Egg-laying hen premix with high-efficiency enrichment of dha and egg preservation and application thereof

Abstract

The present application relates to a kind of egg-laying hens premix with DHA efficient enrichment and egg preservation, and application thereof, belong to animal nutrition and feed technology field.The present application first clearly that phospholipid deficiency is the core reason of limiting DHA enrichment efficiency, reveals that soy lecithin promotes the molecular mechanism of DHA deposition by up-regulating FABP, PCTP expression, and proves that astaxanthin through direct free radical scavenging and the double synergistic antioxidant mechanism of activating endogenous antioxidant enzyme system.Based on this, with microencapsulated schizochytrium sp powder, high-purity soy lecithin, astaxanthin as core component, compound probiotics, lutein, organic selenium, construct four-effect synergistic premix formula, and establish "DHA enrichment-egg quality-shelf life" linkage parameter system.Application of the present application, egg DHA enrichment amount reaches 500~1000 mg / 100 g yolk, feeding 2 weeks stable enrichment, egg production rate keeps more than 85%, shelf life is extended by 15~20 days, with efficient nutrition fortification and preservation function.

Description

Technical Field

[0001] This invention belongs to the field of animal nutrition and feed technology, specifically relating to a premixed feed for laying hens that combines efficient DHA enrichment with egg preservation and its application. Background Technology

[0002] Docosahexaenoic acid (DHA), an essential omega-3 polyunsaturated fatty acid with extremely low bioavailability, plays a crucial role in brain development, vision protection, and cardiovascular health. DHA intake levels in the diets of Chinese residents are generally low, far below the recommended intake. Fortifying poultry eggs with DHA is an effective way to supplement DHA due to its palatability, high absorption rate, and wide applicability. Therefore, developing specialized laying hen feed that can efficiently increase DHA deposition in eggs has significant market application value and industrial promotion significance.

[0003] Currently, technologies and products used to produce DHA-fortified eggs generally suffer from core drawbacks that are difficult to avoid. On the one hand, simply adding DHA-rich algal oil or fish oil directly to feed can enrich eggs with DHA, but high-dose additions can easily lead to decreased egg production and reduced egg quality, and the bioavailability and deposition efficiency of DHA are limited. On the other hand, although existing research has found that phospholipids have a certain promoting effect on DHA enrichment, it has not yet been clarified that phospholipid deficiency is the core cause of reduced DHA enrichment efficiency, nor has it explored in depth the molecular mechanism by which phospholipids promote DHA deposition, resulting in a lack of targeted guidance for improving DHA enrichment efficiency. In addition, as an unsaturated fatty acid, DHA is highly susceptible to lipid peroxidation during egg storage, causing a decline in egg quality and a shortened shelf life. Existing antioxidant preservation methods are mostly single-component additions, lacking systematic research on synergistic antioxidant mechanisms, resulting in very limited preservation effects.

[0004] Currently, most DHA-fortified egg feeds are single-nutrient fortification formulas with limited functionality. They are often simple blends of conventional ingredients, lacking carrier processes to protect the stability of active ingredients and failing to offer refined formulations tailored to different growth stages of laying hens. More importantly, existing research focuses solely on the direct link between feed formulation and egg quality, neglecting to incorporate concepts such as gut nutrition regulation and green ecological farming into the technical system, nor extending to the deep processing industry chain. The overall technical solutions are relatively simplistic. Therefore, while ensuring laying performance and egg quality, the core technical challenge that urgently needs to be addressed in the industry is how to achieve targeted and efficient enrichment of DHA at the molecular mechanism level, constructing a multi-effect synergistic technical solution integrating "nutrient fortification, preservation, gut regulation, and ecological farming," and forming a standardized and scalable full-industry chain DHA-fortified egg production system. Summary of the Invention

[0005] This invention discloses a premix for laying hens that combines efficient DHA enrichment with egg preservation and its application. The premix uses microencapsulated Schizochytrium powder, high-purity soybean lecithin (phosphatidylcholine content ≥70%), and natural astaxanthin as core components, compounded with Bacillus subtilis and Lactobacillus plantarum probiotics, lutein, and organic selenium, and supplemented with conventional feed additives required during the laying period of laying hens. This invention reveals for the first time that phospholipid deficiency is the core reason limiting DHA enrichment efficiency, elucidates the molecular mechanism by which soybean lecithin promotes DHA transport and deposition by upregulating FABP and PCTP expression, and explores the dual synergistic antioxidant mechanism of astaxanthin through direct free radical scavenging and activation of endogenous antioxidant enzyme systems. Through a four-effect synergistic formula and a linkage parameter system of "DHA enrichment-egg quality-shelf life", this premix can achieve a DHA enrichment of 500-1000 mg / 100g egg yolk in eggs. The enrichment is stable after 2 weeks of feeding, the egg production rate of laying hens is maintained at over 85%, the shelf life is extended by 15-20 days under 4℃ storage conditions, and it can also improve the intestinal health of laying hens and reduce manure emissions. It is suitable for a variety of laying hen breeds and can be extended to other livestock and poultry, and has important industrial and ecological application value.

[0006] On the one hand, this invention provides a premixed feed for laying hens that combines efficient DHA enrichment with egg preservation, using the following technical solution: A premixed feed for laying hens that combines efficient DHA enrichment with egg preservation consists of core active components and functional compound components. The core active components are microencapsulated Schizochytrium powder, high-purity soybean lecithin, and astaxanthin. The functional compound components are compound probiotics, lutein, and organic selenium. Based on the total mass of the laying hen compound feed, the DHA content provided by the microencapsulated Schizochytrium powder is 2-4 mg / g, the addition amount of high-purity soybean lecithin is 1%-2%, and the addition amount of astaxanthin is 0.05%.

[0007] Preferably, the phosphatidylcholine content in high-purity soybean lecithin is ≥70%.

[0008] Preferably, the compound probiotics consist of Bacillus subtilis and Lactobacillus plantarum; the total live bacteria count of the compound probiotics is ≥1×10⁻⁶ based on the total mass of the laying hen compound feed. 9 CFU / kg.

[0009] Preferably, the amount of lutein added is 20-50 mg / kg and the amount of organic selenium added is 0.1-0.3 mg / kg, based on the total mass of the compound feed for laying hens.

[0010] Preferably, the amount of lutein added is 30 mg / kg and the amount of organic selenium added is 0.2 mg / kg, based on the total mass of the compound feed for laying hens.

[0011] On the other hand, the present invention also provides the application of a premixed feed for laying hens in the preparation of compound feed for laying hens during the laying period.

[0012] A method for producing eggs that combine efficient DHA enrichment and preservation functions involves adding the above-mentioned laying hen premix to the basal diet of laying hens aged 30 weeks and above, and feeding them continuously for at least 2 weeks.

[0013] Preferred laying hens are Hy-Line Brown hens.

[0014] Preferred feed results in DHA enrichment in egg yolks of eggs reaching 500-1000 mg / 100 g, and the egg production rate of laying hens remains above 85% throughout the entire laying process.

[0015] Preferably, the resulting eggs have a shelf life extended by 15-20 days under storage conditions at 4°C, a DHA oxidation inhibition rate of over 75%, and a 36% reduction in egg weight loss.

[0016] In summary, the beneficial effects of the present invention are as follows: First, this invention is the first to demonstrate at the molecular mechanism level that phospholipid deficiency is the core reason limiting DHA enrichment efficiency. It elucidates the mechanism by which soybean lecithin promotes DHA transport and deposition by upregulating FABP and PCTP expression, and reveals the dual synergistic antioxidant mechanism of astaxanthin through direct free radical scavenging and activation of endogenous antioxidant enzyme systems. Based on this, the premix can enrich eggs with 500-1000 mg / 100 g egg yolk DHA, achieving stable enrichment after 2 weeks of feeding, with an enrichment efficiency 30%-45% higher than existing technologies. Simultaneously, under 4°C storage conditions, the shelf life is extended by 15-20 days, the DHA oxidation inhibition rate exceeds 75%, and the egg weight loss rate is reduced by 36%, significantly improving the nutritional quality and preservation performance of DHA-fortified eggs. Secondly, this invention constructs a four-effect synergistic formula of "high-efficiency DHA enrichment + egg preservation + compound nutrition fortification + intestinal regulation", which is compounded with probiotics, lutein and organic selenium to achieve compound nutrition fortification of lutein and selenium in egg yolks and improve the intestinal flora structure of laying hens, thereby increasing the digestibility and absorption rate by 10% and reducing nitrogen and phosphorus emissions from manure by 18% to 22%. At the same time, the established linkage parameter system of "DHA enrichment-egg quality-shelf life" ensures the standardized design and scalability of the formula, which is suitable for a variety of laying hen breeds and can be extended to broilers, waterfowl, etc., maintaining the egg production rate at over 85% throughout the process, taking into account breeding efficiency, product safety and ecological application. Detailed Implementation

[0017] The present invention will be further described in detail below with reference to the embodiments.

[0018] Example Example 1 The specific steps for testing the effects of different DHA sources on laying performance and egg quality in laying hens are as follows: Seventy-twoty Hy-Line Brown chickens aged 30 weeks in good condition and with similar egg production rates were randomly divided into 15 groups, with six replicates per group and eight chickens per replicate. The control group was fed a basal diet, while the experimental groups were supplemented with 1, 2, 4, 8, and 10 mg / g DHA sources, respectively, using Schizochytrium powder, Schizochytrium oil, and fishmeal. The experiment lasted for eight weeks.

[0019] This study investigated the effects of adding different levels (1, 2, 4, 8, 10 mg / g) of DHA from Schizochytrium powder to the diet on the egg production performance of laying hens. Egg production rate, average egg production, average egg weight, average feed intake, and feed conversion ratio were statistically analyzed in three stages: 1-4 weeks, 5-8 weeks, and 1-8 weeks.

[0020] As shown in Table 1, compared with the control group, there were no significant differences in egg production rate, average egg production, average feed intake, and feed conversion ratio among the algae powder-added groups at each stage. p >0.05), although the average egg weight showed a linear decreasing trend from 1 to 8 weeks, the 2 mg / g group had the highest egg weight (60.62 g) and there was no significant difference from the control group. The egg weight of the 8 mg / g and 10 mg / g groups decreased slightly but was still within the normal range.

[0021] Table 1. Effects of different levels of algae powder supplementation in diet on egg production performance of laying hens.

[0022] To investigate the effects of adding different levels of Schizochytrium powder (1, 2, 4, 8, 10 mg / g DHA) to the diet on egg quality.

[0023] The results are shown in Table 2. Compared with the control group, the protein height and Hardy units of the low-added (1-2 mg / g) algal powder group were significantly increased. p <0.0001), while high addition levels (≥4 mg / g) caused albumen height and Haugh units to return to or slightly lower than the control group level; egg weight showed no significant difference among the groups, but yolk weight was slightly reduced in the 8 mg / g and 10 mg / g groups; there were no significant differences among the groups in yolk color, shell strength, shell thickness, egg shape index, eggshell color, and yolk trimethylamine (TMA) content. p >0.05). Overall, the results show that the addition of Schizochytrium powder in the range of 2-4 mg / g has no adverse effect on egg quality, and moderate addition can improve albumen height and Haugh units.

[0024] Table 2. Effects of different levels of algae meal supplementation in diets on egg quality.

[0025] The study investigated the effects of adding different levels of Schizochytrium powder (1, 2, 4, 8, 10 mg / g DHA) to the diet on the quality of eggs stored for 21 days.

[0026] As shown in Table 3, compared with the control group, the egg yolk color of each algae powder-added group was significantly improved ( p <0.05, with the 2mg / g group showing the highest yolk color (5.49); the eggshell redness a value was significantly higher in all supplemented groups than in the control group (12.50), reaching 14.29-18.03 ( p <0.01); while there were no significant differences among groups in egg weight, albumen height, Haugh unit, eggshell strength, eggshell thickness, egg shape index, eggshell weight, yolk weight, and eggshell brightness L value. p >0.05).

[0027] Table 3. Effects of different levels of algae powder added to the diet on egg quality after 21 days of storage.

[0028] The study investigated the effects of adding different levels of Schizochytrium powder (1, 2, 4, 8, 10 mg / g DHA) to the diet on the quality of eggs after 42 days of storage.

[0029] As shown in Table 4, compared with the control group, the egg yolk color of each algae powder-added group was significantly improved ( p <0.0001, with the 10 mg / g group showing the highest yolk color (5.99); the eggshell redness a value also increased significantly in all added groups ( p =0.001), reaching 15.42-18.38; the egg weight of the 1 mg / g and 8 mg / g groups was significantly higher than that of the control group ( p =0.010). However, there were no significant differences among the groups in albumen height, Haugh unit, shell strength, shell thickness, egg shape index, yolk weight, yolk ratio, and eggshell color L and b values. p >0.05). This indicates that even after 42 days of storage, the addition of Schizochytrium powder can still effectively improve the yolk color and shell redness, and has no adverse effects on other egg quality indicators.

[0030] Table 4. Effects of different levels of algae powder added to the diet on egg quality after 42 days of storage.

[0031] To investigate the effects of adding different levels of Schizochytrium powder (1, 2, 4, 8, 10 mg / g DHA) to the diet on the fatty acid composition of egg yolk.

[0032] As shown in Table 5, with the increase of algae powder addition, the content of saturated fatty acids (SFA) and monounsaturated fatty acids in egg yolk showed a decreasing trend, with the SFA content in the 10 mg / g group being significantly lower than that in the control group. p =0.003), and the MUFA in each added group was significantly lower than that in the control group ( p =0.002); the contents of polyunsaturated fatty acids (PUFA), n-3 PUFA, total n-3+n-6 PUFA, and DHA increased significantly with increasing algae powder addition. p <0.01), with the DHA content increasing from a maximum of 3.58 mg / g in the control group to 30.98 mg / g (10 mg / g group), and the corresponding DHA enrichment increasing from a maximum of 159.29 mg / 100g egg yolk to 1388.59 mg / 100g egg yolk (8 mg / g group); at the same time, the n-6 / n-3 ratio significantly decreased from 14.47 in the control group to 2.13 (10 mg / g group) ( p <0.0001).

[0033] Table 5. Effects of different levels of algae powder supplementation in diet on fatty acid composition of egg yolk.

[0034] To investigate the effects of adding different levels of Schizochytrium oil (1, 2, 4, 8, 10 mg / g DHA) to the diet on the egg production performance of laying hens.

[0035] As shown in Table 6, low-dose algal oil (1-4 mg / g) had no significant adverse effects on egg production rate, egg quantity, and egg weight within 1-4 weeks. However, with prolonged feeding time (5-8 weeks) and increased dosage (≥8 mg / g), egg production performance significantly declined. Specifically, within 1-4 weeks, the egg production rates of the 8 mg / g and 10 mg / g groups decreased to 84.97% and 77.68%, respectively, significantly lower than the 91.15% of the control group. p <0.0001); During weeks 5-8, the egg production rate of the 8 mg / g and 10 mg / g groups further decreased to 62.43% and 53.79%, respectively, and the average egg production decreased to 35.82 g / h·d and 31.33 g / h·d, respectively. The feed conversion ratio increased to 2.77 and 3.13, respectively, all of which were significantly worse than the control group. p <0.01). Overall data from 1 to 8 weeks also confirmed that the egg production rate, egg production, egg weight, and feed intake were significantly reduced in the 8 mg / g and 10 mg / g algal oil groups, while the feed conversion ratio increased.

[0036] Table 6. Effects of different levels of algal oil supplementation in diets on egg production performance of laying hens.

[0037] To investigate the effects of adding different levels of Schizochytrium oil (1, 2, 4, 8, 10 mg / g DHA) to the diet on egg quality.

[0038] As shown in Table 7, with the increase of algal oil addition, the egg yolk color showed a significant decreasing trend. The egg yolk color of the 2-10 mg / g group (3.42-3.47) was significantly lower than that of the control group (4.15). p <0.0001); albumen height was significantly decreased in the 1 mg / g group (5.93 mm vs. control group 7.16 mm), but increased to 7.12 mm in the 10 mg / g group with no significant difference from the control group; Haugh units were significantly decreased in the 1 mg / g group (75.73 vs. control group 84.33), but recovered to 84.88 in the 10 mg / g group; eggshell weight was significantly decreased in the 8 mg / g group (6.13 g vs. control group 6.69 g) ( p =0.047); the content of trimethylamine (TMA) in egg yolk was significantly higher in all algal oil-added groups than in the control group (2.12-2.32 μg / g vs 1.85 μg / g). p =0.024). Other indicators such as egg weight, eggshell strength, eggshell thickness, egg shape index, yolk weight, and eggshell color showed no significant differences.

[0039] Table 7. Effects of different levels of algal oil supplementation in diets on egg quality.

[0040] The study investigated the effects of adding different levels of Schizochytrium oil (1, 2, 4, 8, 10 mg / g DHA) to the diet on the quality of eggs after 21 days of storage.

[0041] As shown in Table 8, compared with the control group, the egg yolk color of the 2 mg / g algal oil group was significantly reduced (1.89 vs 3.72). p =0.001), while the yolk color of other groups showed no significant difference from the control group; eggshell strength, eggshell thickness, egg shape index, eggshell weight, yolk weight, yolk ratio, and eggshell color L, a, b* values ​​showed no significant differences among the groups ( p >0.05); Although protein levels and Haugh units were slightly increased in the 8 mg / g and 10 mg / g groups, the differences did not reach a statistically significant level. p >0.05). This indicates that, except for the 2 mg / g addition which had a significant adverse effect on the yolk color of eggs stored for 21 days, the other doses of algal oil had no significant negative effects on the quality indicators of eggs during storage.

[0042] Table 8. Effects of different levels of algal oil supplementation in diets on egg quality after 21 days of storage.

[0043] The study investigated the effects of adding different levels of Schizochytrium oil (1, 2, 4, 8, 10 mg / g DHA) to the diet on the quality of eggs after 42 days of storage.

[0044] As shown in Table 9, compared with the control group, the protein height of the 4 mg / g and 8 mg / g groups (4.34 mm and 4.78 mm, respectively) was significantly lower than that of the control group (5.63 mm). p =0.003), and the Hastings units in the 4 mg / g group (65.23) and 8 mg / g group (67.18) were also significantly lower than those in the control group (74.30). p =0.013); the eggshell redness a value in the 2 mg / g (14.62), 4 mg / g (14.98), and 10 mg / g (14.49) groups was significantly lower than that in the control group (16.36, ). p =0.022). However, there were no significant differences among the groups in egg weight, yolk color, eggshell strength, eggshell thickness, egg shape index, yolk weight, yolk ratio, eggshell brightness (L-value), and yellowness (b-value). p >0.05). This indicates that high doses of algal oil (≥4 mg / g) significantly reduce albumen height, Haugh unit, and eggshell redness after long-term storage (42 days), adversely affecting egg quality.

[0045] Table 9. Effects of different levels of algal oil supplementation in diets on egg quality after 42 days of storage.

[0046] To investigate the effects of adding different levels of Schizochytrium oil (1, 2, 4, 8, 10 mg / g DHA) to the diet on the fatty acid composition of egg yolk.

[0047] As shown in Table 10, with the increase of algal oil addition, the content of saturated fatty acids (SFA) and monounsaturated fatty acids (MUFA) in egg yolks both showed a significant decreasing trend. p <0.01, among which, the SFA in the 10 mg / g group decreased to 168.15 mg / g and MUFA decreased to 190.25 mg / g, both significantly lower than the 219.63 mg / g and 287.33 mg / g in the control group; the contents of n-3 PUFA and DHA increased significantly with increasing algal oil addition ( p <0.0001), DHA content increased from 3.58 mg / g in the control group to 25.97 mg / g in the 10 mg / g group, corresponding to an increase in DHA enrichment from 159.29 mg / 100g egg yolk to 1148.94 mg / 100g egg yolk; the n-6 / n-3 ratio decreased significantly from 14.47 in the control group to 2.73 (p <0.0001). However, there was no significant difference in n-6 PUFA content among the groups ( p >0.05, although the total PUFA content increased, it did not reach a statistically significant level ( p =0.127). This indicates that adding Schizochytrium oil can significantly increase the enrichment of DHA in egg yolk and optimize the fatty acid composition, and the reduction effect of high dose algal oil on SFA and MUFA is more obvious.

[0048] Table 10 Effects of different levels of algal oil supplementation in diet on fatty acid composition of egg yolk

[0049] To investigate the effects of adding different levels of fishmeal (1, 2, 4, 6 mg / g DHA) to the diet on the egg production performance of laying hens.

[0050] As shown in Table 11, during weeks 1-4, there were no significant differences in egg production rate, average egg production, average egg weight, average feed intake, and feed conversion ratio between the fishmeal-supplemented groups and the control group. p >0.05). During weeks 5-8, the average egg production (48.33 g / h·d) and average egg weight (56.70 g) of the 6 mg / g fishmeal group were significantly lower than those of the control group (51.43 g / h·d and 59.69 g), while the feed conversion ratio (2.27) was significantly higher than that of the control group (2.10). p =0.044), while there were no significant differences between the 1, 2, and 4 mg / g groups and the control group. During the overall 1-8 week period, only the 6 mg / g group had a significantly lower average egg weight (57.48 g) than the control group (59.91 g). p =0.014), and there were no significant differences in other indicators.

[0051] Table 11 Effects of different levels of fishmeal supplementation in diet on egg production performance of laying hens

[0052] To investigate the effects of different levels of fishmeal (1, 2, 4, 6 mg / g DHA) on egg quality.

[0053] As shown in Table 12, there were significant differences in egg yolk color among the groups. p =0.038), where the yolk color of the 1 mg / g group (3.78) was significantly lower than that of the control group (4.15), while the yolk color of the 2 mg / g (4.69) and 4 mg / g (4.50) groups was significantly higher than that of the control group, and the 6 mg / g group (4.24) had no significant difference from the control group; there were also significant differences in Haugh units ( p=0.048), the 1 mg / g group (79.04) was significantly lower than the control group (84.33), while the 4 mg / g (87.42) and 6 mg / g (86.36) groups were significantly higher than the control group. Other indicators such as egg weight, albumen height, eggshell strength, eggshell thickness, egg shape index, eggshell weight, eggshell ratio, yolk weight, yolk ratio, and eggshell color L, a, b* values ​​showed no significant differences among the groups. p >0.05). This indicates that low doses of fishmeal (1 mg / g) reduce yolk color and Haugh units, while medium to high doses (2-6 mg / g) improve yolk color and Haugh units.

[0054] Table 12 Effects of different levels of fishmeal supplementation in diet on egg quality

[0055] To investigate the effects of different levels of fishmeal (1, 2, 4, 6 mg / g DHA) on the quality of eggs after 21 days of storage.

[0056] As shown in Table 13, there were significant differences in eggshell strength among the groups (P=0.023), with the 2 mg / g group (5.21 kg / cm²) being significantly higher than the control group (4.04 kg / cm²). The proportion of eggshell increased linearly with the increase of fishmeal addition. p =0.019), the 6 mg / g group had the highest proportion of eggshell (11.82%); eggshell brightness L value (P=0.004) and eggshell redness a value ( p =0.013) also showed significant differences among groups, with the eggshell redness in the 2 mg / g and 6 mg / g groups (16.98, 17.13) being significantly higher than that in the control group (14.20); the eggshell yellowness b value ( p =0.020) with the highest value in the 2 mg / g group (27.84). There were no significant differences among the groups in egg weight, albumen height, yolk color, Haugh unit, shell thickness, egg shape index, shell weight, yolk weight, and yolk ratio. p >0.05).

[0057] Table 13 Effects of different levels of fishmeal supplementation in diet on egg quality after 21 days of storage.

[0058] To investigate the effects of adding different levels of fishmeal (1, 2, 4, 6 mg / g DHA) to the diet on the quality of eggs after 42 days of storage.

[0059] As shown in Table 14, the protein content in each fishmeal-added group was significantly lower than that in the control group. p=0.0010), with the 4 mg / g group having the lowest (4.06 mm), while the control group had 5.63 mm; there was a highly significant difference in eggshell redness a value ( p =0.0005), the 4 mg / g group (12.63) was significantly lower than the control group (16.36), while the 1, 2, and 6 mg / g groups (16.57, 16.05, 16.45) showed no significant difference from the control group; although the Haugh unit decreased to 64.24 in the 4 mg / g group, the difference between the groups did not reach the level of statistical significance. p =0.0798). Other indicators such as egg weight, yolk color, eggshell strength, eggshell thickness, egg shape index, eggshell weight, yolk weight, yolk ratio, and eggshell L value and b value showed no significant differences among the groups. p >0.05).

[0060] Table 14 Effects of different levels of fishmeal supplementation in diet on egg quality after 42 days of storage.

[0061] To investigate the effects of different levels of fishmeal (1, 2, 4, 6 mg / g DHA) on the fatty acid composition of egg yolk.

[0062] As shown in Table 15, fishmeal addition had no significant effect on the content of saturated fatty acids (SFA) and monounsaturated fatty acids (MUFA). p >0.05), but significantly increased the content of polyunsaturated fatty acids (PUFAs). p =0.0285). With the increase of fishmeal addition, the contents of n-3 PUFA and DHA both increased significantly ( p <0.0001), the DHA content in the 6 mg / g group reached 18.81 mg / g, and the DHA enrichment was 854.56 mg / 100g egg yolk; the n-6 PUFA content fluctuated in each supplementation group, with the 1 mg / g and 4 mg / g groups being significantly higher than the control group ( p =0.0069); the n-6 / n-3 ratio decreased significantly from 14.47 in the control group to 3.73 ( p <0.0001). This indicates that fishmeal as a DHA source can effectively increase the DHA enrichment in egg yolks and optimize the fatty acid composition, but the enrichment efficiency is lower than that of algae powder and algae oil at the same added level.

[0063] Table 15 Effects of different levels of fishmeal supplementation in diet on fatty acid composition of egg yolk

[0064] Example 2 The effects of microencapsulated DHA on bioavailability and the expression of related genes in the liver of laying hens were investigated using the following steps: One hundred and ninety-two Hy-Line Brown laying hens of good condition and similar egg production rates at 30 weeks of age were randomly divided into four groups, with six replicates per group and eight hens per replicate. Microencapsulated Schizochytrium powder was added to the diets at concentrations of 0, 2, 4, and 8 mg / g DHA, respectively. The experiment lasted for eight weeks. Egg yolk DHA content (mg / g), DHA conversion rate, liver gene expression, FABP, and PCTP were measured.

[0065] As shown in Table 16, there were no significant differences in egg production rate and egg quality between the unmodified algal powder group and the control group. However, in the algal oil group and fish meal group, egg production rate decreased significantly, and eggshell strength and yolk color also decreased significantly when DHA levels were higher than 4 mg / g. The bioavailability of DHA in the microencapsulated algal powder group was 6%-10% higher than that in the unmodified algal powder group, and the expression levels of FABP and PCTP in the liver of laying hens were significantly increased. Therefore, microencapsulated Schizochypterygium powder was selected as the core DHA source, with an optimal addition level of 2-4 mg / g.

[0066] Table 16 Effects of dietary supplementation with microencapsulated DHA on DHA bioavailability and expression of related genes in the liver of laying hens

[0067] Example 3 The specific steps for investigating the effect of soybean lecithin on DHA enrichment efficiency are as follows: Three hundred and sixty Hy-Line Brown laying hens aged 30 weeks in good condition and with similar egg production rates were randomly divided into nine groups, with six replicates per group and ten hens per replicate. The control group was fed a basal diet, while the experimental groups were supplemented with 2 mg / g microencapsulated algae powder DHA. The following groups were also included: ① Ordinary soybean lecithin group (phosphatidylcholine content 50%): supplemented with 1% and 2%; ② High-purity soybean lecithin group (phosphatidylcholine content ≥70%): supplemented with 1% and 2%; ③ Blank control group: supplemented only with the basal diet. The experiment lasted for eight weeks. The DHA content and enrichment efficiency in eggs were measured, as well as the expression level of PCTP in the ovaries and oviducts of the laying hens and the DHA binding form in the yolk.

[0068] As shown in Table 17, phosphatidylcholine content is a key factor affecting DHA enrichment efficiency. The DHA enrichment efficiency of the high-purity soybean lecithin group was significantly higher than that of the ordinary soybean lecithin group with the same amount of added lecithin. The DHA content in the eggs of the experimental group with 1% high-purity soybean lecithin was 30% higher than that of the control group, and the DHA enrichment efficiency reached 65.25%. The DHA content in the experimental group with 2% high-purity soybean lecithin was 45% higher, and the enrichment efficiency reached 70.61%. Moreover, the proportion of DHA in the yolk in the form of phospholipid complex was more than 80%. The expression level of PCTP in the ovary and oviduct of laying hens was 2.0-2.5 times higher than that of the control group and 0.28-0.73 times higher than that of the ordinary phospholipid group. The expression level of FABP in the liver, oviduct and ovary was 1.6-2.3 times higher than that of the control group and 0.32-0.72 times higher than that of the ordinary phospholipid group. Therefore, the optimal addition amount of soybean lecithin is 1%-2% of the total mass of the premix, and it is preferred to use a high-purity product with a phosphatidylcholine content of ≥70%.

[0069] Table 17 Effects of soybean lecithin of different purities on DHA enrichment and PCTP expression in egg yolk

[0070] Example 4 The specific steps for investigating the effect of astaxanthin on the shelf life of eggs are as follows: One hundred and eighty Hy-Line Brown hens aged 30 weeks in good condition and with similar egg production rates were randomly divided into three groups, with six replicates per group and ten hens per replicate. The control group was fed a basal diet, while experimental group 1 was fed a basal diet plus algal powder DHA (2 mg / g) and high-purity soybean lecithin (1%). Experimental groups 2 and 3 were fed the same diet as experimental group 1, with the addition of 0.05% and 0.10% astaxanthin, respectively. The experiment lasted for eight weeks. Eggs were collected and stored at 4°C, and Haugh units, yolk index, and other indicators were measured periodically.

[0071] As shown in Table 18, the SOD and GSH-Px activities of group 2 (with added astaxanthin) at each time point were significantly higher than those of group 1 and the control group, while the MDA content was significantly lower. This indicates that astaxanthin can achieve antioxidant effects by activating the antioxidant enzyme system and directly scavenging free radicals. Group 1, which only added DHA and lecithin, had a high degree of DHA oxidation; after 45 days of storage, the Huff unit was below 60 and the yolk index was below 0.25, rendering it commercially worthless. In contrast, groups 2 and 3, after 50 days of storage, maintained Huff units of 62.76-68.55 and a yolk index of 25.38-24.66%, far superior to groups 1 and the control group. Egg weight loss was reduced by more than 36% compared to group 1, yolk pH decreased by 0.87, and shelf life was extended by 15-20 days under 4℃ storage conditions compared to the control group. Furthermore, the DHA oxidation inhibition efficiency reached over 75%. Therefore, the optimal addition amount of astaxanthin is 0.05% of the total mass of the premix, which can form a good synergistic antioxidant effect with DHA.

[0072] Table 18 Effects of dietary astaxanthin supplementation on egg quality and antioxidant indicators during storage

[0073] Time-series observation of antioxidant indicators in the livers of laying hens during the experiment revealed that the activities of SOD and GSH-Px in the astaxanthin-supplemented group significantly increased with prolonged feeding time (P<0.05), reaching a peak at week 8 and then stabilizing; while the MDA content significantly decreased with prolonged feeding time (P<0.05). In terms of dose-response, SOD and GSH-Px activities showed a significant increase followed by a plateauing trend with increasing astaxanthin dosage, while MDA content showed an initial decrease followed by a plateauing trend. These results indicate that astaxanthin can activate the endogenous antioxidant enzyme system in laying hens in a time-dependent manner, while simultaneously reducing lipid peroxide content in a dose-dependent manner, achieving a dual antioxidant effect. The 0.05% astaxanthin supplementation group was the optimal dosage (Table 19).

[0074] Table 19 Effects of different doses of astaxanthin on antioxidant indices in the liver of laying hens during the experiment.

[0075] Example 5 The effects of functional ingredient blending and microencapsulation processes on the overall performance of premixes were investigated. The specific steps are as follows: Three hundred and sixty Hy-Line Brown hens aged 30 weeks in good condition and with similar egg production rates were randomly divided into six groups, with six replicates per group and ten hens per replicate. The control group was fed a basal diet. Experimental group 1 was fed a basal diet plus microencapsulated core components [algal powder DHA (2 mg / g DHA) + high-purity soybean lecithin (1%) + astaxanthin (0.05%)]. Experimental groups 2-4 were fed an additional compound probiotic (1 × 10⁻⁶) on top of the diet of experimental group 1. 9 CFU / kg), lutein (30mg / kg) + organic selenium (0.2mg / kg), compound probiotics (1×10 9 The experimental groups were treated with a combination of CFU / kg, lutein (30 mg / kg), and organic selenium (0.2 mg / kg). In addition to the control group, the five experimental groups were supplemented with compound probiotics (1×10⁻⁶ CFU / kg). 9 The study included CFU / kg of lutein (30 mg / kg) and organic selenium (0.2 mg / kg). The experiment lasted 8 weeks, assessing egg production performance, gut microbiota diversity, DHA digestibility and absorption, and yolk nutritional composition in laying hens. The effects of each component on DHA accumulation and production performance were compared.

[0076] The results showed that the egg production rate and DHA enrichment of the laying hens in experimental group 2 (microencapsulated composite encapsulation) were 5%-8% higher than those in experimental group 1 (unencapsulated), and the storage stability of eggs was significantly improved. The intestinal flora diversity of the laying hens in experimental group 3 (compound functional ingredients) was more than 20% higher than that in experimental group 2, with Bacillus subtilis and Lactobacillus plantarum becoming the dominant flora. The digestibility and absorption rate of feed by the laying hens increased by 10%, and nitrogen emissions in feces decreased by 18%. In addition to DHA, the lutein content in the egg yolk reached 8.5 mg / 100g, and the selenium content reached 0.32 mg / 100g.

[0077] The combination of functional components significantly improved the egg production rate and DHA accumulation in egg yolks of laying hens, confirming that probiotics, lutein, and organic selenium had no antagonistic effect with the core components. In Experiment 2, the DHA digestibility was 14.3% higher than in Experiment 1, the gut microbiota diversity index was significantly increased, and the rate of reduction in fecal nitrogen emissions was significantly increased, confirming that probiotics indirectly promote DHA accumulation by improving the gut health of laying hens and enhancing the intestinal digestibility and absorption of DHA. In Experiment 3, the egg production rate and DHA accumulation were slightly higher than in Experiment 1, confirming that lutein and selenium have a slight positive promoting effect on laying hen performance and DHA accumulation, with no adverse effects. In Experiment 4, all indicators were optimal, with egg production rate, DHA accumulation rate, and DHA accumulation in egg yolks all significantly higher than other groups, confirming the synergistic effect of the functional components, achieving a positive cycle of "gut regulation - nutrient absorption - efficient accumulation".

[0078] Table 20 Effects of functional component compounding and microencapsulation process on egg production performance and yolk nutrition in laying hens

[0079] Example 6 The specific steps for constructing a parameter system linking "DHA enrichment - egg quality - shelf life" are as follows: Using the amount of Schizochytrium powder added (X1, mg / g), the proportion of high-purity soybean lecithin added (X2, %), and the proportion of astaxanthin added (X3, ‰) as independent variables, and the amount of DHA enriched in egg yolk (Y1, mg / 100g), the egg production rate of laying hens (Y2, %), and the shelf life at 4℃ (Y3, d) as dependent variables, SPSS 26.0 was used to conduct univariate linear regression and multivariate stepwise regression analysis to construct a mathematical model of linkage parameters, clarify the quantitative relationship between each variable and the optimal threshold.

[0080] (1) Univariate linear regression analysis (the effect of a single factor on the dependent variable) a: Y1=218.65X1+89.32, R²=0.926, P<0.0001, indicating that the amount of Schizochytrium powder added is strongly linearly positively correlated with the amount of DHA enriched in egg yolk. When X1 is 2-4 mg / g, Y1 is in the optimal range of 500-1000 mg / 100g. b: Y1=156.82X2+725.36, R²=0.895, P<0.0001, indicating that the lecithin addition ratio is strongly linearly positively correlated with the DHA enrichment, and the increase of Y1 is most significant when X2 is 1%-2%; c: Y3=385.62X3+30.85, R²=0.918, P<0.0001, indicating that the astaxanthin addition ratio has a strong linear positive correlation with the shelf life. When X3 is 0.5‰ (0.05%), Y3 reaches more than 50 days. d: Y2 showed no significant linear negative correlation with X1, X2, and X3 (P>0.05), indicating that within the optimal addition range, the amount of core component added had no significant adverse effect on the egg production rate, and Y2 remained stable above 85%.

[0081] (2) Multiple stepwise regression analysis (the combined effect of multiple factors on the dependent variable) Core regression model: Comprehensive model of DHA enrichment in egg yolks: Y1=205.36X1+148.62X2+45.89X3+68.52, R²=0.958, p <0.0001 Model explanation: The amount of Schizochytrium powder added contributed the most to the enrichment of DHA, followed by lecithin. Astaxanthin had a slight positive promoting effect on the enrichment of DHA. The synergistic effect of the three factors can significantly improve the model fit. Comprehensive model for egg shelf life: Y3 = 25.68X1 + 32.56X2 + 378.95X3 + 28.65, R² = 0.942. p <0.0001 Model explanation: Astaxanthin contributes more than 85% to shelf life, while the addition of DHA and lecithin has a slight impact on shelf life, forming a synergistic preservation effect with astaxanthin; Comprehensive model for egg production rate of laying hens: Y2 = -5.23X1 + 3.86X2 - 2.58X3 + 98.65, R² = 0.825. p <0.001 Model interpretation: Within the optimal range (X1=2-4, X2=1-2, X3=0.5), the calculated value of Y2 is 89.5%-94.8%, which is consistent with the actual experimental value, confirming that the core component has no significant inhibitory effect on egg production rate within the optimal range.

[0082] (3) Determination of the optimal threshold of the linkage parameter system Based on the regression model and actual experimental data, the core thresholds of the linkage parameter system were determined: Compound probiotics of Bacillus subtilis and Lactobacillus plantarum (total live bacteria ≥ 1×10⁻⁶) 9 The addition of CFU / kg regulates the intestinal flora structure of laying hens, improves the digestion and absorption efficiency of DHA and lecithin, and improves intestinal health; the addition of lutein (20-50mg / kg) and organic selenium (0.1-0.3mg / kg) achieves compound nutritional fortification of "DHA + lutein + selenium", which enhances the nutritional value of eggs. Moreover, the added ingredients have no antagonistic effect with the core components and do not affect the egg production performance of laying hens or the efficiency of DHA enrichment.

[0083] Table 21 Optimal addition amount and target threshold of core components

[0084] The above are all preferred embodiments of the present invention and are not intended to limit the scope of protection of the present invention. Therefore, all equivalent changes made in accordance with the structure, shape and principle of the present invention should be covered within the scope of protection of the present invention.

Claims

1. A premixed feed for laying hens that combines efficient DHA enrichment with egg preservation, characterized in that, It consists of a core active component and a functional compound component; the core active component is microencapsulated Schizochytrium powder, high-purity soybean lecithin and astaxanthin, and the functional compound component is compound probiotics, lutein and organic selenium; based on the total mass of laying hen compound feed, the DHA content provided by microencapsulated Schizochytrium powder is 2~4 mg / g, the addition amount of high-purity soybean lecithin is 1%~2%, and the addition amount of astaxanthin is 0.05%.

2. The laying hen premix according to claim 1, characterized in that, The high-purity soybean lecithin contains ≥70% phosphatidylcholine.

3. The laying hen premix according to claim 1, characterized in that, The compound probiotics consist of Bacillus subtilis and Lactobacillus plantarum; based on the total mass of the laying hen compound feed, the total live bacteria count of the compound probiotics is ≥1×10⁻⁶. 9 CFU / kg.

4. The laying hen premix according to claim 1, characterized in that, Based on the total weight of the compound feed for laying hens, the amount of lutein added should be 20-50 mg / kg, and the amount of organic selenium added should be 0.1-0.3 mg / kg.

5. The laying hen premix according to claim 4, characterized in that, Based on the total mass of the compound feed for laying hens, the amount of lutein added is 30 mg / kg, and the amount of organic selenium added is 0.2 mg / kg.

6. The use of the premixed feed for laying hens according to any one of claims 1-5 in the preparation of compound feed for laying hens during the laying period.

7. A method for producing eggs that combine efficient DHA enrichment and preservation functions, characterized in that, Add the premixed feed for laying hens as described in any one of claims 1-5 to the basal diet of laying hens aged 30 weeks and above, and feed continuously for at least 2 weeks.

8. The production method according to claim 7, characterized in that, The laying hens mentioned are Hy-Line Brown laying hens.

9. The production method according to claim 7, characterized in that, After feeding, the DHA enrichment in egg yolks reaches 500~1000 mg / 100 g, and the egg production rate of laying hens remains above 85% throughout the entire laying process.

10. The production method according to claim 7, characterized in that, The resulting eggs had a shelf life extended by 15-20 days when stored at 4°C, with a DHA oxidation inhibition rate exceeding 75% and an egg weight loss rate reduced by 36%.

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