Feed for promoting growth of penaeus vannamei boone and preparation method of feed

Abstract

The invention relates to the technical field of feeds, in particular to a penaeus vannamei boone growth promoting feed and a preparation method thereof. The feed comprises specially-made chitosan shell microcapsule particles, each chitosan shell microcapsule particle has a double-layer structure of a sodium alginate gel inner core and an oxidative degradation chitosan thin shell, and oligosaccharide and nano / micron calcium carbonate are embedded in the inner core. Through the design, the microcapsules are kept stable in the initial stage of processing and water entry, and loss is reduced; in the intestinal tracts of the prawns, thin shells are corroded in weakly alkaline environments of midguts and posterior guts, gel disintegration is controlled in cooperation with an inner core differentiation calcium release skeleton, targeted slow release of active ingredients is achieved, and therefore the feed utilization rate is remarkably increased, prawn growth is promoted, and intestinal health is improved.

Description

Technical Field

[0001] This invention relates to the field of feed technology, and in particular to a growth-promoting feed for Litopenaeus vannamei and its preparation method. Background Technology

[0002] In the farming of Litopenaeus vannamei, the application of feed additives is crucial for improving farming efficiency. Oligosaccharides, such as xylooligosaccharides and mannooligosaccharides, have become a research hotspot due to their potential functions in regulating intestinal flora, enhancing immunity, and promoting growth. The common practice is to add these active oligosaccharides directly to the feed in free form. However, oligosaccharides are highly water-soluble and are easily lost, migrated, or degraded during the high-temperature, high-humidity pelleting process of feed processing, as well as in the initial water phase after feed introduction. This leads to a significant reduction in the effective dose actually entering the shrimp's digestive tract, resulting in waste of raw materials and uncertainty in efficacy.

[0003] To reduce losses during processing and feeding, existing technologies attempt to protect active ingredients through encapsulation. A common approach is to use natural polymers such as sodium alginate and chitosan to form microcapsules via ionogels. However, these conventional encapsulation techniques often focus on increasing encapsulation efficiency or enhancing capsule wall density, with a tendency to construct thicker, denser walls to block moisture and the external environment. While this heavily protective strategy can reduce losses during processing and initial water exposure to some extent, it can lead to another problem: excessively thick capsule walls or overly dense structures are difficult to degrade effectively in the animal's intestines. This prevents the encapsulated active ingredients from being released in time at their intended digestive tract sites (especially the mid- and hindgut, the main areas for nutrient absorption and microecological regulation), thus limiting their full physiological function.

[0004] Therefore, existing technologies face a dilemma: if oligosaccharides are not protected, they suffer significant losses during processing and initial feeding; however, if conventional encapsulation techniques are used for strong protection, release difficulties may affect their final efficacy. Most existing publicly available technologies focus on the encapsulation itself or the direct efficacy of the oligosaccharides, lacking a comprehensive solution that precisely coordinates the contradictions between processing stability in water and targeted release in the gut. Designing a feed additive carrier system that can withstand the challenges of feed processing and initial water exposure while ensuring effective release of active oligosaccharides in the hindgut of shrimp has become a key technological bottleneck for enhancing the practical application value of such additives. Summary of the Invention

[0005] In view of this, the purpose of this invention is to provide a growth-promoting feed for Litopenaeus vannamei and its preparation method, in order to solve the problems in the prior art where the active oligosaccharides that are directly added or conventionally encapsulated are easily lost in the early stages of feed processing and feeding, and the release location in the digestive tract is uncontrollable, making it difficult to balance processing stability, water stability and effective intestinal targeted release.

[0006] To achieve the above objectives, the present invention provides a growth-promoting feed for Litopenaeus vannamei, which is prepared from the following raw materials by weight: 315-350 parts casein, 95-130 parts gelatin, 265-320 parts dextrin, 90-115 parts microcrystalline cellulose, 8-12 parts taurine, 8-12 parts betaine hydrochloride, 12-18 parts aquatic feed vitamin premix, 12-18 aquatic feed mineral premix, 20-30 parts chitosan shell microcapsule particles, 60-80 parts fish oil, and 15-25 parts soybean lecithin.

[0007] Furthermore, the chitosan shell microcapsule particles comprise a sodium alginate gel core and a chitosan shell coating the sodium alginate gel core. The sodium alginate gel core contains oligosaccharides, nano-sized calcium carbonate, and micron-sized calcium carbonate. By mass fraction, the chitosan shell microcapsule particles are prepared from the following raw materials: 7-9 parts sodium alginate, 8-12 parts oligosaccharides, 0.8-1.2 parts nano-sized calcium carbonate, 1.8-2.2 parts micron-sized calcium carbonate, 2.5-3.5 parts glucono-δ-lactone, and 2.5-3.5 parts chitosan shell precursor. The oligosaccharides are xylooligosaccharides and / or mannotriose. The chitosan shell precursor is obtained by precipitating chitosan after oxidative degradation.

[0008] Preferably, the chitosan shell precursor is prepared by the following method: chitosan is added to an aqueous solution of glacial acetic acid with a mass concentration of 8-12 g / L until the solution is clear, then 30% hydrogen peroxide is added and oxidative degradation is carried out at 52-58°C for 40-60 min, then the pH is adjusted to 7.8-8.3 to precipitate chitosan, and then washed with ethanol and dried to obtain the product.

[0009] Preferably, when preparing the chitosan shell precursor, the ratio of the aqueous glacial acetic acid solution, chitosan, and hydrogen peroxide with a mass fraction of 30% is 380-420 mL: 3.5-4.5 g: 1.5-2.5 g.

[0010] Preferably, the particle size D50 of the nano-sized calcium carbonate is 50 nm, and the particle size D50 of the micron-sized calcium carbonate is 5 μm.

[0011] Preferably, the chitosan used to prepare the chitosan shell precursor has a degree of deacetylation of not less than 95% and a viscosity of 100-200 mPa·s.

[0012] Preferably, the feed is pelleted feed with a pellet diameter of 1.0 mm to 1.5 mm and a moisture content of 8.0 wt% to 9.0 wt%.

[0013] Furthermore, the present invention also provides a method for preparing a growth-promoting feed for Litopenaeus vannamei, comprising the following steps: (1) Chitosan is precipitated after oxidative degradation to obtain chitosan shell precursor; (2) Sodium alginate was mixed with oligosaccharides, and nano-sized calcium carbonate and micron-sized calcium carbonate were added in sequence to obtain the kernel precursor solution; (3) The kernel precursor solution was induced by gluconate-δ-lactone and then dripped into calcium chloride solution for calcification. It was then transferred to calcium chloride maintenance solution and allowed to stand to obtain oligosaccharide kernel wet particles. (4) Dissolve the chitosan shell precursor in glacial acetic acid aqueous solution and adjust the pH to 5.2-5.8, then coat the wet particles of oligosaccharide core and dry them to obtain chitosan shell microcapsule particles; (5) Mix casein, gelatin, dextrin, microcrystalline cellulose, taurine, betaine hydrochloride, aquatic feed vitamin premix, aquatic feed mineral premix, chitosan shell microcapsule particles, fish oil and soybean lecithin, add water and knead, cold extrude granulation and dry to obtain the growth-promoting feed for whiteleg shrimp.

[0014] Preferably, in step (3), the concentration of the calcium chloride solution is 18-22 g / L, and the concentration of the calcium chloride maintenance solution is 0.8-1.2 g / L.

[0015] Preferably, the diameter of the die hole for cold extrusion granulation in step (5) is 1.0-1.5 mm.

[0016] The beneficial effects of this invention are: Significantly improves the processing and storage stability of active ingredients: By constructing a gel core based on sodium alginate, active oligosaccharides such as xylooligosaccharides and / or mannotriose are pre-enriched and fixed in the core network. This structure can effectively buffer the heat, moisture and mechanical shear stress during processing, prevent the oligosaccharides from migrating or denaturing during the granulation and drying stage, and retain sufficient active material basis for subsequent functional performance.

[0017] This method effectively improves the water stability of feed pellets and reduces feeding losses: A low molecular weight chitosan shell is constructed outside the core through pH-controlled ionic cross-linking. This dense, positively charged shell structure can tightly bind with the negatively charged sodium alginate core, effectively preventing rapid water penetration and instantaneous dissolution of the core material in the initial stage of feed introduction into water. This significantly reduces the feed's solubility in water and improves feed utilization.

[0018] This invention achieves delayed and targeted release of active ingredients in the shrimp gut: the core-enriched, thin-shell structure possesses unique release characteristics. The chitosan shell remains stable in the acidic environment of the shrimp foregut but gradually dissolves in the weakly alkaline environment of the midgut and hindgut. Simultaneously, the differentiated calcium-releasing framework constructed from nano- and micro-sized calcium carbonate within the core responds to the intestinal environment, synergistically controlling the disintegration rhythm of the sodium alginate gel within the core. This allows the release of active oligosaccharides to be primarily localized and sustained in the midgut and hindgut segments, more precisely targeting the intestinal flora and villus absorption surface in these areas. Detailed Implementation

[0019] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to specific embodiments.

[0020] Raw materials: Xylooligosaccharides are XOS 95 powder from Shandong Bailong Chuangyuan Biotechnology Co., Ltd., with an XOS content of not less than 95%, of which XOS2-4 content is not less than 65%; Mannotriose is O-MTR from Neogen Megazyme; Chitosan is C105799 from Shanghai Aladdin Biochemical Technology Co., Ltd., with a degree of deacetylation of not less than 95% and a viscosity of 100-200 mPa·s; Sodium alginate is A2033 from Sigma-Aldrich, derived from brown algae, with a 2% solution viscosity of not less than 2000 cP and an M / G ratio of approximately 1.56; Casein is C3400 from Sigma-Aldrich; Gelatin is G2625 from Sigma-Aldrich, Type A, with a gel strength of approximately 175g. Bloom; the fish oil used is Sigma-Aldrich F8020, containing approximately 30% ω-3 fatty acids; the soybean lecithin used is Merck 429415, with a phosphatidylcholine content of no less than 94%; the particle size D50 of nano-sized calcium carbonate is 50nm; the particle size D50 of micron-sized calcium carbonate is 5μm; the remaining dextrin, microcrystalline cellulose, taurine, betaine hydrochloride, glacial acetic acid, hydrogen peroxide, sodium hydroxide, calcium chloride, and deionized water are all commercially available and readily available substances; the aquatic feed vitamin premix and mineral premix both use conventional commercially available aquatic-specific premixes.

[0021] Example 1: (1) Add 4g of chitosan to 400mL of glacial acetic acid aqueous solution with a mass concentration of 10g / L, stir at 45℃ for 60min until the solution is clear, then add 2g of hydrogen peroxide with a mass fraction of 30%, react at 55℃ for 50min, then cool to 25℃, add sodium hydroxide solution with a concentration of 2mol / L to adjust the pH to 8, so that chitosan precipitates out; then add 400mL of anhydrous ethanol to wash, filter and dry at 40℃ to obtain chitosan shell precursor; (2) Add 8g of sodium alginate to 300mL of deionized water and stir at 45℃ and 600rpm for 40min; after the sodium alginate has completely swollen, add 10g of xylooligosaccharide and continue stirring for 15min; then add 1g of nano-sized calcium carbonate and stir for 5min, then add 2g of micron-sized calcium carbonate and stir for 8min, and finally let stand for 10min to remove bubbles to obtain the kernel precursor solution. (3) Add 3g of gluconate-δ-lactone to the kernel precursor solution obtained in step (2), stir for 30s and let stand for 2min, and drop 800mL of calcium chloride solution with a mass concentration of 20g / L into the solution within 8min. The stirring speed of the calcium chloride solution is controlled at 250rpm. After standing for 30s, remove the solution and transfer it to 300mL of calcium chloride maintenance solution with a mass concentration of 1g / L. Let it stand for 10min to obtain oligosaccharide kernel wet particles. (4) Add 3g of the chitosan shell precursor obtained in step (1) to 500mL of glacial acetic acid aqueous solution with a mass concentration of 6g / L, stir at 45℃ for 30min, and then add sodium hydroxide solution with a mass concentration of 1mol / L to adjust the pH to 5.5; then add the oligosaccharide core wet particles obtained in step (3), coat at 100rpm for 8min, take it out and rinse with 200mL of calcium chloride solution with a mass concentration of 1g / L for 10s, and dry at 38℃ to constant weight to obtain chitosan thin-shell microcapsule particles; (4) Mix 330g casein, 110g gelatin, 290g dextrin, 105g microcrystalline cellulose, 10g taurine, 10g betaine hydrochloride, 15g aquatic feed vitamin premix and 15g aquatic feed mineral premix dry for 6 minutes, then add 25g chitosan thin-shell microcapsule particles obtained in step (4) and mix for 2 minutes; then add 70g fish oil and 20g soybean lecithin and mix for 5 minutes, then add 250g deionized water and knead for 10 minutes, controlling the material temperature not to exceed 40℃; then perform conventional cold extrusion granulation on the obtained soft material with a die diameter of 1.2mm, and finally dry at 40℃ to a moisture content of 8.5wt%, and after cooling, obtain the growth-promoting feed for Litopenaeus vannamei.

[0022] Example 2: (1) Add 3.5g of chitosan to 380mL of glacial acetic acid aqueous solution with a mass concentration of 8g / L, stir at 43℃ for 50min until the solution is clear, then add 1.5g of hydrogen peroxide with a mass fraction of 30%, react at 52℃ for 40min, then cool to 25℃, add sodium hydroxide solution with a concentration of 2mol / L to adjust the pH to 7.8, so that chitosan precipitates out; then add 350mL of anhydrous ethanol to wash, filter and dry at 38℃ to obtain chitosan shell precursor; (2) Add 7g of sodium alginate to 280mL of deionized water and stir at 42℃ and 550rpm for 35min; after the sodium alginate has completely swollen, add 8g of xylooligosaccharide and continue stirring for 12min; then add 0.8g of nano-sized calcium carbonate and stir for 5min, then add 1.8g of micron-sized calcium carbonate and stir for 7min, and finally let stand for 8min to remove bubbles to obtain the kernel precursor solution; (3) Add 2.5g gluconate-δ-lactone to the kernel precursor solution obtained in step (2), stir for 30s and let stand for 1min, and drop 700mL of calcium chloride solution with a mass concentration of 18g / L into the solution within 6min. The stirring speed of the calcium chloride solution is controlled at 220rpm. After standing for 20s, remove the solution and transfer it to 250mL of calcium chloride maintenance solution with a mass concentration of 0.8g / L. Let stand for 8min to obtain oligosaccharide kernel wet particles. (4) Add 2.5g of the chitosan shell precursor obtained in step (1) to 450mL of glacial acetic acid aqueous solution with a mass concentration of 5g / L, stir at 42℃ for 25min, and then add sodium hydroxide solution with a mass concentration of 1mol / L to adjust the pH to 5.2; then add the oligosaccharide core wet particles obtained in step (3), coat at 80rpm for 6min, take it out and rinse with 150mL of calcium chloride solution with a mass concentration of 0.8g / L for 8s, and dry at 36℃ to constant weight to obtain chitosan thin-shell microcapsule particles; (5) Dry mix 315g casein, 95g gelatin, 320g dextrin, 115g microcrystalline cellulose, 8g taurine, 8g betaine hydrochloride, 12g aquatic feed vitamin premix and 12g aquatic feed mineral premix for 5 minutes, then add 20g chitosan thin-shell microcapsule particles obtained in step (4) and mix for 1 minute; then add 60g fish oil and 15g soybean lecithin and mix for 4 minutes, then add 270g deionized water and knead for 8 minutes, controlling the material temperature not to exceed 38℃; then perform conventional cold extrusion granulation on the obtained soft material with a die diameter of 1.0mm, and finally dry at 38℃ to a moisture content of 8.0wt%, and after cooling, obtain the growth-promoting feed for Litopenaeus vannamei.

[0023] Example 3: (1) Add 3.8 g of chitosan to 390 mL of glacial acetic acid aqueous solution with a mass concentration of 9 g / L, stir at 44 °C for 55 min until the solution is clear, then add 1.8 g of hydrogen peroxide with a mass fraction of 30%, react at 54 °C for 45 min, then cool to 25 °C, add sodium hydroxide solution with a concentration of 2 mol / L to adjust the pH to 7.9, so that chitosan precipitates out; then add 380 mL of anhydrous ethanol to wash, filter and dry at 39 °C to obtain chitosan shell precursor; (2) Add 7.5g sodium alginate to 290mL of deionized water and stir at 44℃ and 580rpm for 38min; after the sodium alginate has completely swollen, add 10g mannotriose and continue stirring for 14min; then add 0.9g nano-sized calcium carbonate and stir for 5min, then add 1.9g micron-sized calcium carbonate and stir for 8min, and finally let stand for 9min to remove bubbles to obtain the kernel precursor solution; (3) Add 2.8g gluconate-δ-lactone to the kernel precursor solution obtained in step (2), stir for 30s and let stand for 2min, and drop 750mL of calcium chloride solution with a mass concentration of 19g / L into the solution within 7min. The stirring speed of the calcium chloride solution is controlled at 230rpm. After standing for 25s, remove the solution and transfer it to 280mL of calcium chloride maintenance solution with a mass concentration of 0.9g / L. Let stand for 9min to obtain oligosaccharide kernel wet particles. (4) Add 2.8g of the chitosan shell precursor obtained in step (1) to 480mL of glacial acetic acid aqueous solution with a mass concentration of 5.5g / L, stir at 44℃ for 28min, then add sodium hydroxide solution with a mass concentration of 1mol / L to adjust the pH to 5.3; then add the oligosaccharide core wet particles obtained in step (3), coat at 90rpm for 7min, take out and rinse with 180mL of calcium chloride solution with a mass concentration of 0.9g / L for 9s, and dry at 37℃ to constant weight to obtain chitosan thin-shell microcapsule particles; (5) Dry mix 325g casein, 105g gelatin, 300g dextrin, 100g microcrystalline cellulose, 10g taurine, 10g betaine hydrochloride, 15g aquatic feed vitamin premix and 15g aquatic feed mineral premix for 6 minutes, then add 25g chitosan thin-shell microcapsule particles obtained in step (4) and mix for 2 minutes; then add 70g fish oil and 20g soybean lecithin and mix for 5 minutes, then add 255g deionized water and knead for 10 minutes, controlling the material temperature not to exceed 39℃; then perform conventional cold extrusion granulation on the obtained soft material with a die diameter of 1.2mm, and finally dry at 39℃ to a moisture content of 8.3wt%, and after cooling, obtain the growth-promoting feed for Litopenaeus vannamei.

[0024] Example 4: (1) Add 4.2 g of chitosan to 410 mL of glacial acetic acid aqueous solution with a mass concentration of 11 g / L, stir at 45 °C for 60 min until the solution is clear, then add 2.2 g of hydrogen peroxide with a mass fraction of 30%, react at 56 °C for 55 min, then cool to 25 °C, add sodium hydroxide solution with a concentration of 2 mol / L to adjust the pH to 8.1, so that chitosan precipitates out; then add 420 mL of anhydrous ethanol to wash, filter and dry at 41 °C to obtain chitosan shell precursor; (2) Add 8.5g sodium alginate to 310mL of deionized water and stir at 46℃ and 620rpm for 42min; after the sodium alginate has completely swollen, add 6g xylooligosaccharide and 4g mannotriose and continue stirring for 18min; then add 1.1g of nano-sized calcium carbonate and stir for 5min, then add 2.1g of micron-sized calcium carbonate and stir for 8min, and finally let stand for 10min to remove bubbles to obtain the kernel precursor solution; (3) Add 3.2g gluconate-δ-lactone to the kernel precursor solution obtained in step (2), stir for 30s and let stand for 2min, and drop 850mL of calcium chloride solution with a mass concentration of 21g / L into the solution within 9min. The stirring speed of the calcium chloride solution is controlled at 260rpm. After standing for 35s, remove the solution and transfer it to 320mL of calcium chloride maintenance solution with a mass concentration of 1.1g / L. Let stand for 11min to obtain oligosaccharide kernel wet particles. (4) Add 3.2g of the chitosan shell precursor obtained in step (1) to 520mL of glacial acetic acid aqueous solution with a mass concentration of 6.5g / L, stir at 46℃ for 32min, then add sodium hydroxide solution with a mass concentration of 1mol / L to adjust the pH to 5.6; then add the oligosaccharide core wet particles obtained in step (3), coat at 110rpm for 9min, take it out and rinse with 220mL of calcium chloride solution with a mass concentration of 1.1g / L for 10s, and dry at 39℃ to constant weight to obtain chitosan thin-shell microcapsule particles; (5) Dry mix 340g casein, 120g gelatin, 280g dextrin, 95g microcrystalline cellulose, 11g taurine, 11g betaine hydrochloride, 16g aquatic feed vitamin premix and 16g aquatic feed mineral premix for 6 minutes, then add 28g chitosan thin-shell microcapsule particles obtained in step (4) and mix for 2 minutes; then add 68g fish oil and 22g soybean lecithin and mix for 5 minutes, then add 243g deionized water and knead for 10 minutes, controlling the material temperature not to exceed 39℃; then perform conventional cold extrusion granulation on the obtained soft material with a die diameter of 1.3mm, and finally dry at 39℃ to a moisture content of 8.7wt%, and after cooling, obtain the growth-promoting feed for whiteleg shrimp.

[0025] Example 5: (1) Add 4.5 g of chitosan to 420 mL of glacial acetic acid aqueous solution with a mass concentration of 12 g / L, stir at 45 °C for 60 min until the solution is clear, then add 2.5 g of hydrogen peroxide with a mass fraction of 30%, react at 58 °C for 60 min, then cool to 25 °C, add sodium hydroxide solution with a concentration of 2 mol / L to adjust the pH to 8.3, so that chitosan precipitates out; then add 450 mL of anhydrous ethanol to wash, filter and dry at 45 °C to obtain chitosan shell precursor; (2) Add 9g of sodium alginate to 320mL of deionized water and stir at 48℃ and 650rpm for 45min; after the sodium alginate has completely swollen, add 12g of xylooligosaccharide and continue stirring for 20min; then add 1.2g of nano-sized calcium carbonate and stir for 5min, then add 2.2g of micron-sized calcium carbonate and stir for 9min, and finally let stand for 12min to remove bubbles to obtain the kernel precursor solution; (3) Add 3.5g gluconate-δ-lactone to the kernel precursor solution obtained in step (2), stir for 30s and let stand for 3min, and drop 900mL of calcium chloride solution with a mass concentration of 22g / L into the solution within 10min. The stirring speed of the calcium chloride solution is controlled at 280rpm. After standing for 40s, remove the solution and transfer it to 350mL of calcium chloride maintenance solution with a mass concentration of 1.2g / L. Let stand for 12min to obtain oligosaccharide kernel wet particles. (4) Add 3.5g of the chitosan shell precursor obtained in step (1) to 550mL of glacial acetic acid aqueous solution with a mass concentration of 7g / L, stir at 48℃ for 35min, and then add sodium hydroxide solution with a mass concentration of 1mol / L to adjust the pH to 5.8; then add the oligosaccharide core wet particles obtained in step (3), coat at 120rpm for 10min, take it out and rinse with 250mL of calcium chloride solution with a mass concentration of 1.2g / L for 12s, and dry at 40℃ to constant weight to obtain chitosan thin-shell microcapsule particles; (5) Dry mix 350g casein, 130g gelatin, 265g dextrin, 90g microcrystalline cellulose, 12g taurine, 12g betaine hydrochloride, 18g aquatic feed vitamin premix and 18g aquatic feed mineral premix for 7 minutes, then add 30g chitosan thin-shell microcapsule particles obtained in step (4) and mix for 3 minutes; then add 80g fish oil and 25g soybean lecithin and mix for 6 minutes, then add 220g deionized water and knead for 12 minutes, controlling the material temperature not to exceed 40℃; then perform conventional cold extrusion granulation on the obtained soft material with a die diameter of 1.5mm, and finally dry at 40℃ to a moisture content of 9.0wt%, and after cooling, obtain the growth-promoting feed for Litopenaeus vannamei.

[0026] Comparative Example 1: The difference from Example 1 is that xylooligosaccharide is not added in step (2); 25g of chitosan thin-shell microcapsule particles are not added in step (5), but instead 10g of xylooligosaccharide and 15g of dextrin are added directly, and the other conditions are the same as in Example 1.

[0027] Comparative Example 2: The difference from Example 1 is that chitosan coating is no longer performed in step (4); instead of adding 25g of chitosan thin-shell microcapsule particles in step (5), 25g of particles obtained by drying the oligosaccharide core wet particles obtained in step (3) under the same drying conditions as in Example 1 are added, and the other conditions are the same as in Example 1.

[0028] Comparative Example 3: The difference from Example 1 is that: in step (4), the oligosaccharide kernel wet particles obtained in step (3) are no longer coated; the chitosan solution prepared in step (4) is added to the kernel precursor solution obtained in step (2) before the start of step (3), and after mixing for 30 seconds, it is directly dropped into a calcium chloride solution with a mass concentration of 20 g / L, and dried under the same conditions as in Example 1. In step (5), 25 g of the obtained particles are added, and the remaining conditions are the same as in Example 1.

[0029] Comparative Example 4: The difference from Example 1 is that hydrogen peroxide is not added in step (1). After the chitosan is completely dissolved, it is directly cooled to 25°C and the pH is adjusted to 8 to precipitate the chitosan shell precursor. The other conditions are the same as in Example 1.

[0030] Comparative Example 5: The difference from Example 1 is that in step (2), instead of adding 1g of nano-sized calcium carbonate, 3g of micron-sized calcium carbonate is added at once to keep the total amount of calcium carbonate added consistent. The other conditions are the same as in Example 1.

[0031] Comparative Example 6: The difference from Example 1 is that in step (2), instead of adding 2g of micron-sized calcium carbonate, 2g of nano-sized calcium carbonate is added on the basis of the original 1g of nano-sized calcium carbonate to keep the total amount of calcium carbonate added consistent. The other conditions are the same as in Example 1.

[0032] Comparative Example 7: The difference from Example 1 is that gluconate-δ-lactone is not added in step (3), and the particles are not transferred into a calcium chloride maintenance solution with a mass concentration of 1 g / L and left to stand for 10 min. Instead, they are directly soaked in a calcium chloride solution with a mass concentration of 20 g / L for 10 min and then taken out. The other conditions are the same as in Example 1.

[0033] Performance testing: Preparation and source of test samples: The test samples included the finished feed obtained in step (5) of Examples 1-5 and the finished feed obtained in Comparative Examples 1-7. All finished feeds were dried at below 40℃ to a moisture content of 8.0wt%-9.0wt% and then cooled for 24h. Particles with a particle size of 1.0-1.5mm, intact appearance and no obvious powdering were screened, packed into aluminum foil composite bags, and sealed and stored at 4℃ in the dark. The whiteleg shrimp used in the aquaculture experiment were purchased from the same batch of healthy seedlings from the same hatchery. Before the test, white spot syndrome was tested according to GB / T 28630.2-2012 and the result was negative. After temporary rearing for 7 days, individuals with an initial body weight of 0.80g±0.05g were selected for subsequent experiments.

[0034] The standard composition of finished feed: all finished feeds were processed by crushing and passing through a 0.425mm sieve. The moisture content was determined according to GB / T6435-2014, the crude protein according to GB / T 6432-2018, the crude fat according to GB / T 6433-2025, and the crude fiber according to GB / T 6434-2022. Each sample was tested in triplicate.

[0035] 20-minute stability in water: The 20-minute immersion test was conducted according to the standard of GB / T 22919.5-2008. 5.000 g of intact particles from each sample were weighed, placed in a 100-mesh polyester sieve bag, and immersed in 500 mL of artificial seawater with a salinity of 25‰±1‰ and a temperature of 28.0℃±1.0℃. The mixture was allowed to stand for 20 minutes, drained for 30 seconds, dried at 105℃ for 2 hours, cooled to room temperature, and weighed. The 20-minute solubility loss rate was calculated based on the difference in dry weight before and after immersion. Six replicates were performed for each group.

[0036] Growth performance of Litopenaeus vannamei: The effectiveness of the target animal was evaluated according to GB / Z 31812-2015. A total of 12 groups were set up, including Examples 1-5 and Comparative Examples 1-7, with 3 replicates per group and 30 shrimp per replicate. The culture containers were 300L fiberglass tanks with an effective water volume of 250L. Continuous aeration was maintained. The temperature was kept at 28.0℃±1.0℃, the salinity at 25‰±1‰, the pH was monitored and controlled at 7.8-8.2 according to GB / T 6920-1986, dissolved oxygen was monitored and maintained at no less than 6.0 mg / L according to HJ 506-2009, and ammonia nitrogen was monitored and maintained at no more than 0.20 mg / L according to HJ 535-2009. Feeding was conducted daily at 6:00, 11:00, 17:00, and 22:00 until apparent saturation. The experiment lasted 56 days, and feed intake, mortality, water exchange volume, and any abnormalities were recorded. After the experiment, the final average body weight was measured, and the weight gain rate, specific growth rate, survival rate and feed conversion ratio were calculated. The weight gain rate was calculated as follows: weight gain rate = (final average body weight - initial average body weight) / initial average body weight × 100%; specific growth rate = [ln final average body weight - ln initial average body weight] / 56 × 100%; survival rate = final number of tails / initial number of tails × 100%; feed conversion ratio = feed intake / weight gain.

[0037] Survival rate after ammonia nitrogen stress: After the growth experiment, 15 healthy surviving Litopenaeus vannamei shrimp were randomly selected from each parallel tank. After 24 hours of fasting, they were transferred to 30L glass tanks. 20L of artificial seawater with a salinity of 25‰±1‰ was added to each tank. The stress water with an ammonia nitrogen concentration of 20.0mg / L was prepared using ammonium chloride. The temperature was maintained at 28.0℃±1.0℃, the pH was maintained at 8.0±0.2, and continuous aeration was provided. No feeding was provided within 24 hours. Ammonia nitrogen was measured according to HJ 535-2009 before and at the end of the stress to verify the concentration, and the 24-hour survival rate was calculated.

[0038] Intestinal tissue morphology: After the growth experiment, three Pacific white shrimp were randomly selected from each parallel study. After fasting for 24 hours, the midgut was dissected under ice anesthesia. The midgut segment was fixed, dehydrated, embedded in paraffin, sectioned, and stained according to GB / T 28630.4-2012. 10% neutral formaldehyde was used as the fixative, and fixation was performed for 24 hours. Paraffin sections were 5 μm thick, stained with hematoxylin and eosin, and observed under an optical microscope. Five complete fields of view were randomly selected from each shrimp to measure the height of the intestinal villi and the thickness of the muscle layer.

[0039] Table 1 Performance Test Results Data Analysis: As can be seen from the data in Table 1, the growth-promoting feed for Litopenaeus vannamei prepared in this invention, while maintaining a low 20-minute solubility loss rate and good particle integrity, still achieves a high weight gain rate, a low feed conversion ratio, a high survival rate after ammonia nitrogen stress, and a superior intestinal tissue morphology. This indicates that the system does not simply pursue water stability, but achieves a good balance between water stability protection and intestinal release. The reason for this is presumably that the sodium alginate core first enriches xylooligosaccharides and / or mannotrioses; nano-sized calcium carbonate and micron-sized calcium carbonate construct a fast-slow calcium release framework; gluconate-δ-lactone, combined with exogenous calcium chloride and low-concentration calcium chloride maintenance steps, makes the core cross-linking more balanced; and the subsequently formed low-molecular-weight chitosan thin shell reduces loss during processing and initial water entry without excessively hindering subsequent diffusion, thus making the active ingredients more conducive to their sustained effect in the intestines of Litopenaeus vannamei. Especially when xylooligosaccharides and mannotrisaccharides are used simultaneously with appropriate coating and calcification conditions, the overall growth performance, ammonia tolerance and intestinal morphology improvement are more coordinated, demonstrating the overall promoting effect after the coupling of multiple factors.

[0040] As can be seen from the data in Table 1 for Example 1 and Comparative Example 1, when xylooligosaccharides were added directly to the feed without microencapsulation, the 20-minute solubility rate increased significantly, and the weight gain rate, feed conversion ratio, survival rate after ammonia nitrogen stress, and intestinal morphology indicators all deteriorated simultaneously. The main reason for this is that free xylooligosaccharides are more prone to migration or dissolution during granulation, drying, and the initial stages of water introduction, reducing the effective portion that actually enters the intestine and continues to exert its effect.

[0041] As can be seen from the data in Table 1 for Examples 1, 2, and 3, retaining only the sodium alginate core without the subsequent chitosan shell, or blending the chitosan with the core precursor solution in advance, will reduce the overall performance. The former mainly manifests as insufficient outer barrier and rapid loss in the initial stage of water immersion; the latter easily leads to unclear shell position and disordered diffusion paths within the particles, making it difficult to form a clear core-shell division of labor.

[0042] As can be seen from the data in Table 1 for Example 1 and Comparative Example 4, when using protochitosan directly without oxidative degradation, although the 20-minute solubility loss rate was not the worst, the weight gain, feed conversion ratio, survival rate after ammonia nitrogen stress, and intestinal morphology were still lower than in the examples. It is speculated that this is because higher molecular weight chitosan is more likely to form a thicker and more uneven outer layer, leading to increased local brittleness after drying. Simultaneously, it exhibits greater diffusion resistance in the intestinal environment, which is not conducive to the release of oligosaccharides at the intended location.

[0043] As can be seen from the data in Table 1 for Examples 1, 5, and 6, neither micron-sized nor nano-sized calcium carbonate alone can achieve the comprehensive effects of the examples. When only micron-sized calcium carbonate is used, the core formation speed is slow, the cross-linked framework is coarse, and the protective effect is insufficient; when only nano-sized calcium carbonate is used, local calcium release is too rapid, easily forming a structure with an overly dense surface and uneven interior. When the two particle sizes are used in combination, first nano-sized and then micron-sized, it is beneficial for both initial nucleation and subsequent support. Therefore, it exhibits a significant synergistic gain in water stabilization, release, growth promotion, and intestinal improvement, demonstrating a technical effect greater than simple superposition.

[0044] As can be seen from the data in Table 1 for Example 1 and Comparative Example 7, after eliminating gluconate-δ-lactone and omitting the low-concentration calcium chloride maintenance step, the weight gain rate, survival rate after ammonia nitrogen stress, and the extent of improvement in intestinal morphology were all significantly limited. The main reason is that long-term treatment with exogenous calcium chloride alone is more likely to form a structure that is initially dense on the surface and then confined internally. This makes the particle surface appear stable, but it is not conducive to the effective release of oligosaccharides in the intestine.

[0045] Those skilled in the art should understand that the discussion of any of the above embodiments is merely exemplary and is not intended to imply that the scope of the invention is limited to these examples; within the framework of the invention, the technical features of the above embodiments or different embodiments can also be combined, and there are many other variations of the different aspects of the invention as described above, which are not provided in detail for the sake of brevity.

Claims

1. A growth-promoting feed for Litopenaeus vannamei, characterized in that, The product is prepared from the following raw materials in parts by weight: 315-350 parts casein, 95-130 parts gelatin, 265-320 parts dextrin, 90-115 parts microcrystalline cellulose, 8-12 parts taurine, 8-12 parts betaine hydrochloride, 12-18 parts aquatic feed vitamin premix, 12-18 aquatic feed mineral premix, 20-30 parts chitosan shell microcapsule particles, 60-80 parts fish oil, and 15-25 parts soybean lecithin. The chitosan shell microcapsule particles comprise a sodium alginate gel core and a chitosan shell coating the sodium alginate gel core. The sodium alginate gel core contains oligosaccharides, nano-sized calcium carbonate, and micron-sized calcium carbonate. By mass, the chitosan shell microcapsule particles are prepared from the following raw materials: 7-9 parts sodium alginate, 8-12 parts oligosaccharides, 0.8-1.2 parts nano-sized calcium carbonate, 1.8-2.2 parts micron-sized calcium carbonate, 2.5-3.5 parts glucono-δ-lactone, and 2.5-3.5 parts chitosan shell precursor. The oligosaccharides are xylooligosaccharides and / or mannotriose, and the chitosan shell precursor is obtained by precipitating chitosan after oxidative degradation.

2. The growth-promoting feed for Litopenaeus vannamei according to claim 1, characterized in that, The chitosan shell precursor is prepared by the following method: chitosan is added to an aqueous solution of glacial acetic acid with a mass concentration of 8-12 g / L until the solution is clear, then 30% hydrogen peroxide is added and oxidative degradation is carried out at 52-58℃ for 40-60 min. Subsequently, the pH is adjusted to 7.8-8.3 to precipitate chitosan, which is then washed with ethanol and dried to obtain the product.

3. The growth-promoting feed for Litopenaeus vannamei according to claim 2, characterized in that, When preparing the chitosan shell precursor, the ratio of the aqueous glacial acetic acid solution, chitosan, and hydrogen peroxide with a mass fraction of 30% is 380-420 mL: 3.5-4.5 g: 1.5-2.5 g.

4. The growth-promoting feed for Litopenaeus vannamei according to claim 1, characterized in that, The particle size D50 of the nano-sized calcium carbonate is 50 nm, and the particle size D50 of the micron-sized calcium carbonate is 5 μm.

5. The growth-promoting feed for Litopenaeus vannamei according to claim 1, characterized in that, The degree of deacetylation of the chitosan used to prepare the chitosan shell precursor is not less than 95%, and the viscosity is 100-200 mPa·s.

6. The growth-promoting feed for Litopenaeus vannamei according to claim 1, characterized in that, The feed is pelleted feed with a pellet diameter of 1.0mm-1.5mm and a moisture content of 8.0wt%-9.0wt%.

7. A method for preparing a growth-promoting feed for Litopenaeus vannamei according to any one of claims 1-6, characterized in that, Includes the following steps: (1) Chitosan is precipitated after oxidative degradation to obtain chitosan shell precursor; (2) Sodium alginate was mixed with oligosaccharides, and nano-sized calcium carbonate and micron-sized calcium carbonate were added in sequence to obtain the kernel precursor solution; (3) The kernel precursor solution was induced by gluconate-δ-lactone and then dripped into calcium chloride solution for calcification. It was then transferred to calcium chloride maintenance solution and allowed to stand to obtain oligosaccharide kernel wet particles. (4) Dissolve the chitosan shell precursor in glacial acetic acid aqueous solution and adjust the pH to 5.2-5.8, then coat the wet particles of oligosaccharide core and dry them to obtain chitosan shell microcapsule particles; (5) Mix casein, gelatin, dextrin, microcrystalline cellulose, taurine, betaine hydrochloride, aquatic feed vitamin premix, aquatic feed mineral premix, chitosan shell microcapsule particles, fish oil and soybean lecithin, add water and knead, cold extrude granulation and dry to obtain the growth-promoting feed for whiteleg shrimp.

8. The method for preparing the growth-promoting feed for Litopenaeus vannamei according to claim 7, characterized in that, In step (3), the concentration of the calcium chloride solution is 18-22 g / L, and the concentration of the calcium chloride maintenance solution is 0.8-1.2 g / L.

9. The method for preparing the growth-promoting feed for Litopenaeus vannamei according to claim 7, characterized in that, The die diameter for cold extrusion granulation in step (5) is 1.0-1.5 mm.

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