CLSM (STELLARIS 8, Leica, Wetzlar, Germany), according to the method described by Huang et al. [21] with minor modifications. Briefly, 2 mL of sample was stained with 200 μL of a mixed dye solution containing 0.25% (w/v) fluorescein isothiocyanate (FITC), 0.025% (w/v) Rhodamine B, and 0.1% (w/v) Nile Red in DMSO. Subsequently, 50 μL of the stained sample was placed on a microscope slide, and micrographs were acquired at ×40 magnifications. The laser emission wavelengths were set as follows: 488 nm for FITC, 555 nm for Rhodamine B, and 568 nm for Nile Red.

2.11. Damaged Starch and Soluble Solids Contents

2.11.1. Damaged Starch Content

The degree of starch granule damage was determined using the Megazyme starch damage assay kit (K–SDAM, Megazyme Ltd., Wicklow, Ireland), which is based on the AACC Method 76–31.01. The sample was freeze-dried before analysis, and the results were expressed on a dry weight basis. Partial procedures for measurement and calculation were adapted from the work of Asmeda et al. [22].

2.11.2. Soluble Solids Content

The soluble solids content of the samples was determined using a digital Abbe refractometer (WAY–3S, Shanghai INESA Physico-optical Instrument Co., Ltd., Shanghai, China), according to the method described by Strieder et al. [23]. The instrument was zero-calibrated with distilled water before the measurement. A few drops of each sample were applied to cover the sensing area, and the soluble solids content was read directly and expressed in °Brix, which represents the mass percentage of sucrose in the slurry. All measurements were performed at 20 °C.

2.12. Color

The color parameters of the samples were measured using a colorimeter (CS–800, Hangzhou Caipu Technology Co., Ltd., Zhejiang, China), the color parameters corresponding to the uniform color space CIELAB were obtained directly from the apparatus, following the method described by Zhang et al. [24] with minor modifications. A total of 10 mL of each sample was transferred into a measurement cuvette, and the L*, a*, and b* values were recorded. The measurements were conducted under D65 illuminant with a 10° standard observer. The total color difference (ΔE), chroma (C*), and hue angle (h values) were then calculated using the following formulas:

$$\Delta E=\sqrt{{\left(\Delta L*\right)}^2+ {\left(\Delta a*\right)}^2+ {\left(\Delta b*\right)}^{2 }}$$

(3)

$$C*=\sqrt{\left({a*}^2\right)+\left({b*}^2\right)}$$

(4)

$$h=arctan{\displaystyle \frac{b*}{a*}}$$

(5)

where L* represents lightness, a* represents redness/greenness, b* represents yellowness/blueness.

2.13. Statistical Analysis

All measurements were performed in triplicate, and the data are presented as means ± standard deviation (SD). Statistical analysis was conducted using IBM SPSS Statistics (version 26.0, IBM Corp., Armonk, NY, USA). Differences among groups were evaluated using one-way analysis of variance (ANOVA), and normality and homogeneity of variance were verified prior to ANOVA. When the ANOVA indicated a significant overall difference (p < 0.05), Duncan’s multiple range test (DMRT) was applied for post hoc pairwise comparisons. Furthermore, hierarchical cluster analysis (HCA) was performed based on ten selected physicochemical properties to assess sample similarity. Variables were standardized prior to clustering. During the clustering process, Euclidean distance was selected as the distance metric, and average linkage was employed for clustering. A dendrogram visualizing the clustering results was constructed using Origin (version 2024, OriginLab Co., Northampton, MA, USA).

3. Results and Discussion

3.1. Effects of Different ISM Pressures on Apparent Stability of Whole-Component Dehulled Foxtail Millet Slurry

The physical stability of the whole-component foxtail millet slurries showed a clear dependence on ISM treatment pressure, as visually assessed over 24 h (Figure 1). At 0 h, all slurries appeared homogeneous. Upon standing, distinct differences emerged. The untreated control (ISM–0) exhibited the most rapid and severe phase separation, with a clear supernatant layer observable by 2 h. This separation progressively intensified, resulting in a large serum layer and concentrated sediment by 24 h. In contrast, the slurry treated at 120 MPa (ISM–120) demonstrated the highest stability, showing the slowest phase separation and the least pronounced sediment layer after 24 h. The slurries treated at intermediate pressures displayed a graded response. ISM–60 showed noticeable separation by 2 h, which became more pronounced over time, whereas ISM–90 exhibited a slower onset and lesser degree of separation compared to ISM–60. In summary, the visual stability of the slurries was enhanced with increasing ISM treatment pressure, with ISM–120 showing the greatest resistance to sedimentation and phase separation. The enhanced physical stability observed visually is attributed to modifications in the slurry’s colloidal system. To establish this structure–property relationship, the key physicochemical properties governing stability—particle size distribution, zeta potential, and rheological behavior—were characterized, along with microstructural observations using SEM and CLSM and compositional analyses of changes, including microstructure, damaged starch and soluble solids contents.

3.2. Effects of Different ISM Pressures on Dispersion Characteristics of Whole-Component Dehulled Foxtail Millet Slurry

The whole-component dehulled foxtail millet slurry constitutes a complex colloidal system comprising suspended particulates of varying composition and size. During storage, density differences between these particulates and the continuous aqueous phase can lead to gravitational sedimentation and phase separation, thereby compromising product stability. A bulk dispersion stability analyzer was employed to evaluate physical stability, providing two key metrics: the instability index and change in light transmission. The instability index, calculated from the temporal variation in light transmission across different sample regions during centrifugation, serves as a direct indicator of stability, with lower values denoting higher system stability [25]. As presented in Figure 2, the untreated control slurry (ISM–0) exhibited the highest instability index, confirming its poorest stability. In contrast, ISM treatment progressively enhanced stability, with the instability index values for ISM–60, ISM–90, and ISM–120 samples reduced by 1%, 6%, and 8%, respectively. The evolution of phase separation was further elucidated by the light transmittance finger pattern (Figure 3). Initially, the transmittance for all samples approached zero, indicative of a uniformly opaque and well-dispersed system. During centrifugation, the less dense serum phase migrated upward under centrifugal force, while denser sediment accumulated at the bottom, leading to increased light transmittance over time [26]. The profiles, represented from the initial bottom scan (red) to the final top scan (green), revealed that, with increasing ISM pressure, the change in transmittance diminished, and the area between the transmission curves decreased. A smaller interfacial area between the curves signifies slower serum migration and closer sediment packing, which correlates with greater system stability [27]. This observed enhancement in stability with increasing ISM pressure can be primarily attributed to two synergistic factors: a reduction in particle size and an increase in apparent viscosity. Finer particles experience reduced gravitational settling forces, while a more viscous continuous phase retards particle movement, both of which collectively inhibit phase separation.

3.3. Effects of Different ISM Pressures on Serum Cloudiness of Whole-Component Dehulled Foxtail Millet Slurry

Serum cloudiness is another critical parameter for assessing the physical stability of suspension systems, where higher turbidity often correlates with better stability due to the prolonged suspension of fine particles [28]. As shown in Figure 4A, the serum cloudiness of the whole-component dehulled foxtail millet slurry exhibited a significant upward trend with increasing ISM pressure. Based on these observations, it is hypothesized that this phenomenon may result from the gradual increase of the absolute value in the Zeta potential of the slurry, gradually increasing as the ISM processing pressure rises. This leads to enhanced electrostatic repulsion between small particles in the supernatant after centrifugation [29], making it difficult for them to aggregate and settle, resulting in a marked increase in serum cloudiness.

3.4. Effects of Different ISM Pressures on Zeta Potential of Whole-Component Dehulled Foxtail Millet Slurry

The Zeta potential is widely used to characterize the degree of electrostatic repulsion between adjacent colloidal particles carrying similar charges in suspensions, and it is used to evaluate the potential stability of colloidal systems [29]. A higher absolute value of the Zeta potential is associated with stronger electrostatic repulsion forces between components in the solution, which leads to greater system stability [30]. As illustrated in Figure 4B, the absolute zeta potential value of the whole-component foxtail millet slurry increased progressively with rising ISM processing pressure. Notably, a charge reversal was observed. At processing pressures below 60 MPa, the Zeta potential values were positive, whereas all samples treated at pressures exceeding 60 MPa exhibited negative Zeta potentials. This may be attributed to the high pressure initially increasing the exposure of the slurry’s inherent positive charges. Subsequently, at higher processing pressures, the slurry structure was disrupted, exposing internal charges and hydrophilic groups, thereby imparting a highly negative charge to the slurry [31].

3.5. Effects of Different ISM Pressures on Particle Size and Distribution of Whole-Component Dehulled Foxtail Millet Slurry

According to Stokes’ law, the sedimentation velocity of particles is governed by their size and density [32]. Consequently, particle size and its distribution serve as critical indicators for evaluating the physical stability of beverage systems. The particle size characteristics of the whole-component dehulled foxtail millet slurries, including the volume-weighted mean diameter (D[4,3]), surface-weighted mean diameter (D[3,2]), and the cumulative percentiles D(10), D(50), and D(90), were determined using laser diffraction and were summarized in

Table 1. Effects of different ISM pressures on particle size of whole-component dehulled foxtail millet slurry.

Samples	D[3,2] (μm)	D[4,3] (μm)	D(10) (μm)	D(50) (μm)	D(90) (μm)	Span (μm)
ISM–0	8.41 ± 0.03 a	63.87 ± 0.37 a	4.27 ± 0.02 b	14.23 ± 0.05 a	198.00 ± 0.82 a	13.61 ± 0.07 a
ISM–60	5.31 ± 0.02 d	11.93 ± 0.05 c	2.33 ± 0.01 d	9.01 ± 0.04 d	17.77 ± 0.17 c	1.71 ± 0.01 b
ISM–90	5.95 ± 0.03 c	11.67 ± 0.12 c	3.35 ± 0.04 c	9.54 ± 0.05 c	18.33 ± 0.17 c	1.57 ± 0.01 c
ISM–120	7.23 ± 0.04 b	13.77 ± 0.12 b	4.83 ± 0.03 a	11.27 ± 0.05 b	22.53 ± 0.31 b	1.56 ± 0.01 c

Data are expressed as means ± standard deviation (n = 3). Different superscript letters in the same column indicate significant differences (p < 0.05). ISM–0, ISM–60, ISM–90, and ISM–120 indicate whole-component dehulled foxtail millet slurry prepared with ISM at pressures of 0, 60, 90, and 120 MPa, respectively. Different letters indicate significant differences (p < 0.05); same letters indicate no significant difference.

. The particle size indices exhibited a non-linear trend with increasing ISM treatment pressure, initially decreasing and then increasing. The untreated slurry (control) displayed a D[4,3] of 63.87 µm. In contrast, treatment at 60, 90, and 120 MPa significantly reduced this value to 11.93 µm, 11.67 µm, and 13.77 µm (p < 0.05), respectively. This initial reduction confirms that higher processing pressures generally enhance grinding efficiency, leading to particle size diminution. However, the relationship between pressure and final particle size is not monotonic. While particle size decreased progressively as pressure increased from 0 to 90 MPa, a subsequent increase was observed for the ISM–120 sample. Similarly, the values for D[3,2], D(10), D(50), and D(90) all decreased within the 0–60 MPa range but showed an upward trend beyond 60 MPa. The span value gradually decreased within the 0–90 MPa range but remained nearly unchanged above 90 MPa. This indicates that ISM processing effectively enhances the uniformity of whole-component dehulled foxtail millet slurry, though a higher pressure does not necessarily yield better results. The particle size distribution profiles, presented in Figure 5, further elucidate these trends. All slurries exhibited a multimodal distribution, indicative of the complex, heterogeneous nature of the whole-component system comprising starch, protein, and dietary fiber particles of varying sizes. As the processing pressure increased from 0 to 60 MPa, the distribution curves shifted leftwards, signifying the effective comminution of both large and small constituent particles [11,12,33]. However, from 60 MPa to 120 MPa, the peak of the particle size distribution became less distinct, likely due to the partial repolymerization of small particles under a higher pressure [34].

3.6. Effects of Different ISM Pressures on Rheological Properties of Whole-Component Dehulled Foxtail Millet Slurry

According to Stokes’ law, the higher viscosity of polydisperse systems is known to minimize droplet mobility and collision frequency, as well as reduce droplet flocculation, thereby enhancing system stability [35]. Consequently, viscosity is regarded as a physical barrier against stratification in whole-grain beverages, which makes rheological properties critical to beverage stability. As shown in Figure 6, the apparent viscosity of all slurry samples exhibited pronounced shear-thinning behavior with increasing shear rate, indicating that whole-component dehulled foxtail millet slurry is a typical pseudoplastic non-Newtonian fluid [36]. Furthermore, the strong shear forces, high-speed impacts, high-frequency vibrations, and thermal effects generated during ISM processing caused the apparent viscosity of the sample to gradually increase with rising processing pressure. This may be due to an increase in damaged starch or soluble solids content [37]. Therefore, the increase in system viscosity mitigates particle gravitational settling to a certain extent, thereby enhancing the overall stability of the whole-component dehulled foxtail millet slurry.

3.7. SEM Analysis of Freeze-Dried Powder from Millet Slurry Prepared at Different Pressures

The microstructure of freeze-dried powder from millet slurry was observed using a scanning electron microscope. Pronounced pressure-dependent changes in particle morphology were evident (Figure 7). The control sample (Figure 7A) exhibited relatively large, intact particles with smooth surfaces and distinct edges, characteristic of minimal structural disruption. At 60 MPa (Figure 7B), partial fragmentation occurred, yet the overall particle morphology and surface characteristics remained largely unchanged. When the processing pressure increased to 90 MPa (Figure 7C), the particle breakage became more severe. The microstructure was characterized by extensive fragmentation, alongside noticeable particle adhesion that resulted in the formation of aggregated, blocky structures. At the highest pressure of 120 MPa (Figure 7D), the microstructure was dominated by large, densely packed aggregates with fused boundaries, suggesting extensive structural reorganization. Particle breakage can be attributed to high pressure disrupting hydrogen bonds between starch molecules and reducing starch crystallinity [38]. Protein rupture was found to contribute to particle aggregation, with an increase in disulfide bonds and a decrease in total thiol content inducing pH shifts that further promoted protein aggregation [39].

3.8. CLSM Analysis of Whole-Component Dehulled Foxtail Millet Slurry Prepared at Different Pressures

The microstructure of the slurry samples was observed using CLSM. Starch was stained with FITC (blue), proteins with Rhodamine B (green), and oil bodies with Nile Red (red). A multicolor image was obtained by overlaying the three monochrome images. As ISM pressure increased, progressive microstructural reorganization was observed. Starch granules gradually formed aggregates (Figure 8(A1–D1)). In the control sample (ISM–0), proteins initially aggregated (Figure 8(A2)). With increasing pressure, proteins gradually became dispersed (Figure 8(B2–D2)). A similar trend was observed for oil bodies: their size decreased with rising pressure, and they became more closely associated with proteins, suggesting enhanced protein–lipid interactions (Figure 8(A3–D3)). Ultimately, at 120 MPa, oil body particles were finely dispersed throughout the system, which is known to contribute to emulsion stability [40]. Notably, at 90 MPa, the spatial distribution of all three components exhibited significant changes. Regions of low concentration appeared as dark areas, while high-concentration zones exhibited blurred boundaries (Figure 8(C1–C3)). In the corresponding overlay images (Figure 8(C4)), particle boundaries became less distinct and tended to merge as pressure reached 90 MPa, indicating the formation of an interpenetrating network structure within the system [41]. This observation is consistent with the aggregated structures seen in SEM images (Figure 7C,D). Collectively, these CLSM observations demonstrate that increasing ISM pressure reduces the average particle size of starch, proteins, and oil bodies, while promoting their interaction, permeation, and binding. These changes lead to a more uniform and stable microstructure in the ISM–90 and ISM–120 samples, providing a structural basis for the enhanced physical stability reported in Section 3.1, Section 3.2 and Section 3.3.

3.9. Effects of Different ISM Pressures on Damaged Starch and Soluble Solids Contents of Whole-Component Dehulled Foxtail Millet Slurry

During ISM processing, shear forces and impacts generated by media collisions may cause damage to starch granules in terms of morphology, crystallinity, and molecular structure [42]. Damaged starch is an important factor in starch-based products, as changes in particle size are correlated with variations in damaged starch content, and finer particles typically show higher levels of damaged starch [43]. As shown in Figure 9A, the damaged starch content in whole-component dehulled foxtail millet slurry increased with rising ISM processing pressure. At 120 MPa, the damaged starch content reached 15.35%, representing an approximately 6.80% increase compared to ISM–0. This result highlights the trade-off in damaged starch formation, suggesting that pressure needs to be balanced between particle size reduction and starch integrity. The soluble solids content is a crucial indicator in characterizing beverage quality. As shown in Figure 9B, the soluble solids content of whole-component dehulled foxtail millet slurry increased with rising processing pressure. The soluble solids content of ISM–120 reached 5.13°Brix, representing an increase of approximately 2.71°Brix compared to ISM–0. This is attributed to the fact that the intense mechanical force of ISM promotes the conversion of insoluble components in the slurries into soluble components. This results in an increased soluble solids content, which contributes to enhancing the stability of the whole-component dehulled foxtail millet slurry system.

3.10. Effects of Different ISM Pressures on Color of Whole-Component Dehulled Foxtail Millet Slurry

Color is considered a crucial indicator for evaluating beverage quality and is known to directly influence consumer preference. The ΔE value is used to assess whether color differences are perceptible: a ΔE value less than 0.5 indicates no significant difference, a ΔE value between 0.5 and 1.5 indicates a slight perceptible difference, a ΔE value between 1.5 and 3 indicates a noticeable difference, and a ΔE value greater than 3 indicates a clearly visible difference [44]. As shown in

Table 2. Effects of different ISM pressures on color of whole-component dehulled foxtail millet slurry.

Samples	L*	a*	b*	ΔE	C*	h
ISM–0	77.97 ± 0.02 a	−0.27 ± 0.01 a	19.63 ± 0.03 a	–	19.63 ± 0.03 a	−89.27 ± 0.04 d
ISM–60	75.33 ± 0.01 b	−0.46 ± 0.01 b	10.31 ± 0.01 b	9.70 ± 0.03 c	10.31 ± 0.01 b	−87.51 ± 0.07 c
ISM–90	73.46 ± 0.02 c	−0.55 ± 0.01 c	8.06 ± 0.01 c	12.42 ± 0.02 b	8.08 ± 0.01 c	−86.12 ± 0.09 b
ISM–120	73.32 ± 0.01 d	−0.85 ± 0.01 d	8.02 ± 0.02 c	12.53 ± 0.02 a	8.06 ± 0.01 c	−83.97 ± 0.09 a

Data are expressed as means ± standard deviation (n = 3). Different superscript letters in the same column indicate significant differences (p < 0.05). ISM–0, ISM–60, ISM–90, and ISM–120 indicate whole-component dehulled foxtail millet slurry prepared with ISM at pressures of 0, 60, 90, and 120 MPa, respectively. Different letters indicate significant differences (p < 0.05); same letters indicate no significant difference.

, after ISM processing, the L*, a*, b*, and C* values of the whole-component dehulled foxtail millet slurry were gradually reduced, with the b* values showing the most significant changes (p < 0.05). Meanwhile, the ΔE and h values were gradually increased. This indicates a reduction in product brightness, as well as a greater attenuation of red tones compared to yellow tones, with the overall difference becoming more pronounced. This may be attributed to the formation of smaller particles, which enhance sample transparency and consequently reduce its brightness. Under increased processing pressure in the ISM, carotenoids gradually dissolve and disperse. The resulting dilution of pigment concentration per unit volume or area ultimately leads to diminished color saturation.

3.11. HCA

To describe the differences in the physicochemical properties and quality of whole-component dehulled foxtail millet slurry processed under various ISM pressures, a heatmap analysis using HCA dendrograms was conducted on the selected ten indicators. The selection of indicators primarily focuses on key metrics potentially affected by varying ISM processing pressures. For highly correlated indicators, one or two representative ones are chosen. The HCA dendrogram was used to group all samples into three distinct clusters, as shown in Figure 10. High similarity was observed between ISM–90 and ISM–120, indicating only minor differences in their physicochemical properties and quality. Significantly higher levels of soluble solids content, damaged starch content, and serum cloudiness were observed in ISM–90 and ISM–120 compared to ISM–0 and ISM–60. Conversely, the values of L*, a*, b*, instability index, zeta potential, D[3,2], and D[4,3] were mostly significantly lower in ISM–90 and ISM–120 than in ISM–0 and ISM–60. Therefore, ISM slurries processed under different pressures were clustered into three tiers: low pressure (0–60 MPa), medium pressure (60–90 MPa), and high pressure (90–120 MPa). This demonstrates that the effect of ISM processing pressure on sample properties exhibits continuity and regularity.

Although this study offers valuable insights for the development of whole-grain beverages, several limitations should be acknowledged. Owing to the inherent complexity of the slurry system, certain mechanistic aspects—such as protein conformational changes, disulfide bond formation, and pH shifts—were not directly measured. Consequently, the proposed explanations remain partially hypothetical. Future research should focus on elucidating these structural and physicochemical changes under varying ISM pressure treatments to validate the underlying mechanisms and further support the findings.

4. Conclusions

The effects of different processing pressures (0, 60, 90, and 120 MPa) using the ISM on the physicochemical properties and quality of whole-component dehulled foxtail millet slurry were investigated. ISM treatment significantly enhanced the apparent stability of the slurry, with the stability increasing as the treatment pressure rose. Measurements of dispersion characteristics and serum cloudiness revealed that the instability index was reduced, variation in light transmittance was minimized, and a gradual increase in serum cloudiness was observed following ISM treatment, all of which indicated a significant improvement in the physical stability of the slurry. As the ISM pressure increased, the particle size of the dehulled foxtail millet exhibited an initial sharp decrease, followed by a slight increase. Both the zeta potential and the apparent viscosity of the slurry increased with rising ISM pressure. Furthermore, CLSM imaging revealed that the microstructures of samples treated at 90 and 120 MPa were more uniform and stable overall. The increase in damaged starch content was consistent with the trends observed in particle size analysis and SEM. The soluble solids content also increased with higher ISM processing pressures. However, the color quality of the whole-component slurry was slightly reduced at higher ISM pressures. In conclusion, this study demonstrates that controlling the ISM processing pressure can effectively enhance the stability of the whole-component dehulled foxtail millet slurry system, providing a theoretical basis for the development of dehulled foxtail millet beverages.