Process for high-frequency pulse-assisted extraction of loquat seed polysaccharides

Abstract

This application discloses a process for high-frequency pulse-assisted extraction of loquat seed polysaccharides, comprising the following steps: S1, pretreatment: drying, pulverizing, and defatting loquat seeds to obtain loquat seed powder; S2, high-frequency pulse-assisted extraction: mixing the loquat seed powder from step S1 with an extraction solvent at a material-to-liquid ratio of 1:20 to 1:30 g / mL, and extracting at an extraction temperature of 80 to 95°C for 2.0 to 2.5 hours using high-frequency pulse assistance to obtain an extract, wherein the number of high-frequency pulses is 25 to 35; S3, post-treatment: performing solid-liquid separation, purification, and drying on the extract obtained in step S2 to obtain loquat seed polysaccharides. This application, by employing high-frequency pulse-assisted hot water extraction with specific parameters, can efficiently and gently disrupt the cell wall structure of loquat seeds, significantly promoting polysaccharide dissolution. Compared with traditional hot water extraction, the process of this invention has a shorter processing time, lower temperature, and higher polysaccharide yield. Furthermore, the obtained polysaccharides have good antioxidant activity, providing a green new technology for the high-value utilization of loquat processing by-products.

Description

Technical Field

[0001] This invention relates to the field of polysaccharide extraction technology, specifically to a process for high-frequency pulse-assisted extraction of loquat seed polysaccharides. Background Technology

[0002] Loquat (Eriobotrya japonica), a fruit with both medicinal and edible uses in my country, is popular among consumers for its lung-moistening and cough-relieving effects. With the rapid development of the loquat processing industry (such as loquat paste, loquat beverages, and canned goods), a large amount of processing by-products have been generated, of which loquat seeds (kernels) account for approximately 15%-20% of the fruit's fresh weight. Currently, most of these loquat seeds are disposed of as waste through landfills or incineration, resulting in a huge waste of biomass resources and placing a heavy burden on the environment. Research shows that loquat seeds are rich in polysaccharides, starch, oils, and other active ingredients with high reuse value. In particular, loquat seed polysaccharides have been preliminarily confirmed to possess potential biological activities such as in vitro antioxidant and antibacterial properties.

[0003] Traditional polysaccharide extraction methods mainly rely on hot water extraction. The principle is to use high temperature to accelerate molecular motion, allowing polysaccharides inside the cells to diffuse into the solvent. Although this method is simple, easy to operate, and inexpensive, the cell wall structure of loquat seeds is dense, mainly composed of cellulose, hemicellulose, and pectin. This structure poses a strong mass transfer resistance to the dissolution of polysaccharides inside the cells. Traditional hot water extraction methods cannot effectively destroy this dense structure, resulting in generally low polysaccharide extraction efficiency.

[0004] To address these issues, attempts have been made to introduce external physical fields for assisted extraction. For example, ultrasound-assisted extraction utilizes the shock waves and microjets generated by cavitation to break down cell walls, thus shortening the extraction time; microwave-assisted extraction uses the heat generated by the high-speed rotation and friction of polar molecules in a high-frequency electromagnetic field to achieve rapid heating. However, prolonged ultrasound treatment may lead to the destruction of polysaccharide structures by free radicals; localized overheating of microwaves poses a risk of damaging heat-sensitive active ingredients, and the equipment cost is relatively high.

[0005] High-frequency pulse-assisted extraction (HPE) technology utilizes the release of high-energy pulses over an extremely short time to generate intense instantaneous shock waves, ultraviolet light, and localized high-temperature effects. This combined physical action efficiently penetrates and breaks down plant cell walls, forming instantaneous microporous channels on the cell membrane. This significantly reduces the mass transfer resistance of the cell wall, allowing intracellular polysaccharides to dissolve rapidly under relatively mild conditions. However, there are currently no reports on the optimization of extraction process parameters for HPE technology in loquat seed polysaccharides, nor on the comprehensive research on the effects of this technology on the yield, antioxidant activity, and structural characteristics of the obtained loquat seed polysaccharides. Summary of the Invention

[0006] The present invention aims to provide a process for high-frequency pulse-assisted extraction of loquat seed polysaccharides. By using high-frequency pulse-assisted hot water extraction with specific parameters, the cell wall structure of loquat seeds can be destroyed efficiently and gently, thereby promoting the dissolution of polysaccharides.

[0007] To achieve the above objectives, the first aspect of this application provides the following technical solution: A high-frequency pulse-assisted extraction process for loquat seed polysaccharides includes the following steps: S1. Pretreatment: Loquat seeds are dried, pulverized, and degreased to obtain loquat seed powder; S2. High-frequency pulse-assisted extraction: The loquat seed powder from step S1 is mixed with the extraction solvent at a material-to-liquid ratio of 1:20 to 1:30 g / mL. The mixture is then extracted with high-frequency pulse assistance at an extraction temperature of 80 to 95°C for 2.0 to 2.5 hours to obtain the extract. The number of pulses in the high-frequency pulse is 25 to 35. S3. Post-processing: The extract obtained in step S2 is subjected to solid-liquid separation, purification, and drying to obtain loquat seed polysaccharide.

[0008] Furthermore, the material-to-liquid ratio in step S2 is 1:23 to 1:27 g / mL.

[0009] Furthermore, the material-to-liquid ratio in step S2 is 1:25 g / mL.

[0010] Furthermore, the extraction temperature is 87°C.

[0011] Furthermore, the extraction time is 2.3 hours.

[0012] Furthermore, the high-frequency pulse in step S2 has 30 pulses.

[0013] The second aspect of this application is loquat seed polysaccharide obtained based on the loquat seed polysaccharide extraction process of the first aspect.

[0014] A third aspect of this application provides the use of loquat seed polysaccharide in the preparation of antioxidants or functional foods.

[0015] Working principle and beneficial effects of the present invention: The instantaneous, intense shockwave generated by the pulse acts on the loquat seed cell wall in the form of mechanical stress, producing a micro-explosion-like effect. This causes the cell wall skeletal structure to rupture and loosen, thereby directly eliminating the main physical barrier to polysaccharide release. The pulse energy induces the generation of numerous microbubbles in the liquid medium. These bubbles grow, oscillate, and collapse violently in an instant, releasing localized high temperature, high pressure, and microjets, which intensifies the tearing effect on the cell wall and promotes the penetration of the extraction solvent into the cell interior.

[0016] This application also combines high-frequency pulse extraction with hot water extraction at 80–95℃. On the one hand, the hot water itself provides the basic thermal energy, accelerating molecular diffusion; on the other hand, the intervention of pulses allows for higher extraction efficiency at lower temperatures and in shorter times compared to traditional hot water extraction, effectively avoiding the degradation of polysaccharides caused by prolonged high-temperature cooking, thus protecting the natural structure and bioactivity of the polysaccharides. By optimizing key parameters (material-to-liquid ratio 1:25, temperature 87℃, time 2.3h, pulses 30 times), the yield of loquat seed polysaccharides can be stably reached above 19.36%, significantly higher than that of traditional hot water extraction, indicating that this method can fully release intracellular polysaccharides.

[0017] The obtained polysaccharide is a typical α-pyranopolysaccharide with a wide molecular weight distribution (weight average molecular weight 406.729 kDa). It mainly contains eight monosaccharides, including glucose (80.50%), galactose, and arabinose. Its microstructure is a porous honeycomb network, which can be used as a high-quality raw material for functional foods or pharmaceutical products. Attached Figure Description

[0018] Figure 1 This is a graph showing the effect of the material-to-liquid ratio on the extraction rate of loquat seed polysaccharides in the process described in this application. Figure 2 The graph shows the effect of extraction time on the extraction rate of loquat seed polysaccharides in the process described in this application. Figure 3 The figure shows the effect of extraction temperature on the extraction rate of loquat seed polysaccharides in the process described in this application. Figure 4 The graph shows the effect of the number of high-frequency pulses on the extraction rate of loquat seed polysaccharides in the process of this application. Figure 5 The ability of loquat seed polysaccharide to scavenge DPPH free radicals; Figure 6 The ability of loquat seed polysaccharides to scavenge ABTS free radicals; Figure 7 The ability of loquat seed polysaccharides to scavenge hydroxyl free radicals; Figure 8 Infrared spectrum of loquat seed polysaccharide; Figure 9 This is a scanning electron microscope image of loquat seed polysaccharides. Detailed Implementation

[0019] The following detailed description illustrates the specific implementation method: Example 1: Effect of different material-to-liquid ratios on extraction efficiency Weigh 0.2g of defatted loquat seed powder and add distilled water at material-to-liquid ratios of 1:20, 1:25, 1:30, 1:35, and 1:40 (g / mL). Extraction is carried out at an extraction temperature of 80℃, an extraction time of 2h, and 30 pulses to investigate the effect of material-to-liquid ratio on polysaccharide yield.

[0020] like Figure 1 As shown, the polysaccharide extraction rate reaches its highest value when the solid-liquid ratio is 1:25. When the solid-liquid ratio is too low (e.g., 1:20), the solvent volume is insufficient, the mass transfer driving force is weak, and the polysaccharides are not fully dissolved. When the solid-liquid ratio is too high (e.g., 1:40), although the solvent increases, the number of cells per unit volume decreases, and the energy consumption for subsequent concentration increases, resulting in a slight decrease in the extraction rate. Therefore, the suitable solid-liquid ratio range is 1:20-1:30, with the optimal point being approximately 1:25.

[0021] Example 2: Effect of different extraction times on extraction rate like Figure 2 As shown, 0.2g of defatted loquat seed powder was weighed and added to distilled water at a material-to-liquid ratio of 1:25. The extraction temperature was 80℃ and the number of pulses was 30. Extraction was carried out for 1.0h, 1.5h, 2.0h, 2.5h and 3.0h respectively to investigate the effect of extraction time on polysaccharide yield.

[0022] like Figure 2 As shown, within 2.5 hours, the polysaccharide extraction rate increases significantly with increasing extraction time. The extraction rate reaches its maximum at 2.5 hours; further extending the extraction time to 3.0 hours results in a decrease in the extraction rate. Therefore, the suitable extraction time range is 2.0-2.5 hours.

[0023] Example 3: Effect of different extraction temperatures on extraction rate Weigh 0.2g of defatted loquat seed powder, add distilled water at a material-to-liquid ratio of 1:25, extract for 2 hours, pulse 30 times, and extract at 70℃, 75℃, 80℃, 85℃ and 90℃ respectively to investigate the effect of extraction temperature on polysaccharide yield.

[0024] like Figure 3 As shown, within the temperature range of 70-85℃, the polysaccharide extraction rate increases significantly with increasing temperature. When the temperature reaches 85℃, the extraction rate is already at a relatively high level; further increasing the temperature to 90℃ results in a slight but minimal increase in the extraction rate. Higher temperatures enhance molecular thermal motion, reduce solvent viscosity, and increase mass transfer rate, while the synergistic effect of pulse-assisted extraction is also more pronounced. However, excessively high temperatures (above 90℃) lead to increased energy consumption and damage to heat-sensitive components. Therefore, the suitable extraction temperature range is 80-90℃, with the optimal point being approximately 87℃.

[0025] Example 4: Effect of different high-frequency pulse numbers on extraction rate Weigh 0.2g of defatted loquat seed powder, add distilled water at a material-to-liquid ratio of 1:25, extract at 85℃ for 2 hours, and set the pulse number to 15, 20, 25, 30 and 35 times respectively to investigate the effect of pulse number on polysaccharide yield.

[0026] like Figure 4 As shown, the polysaccharide extraction rate continuously increased from 15 to 30 pulses, reaching its maximum at 30 pulses. However, when the number of pulses continued to increase to 35, the extraction rate decreased. The reason for this is that moderate pulse treatment can effectively disrupt the cell wall structure, form microporous channels, and promote polysaccharide dissolution; however, excessive pulses can lead to excessive mechanical impact and localized instantaneous high temperatures, potentially causing polysaccharide molecular chain breakage or structural damage, thus reducing the effective yield. Therefore, the suitable range for the number of pulses is 25-35, with the optimal point being approximately 30 pulses.

[0027] Example 5: Response Surface Methodology for Optimizing Interaction Analysis Based on the single-factor experiments, a four-factor, three-level response surface methodology was conducted using a Box-Behnken design with the material-to-liquid ratio (A), extraction time (B), extraction temperature (C), and number of pulses (D) as independent variables and the loquat seed polysaccharide extraction rate as the response value. Regression analysis was performed on the response surface experimental results using Design-Expert software, yielding a quadratic polynomial regression model. Analysis of variance showed that the model p-value was <0.0001, the lack-of-fit term p=0.1608>0.05, and the model coefficient of determination R0.05 was [not specified]. 2 =0.9502, signal-to-noise ratio 14.7162, indicating that the model fits well.

[0028] The optimal extraction conditions for loquat seed polysaccharides, predicted by the model, were: a material-to-liquid ratio of 1:25.23, an extraction time of 2.29 h, an extraction temperature of 87.47 ℃, and 29.7 pulses, with a predicted extraction rate of 20.228%. For ease of practical operation, the process parameters were modified to: a material-to-liquid ratio of 1:25 g / mL, an extraction time of 2.3 h, an extraction temperature of 87 ℃, and 30 pulses.

[0029] Example 6: Verification of the antioxidant activity of loquat seed polysaccharides Loquat seed polysaccharides prepared under the optimal conditions in Example 5 were prepared into gradient solutions of 1, 2, 4, 6, 8, and 10 mg / mL, and the scavenging rates of DPPH free radicals, ABTS cation free radicals, and hydroxyl free radicals were determined respectively.

[0030] like Figure 5As shown, when the concentration of loquat seed polysaccharide is between 1-10 mg / ml, the scavenging ability of DPPH free radicals increases significantly. When the concentration reaches 10 mg / ml, the DPPH free radical scavenging rate reaches 88.2%, and the IC50 value is 1.615.

[0031] from Figure 6 As shown, when the concentration of loquat seed polysaccharide is between 1 and 6 mg / ml, the scavenging ability of ABTS free radicals increases with the increase of polysaccharide concentration. The scavenging rate no longer increases when the polysaccharide concentration is between 6 and 10 mg / ml, but tends to level off. It reaches 82.8% at 10 mg / ml with an IC50 value of 2.021.

[0032] like Figure 7 As shown, when the concentration of loquat seed polysaccharide is between 1 and 10 mg / ml, its ability to scavenge hydroxyl radicals increases with increasing concentration, reaching a maximum of 93.2% at 10 mg / ml, with an IC50 value of 2.16.

[0033] The above data proves that the loquat seed polysaccharide extracted by the process of this invention has excellent in vitro antioxidant activity and can be used as a natural antioxidant in functional foods, health products or cosmetics.

[0034] Example 7: Polysaccharide structural characterization (1) Infrared scanning determination of polysaccharides Infrared spectrum of loquat seed polysaccharide as follows Figure 8 As shown, at 3425.06cm -1 The absorption peak at 2927.44 cm⁻¹ is the stretching vibration absorption peak of -OH, which is a characteristic peak of carbohydrates; the absorption peak at 2927.44 cm⁻¹ belongs to the CH stretching vibration. At 1617.92, 1419.1 and 1242.69 cm -1 The absorption peaks at these locations are correlated with the characteristic vibrations of C=O, CH4, and CO, respectively, in the range of 1200-1000 cm⁻¹. -1 The regions show measurements of 1155.88, 1080.43, and 1020.58 cm. -1 Three typical absorption peaks are observed, with the strongest intensity at 1020 cm⁻¹, representing the stretching vibrations of the COC and COH bonds in the pyranose ring, indicating the presence of pyranose unit structures. The peak intensity is at 846.09 cm⁻¹. -1 The presence of an absorption peak indicates that loquat seed polysaccharide contains α-type pyranose polysaccharide.

[0035] (2) Determination of polysaccharide molecular weight distribution Table 1 - Molecular weight distribution of loquat seed polysaccharides

[0036] As shown in Table 1, loquat seed polysaccharide begins to elute at around 20 min with a mass fraction of 100%, indicating that there is only one main peak, but the peak shape should be very broad. The weight-average molecular weight of loquat seed polysaccharide is Mw=406.729 kDa, the peak molecular weight is Mp=3.52 kDa, and the number-average molecular weight is Mn=19.123 kDa. The molecular weight distribution of the loquat seed polysaccharide sample is extremely broad, containing a large number of low molecular weight components as well as some high molecular weight or aggregated components.

[0037] (3) Determination of polysaccharide molecular weight distribution Table 2 - Monosaccharide composition of loquat seed polysaccharides

[0038] As shown in Table 2, the monosaccharide composition of loquat seed polysaccharides includes arabinose, rhamnose, galactose, glucose, xylose, mannose, galacturonic acid, and glucuronic acid. Among them, glucose has the highest content, accounting for 80.5%, and is the main monosaccharide component.

[0039] (4) Scanning electron microscopy analysis The results of scanning electron microscopy are as follows Figure 9 As shown, at 30× magnification, the loquat seed polysaccharide sample exhibits a discrete, amorphous aggregate state, without large-area, dense clusters or regular crystal structures, consistent with the typical characteristics of amorphous polymers of plant-derived polysaccharides. At 1000× magnification, the polysaccharide core region reveals a porous honeycomb network structure assembled from numerous near-spherical microparticles. The particles are relatively densely arranged, forming an irregular sheet-like body. The edges of the polysaccharide body exhibit irregular serrated and dendritic extensions, with some areas forming tubular, rod-shaped, and free protrusion structures. Furthermore, the surface of the polysaccharide particles is generally smooth, without sharp edges.

[0040] It should be noted that those skilled in the art can make various modifications and improvements without departing from the technical solution of this invention. These modifications and improvements should also be considered within the scope of protection of this invention, and will not affect the effectiveness of the invention or the practicality of the patent. The scope of protection claimed in this application shall be determined by the content of its claims, and the specific embodiments described in the specification can be used to interpret the content of the claims.

Claims

1. A process for high frequency pulse assisted extraction of Loquat seed polysaccharides, characterized by, Includes the following steps: S1. Pretreatment: Loquat seeds are dried, pulverized, and degreased to obtain loquat seed powder; S2. High-frequency pulse-assisted extraction: The loquat seed powder from step S1 is mixed with the extraction solvent at a material-to-liquid ratio of 1:20 to 1:30 g / mL. The mixture is then extracted with high-frequency pulse assistance at an extraction temperature of 80 to 95°C for 2.0 to 2.5 hours to obtain the extract. The number of pulses in the high-frequency pulse is 25 to 35. S3. Post-processing: The extract obtained in step S2 is subjected to solid-liquid separation, purification, and drying to obtain loquat seed polysaccharide.

2. The process according to claim 1, characterized in that, The material-to-liquid ratio in step S2 is 1:23 to 1:27 g / mL.

3. The process according to claim 2, characterized in that, The material-to-liquid ratio in step S2 is 1:25 g / mL.

4. The process of claim 3, wherein, The extraction temperature is 87℃.

5. The process of claim 4, wherein, The extraction time was 2.3 hours.

6. The process according to claim 5, characterized in that, The high-frequency pulse in step S2 is pulsed 30 times.

7. Loquat seed polysaccharide prepared according to any one of claims 1 to 6.

8. The application of loquat seed polysaccharide obtained according to claim 7 in the preparation of antioxidants or functional foods.

Images