Application of thiamine in improving sugar utilization rate and growth performance of sturgeons

Abstract

The invention discloses application of thiamine in improving the sugar utilization rate and growth performance of sturgeons, and belongs to the field of aquaculture. It is proved for the first time that by enhancing a glycolytic pathway, inhibiting a gluconeogenesis pathway and promoting glycogen decomposition, the thiamine effectively improves the sugar metabolism capacity of sturgeons and reduces hepatic glycogen deposition and fat accumulation, so that the risk of metabolic disorder is reduced. Meanwhile, thiamine also promotes muscle fiber growth, so that the growth speed of the sturgeons and the feed utilization rate are remarkably increased. Therefore, the thiamine can be used as a feed additive for preparing the feed for improving the sugar utilization rate and the growth performance of the sturgeons. A safe and efficient nutrition regulation and control scheme is provided for sturgeon culture, the feed utilization rate and culture benefits are improved, meanwhile, ecological risks such as antibiotic abuse are avoided, and remarkable economic benefits and application prospects are achieved.

Description

Technical Field

[0001] This invention relates to the field of aquaculture, and in particular to the application of thiamine in improving the utilization of sugar and growth performance of sturgeon. Background Technology

[0002] Sturgeon, as an important economic fish species, often faces the problem of low carbohydrate metabolism efficiency during its aquaculture, resulting in poor feed utilization and limited growth performance. Current technologies often reduce costs by adding carbohydrates to fish feed; however, carnivorous fish like sturgeon have poor carbohydrate utilization capabilities, and excessive intake can easily lead to metabolic disorders such as glycogen deposition and fatty liver, thus affecting their health and growth. Currently, research on improving carbohydrate metabolism in fish mainly focuses on optimizing feed formulations or adding conventional growth promoters (such as yeast extracts and probiotics), but these methods have limited effectiveness and may lead to antibiotic overuse or ecological safety risks.

[0003] Although previous studies have shown that thiamine (vitamin B1) can affect carbohydrate metabolism in shrimp, the significant physiological differences between fish and shrimp, and the varying thiamine requirements and metabolic responses among different fish species, mean that this finding cannot be directly applied to sturgeon farming. Furthermore, the specific mechanism of action of thiamine in regulating sturgeon carbohydrate metabolism and the optimal dosage remain unclear, and current technologies lack precise nutritional regulation programs for sturgeon. Therefore, developing a highly efficient thiamine-based method for regulating sturgeon carbohydrate metabolism is of great significance for improving feed utilization and aquaculture efficiency. Summary of the Invention

[0004] The purpose of this invention is to provide an application of thiamine in improving the utilization rate of sugar and growth performance of sturgeon, thereby solving the problems existing in the prior art. This invention significantly improves the utilization rate of sugar and growth performance of sturgeon by adding a specific dose of thiamine (3.2-4.4 mg / kg) to sturgeon feed, providing a safe and efficient nutritional regulation scheme for sturgeon farming. This helps to improve feed utilization and farming efficiency, while avoiding ecological risks such as antibiotic overuse, and has significant economic benefits and application prospects.

[0005] To achieve the above objectives, the present invention provides the following solution:

[0006] This invention provides the application of thiamine in the preparation of feed additives that improve the utilization of sugar in sturgeon.

[0007] This invention also provides the application of thiamine as a feed additive in the preparation of feed that improves the utilization rate of sugar in sturgeon.

[0008] Furthermore, the thiamine content in the feed is 3.2-4.4 mg / kg.

[0009] Furthermore, thiamine enhances the utilization of sugar by promoting sugar metabolism in sturgeon.

[0010] Furthermore, the promotion of sturgeon's sugar metabolism includes enhancing glycolysis pathways, inhibiting gluconeogenesis pathways, and promoting glycogenolysis.

[0011] This invention also provides the application of thiamine in the preparation of feed additives that improve the growth performance of sturgeon.

[0012] This invention also provides the application of thiamine as a feed additive in the preparation of feed to improve the growth performance of sturgeon.

[0013] Furthermore, the thiamine content in the feed is 3.2-4.4 mg / kg.

[0014] Furthermore, the improvement of sturgeon growth performance includes enhancing the fish's ability to utilize sugar, reducing fat accumulation, and promoting muscle fiber growth.

[0015] The present invention discloses the following technical effects:

[0016] This invention significantly improves the utilization of glucose and growth performance in sturgeon by adding a specific dose of thiamine (3.2-4.4 mg / kg) to their feed. Specifically, thiamine effectively improves the sturgeon's glucose metabolism by enhancing glycolysis, inhibiting gluconeogenesis, and promoting glycogenolysis, thereby reducing liver glycogen deposition and fat accumulation, and thus lowering the risk of metabolic disorders. Simultaneously, thiamine also significantly improves the growth rate and feed utilization of sturgeon by promoting muscle fiber growth. The experimental group supplemented with 3.2 mg / kg thiamine achieved the highest final body weight and specific growth rate, and the lowest feed conversion ratio. Furthermore, the addition of thiamine also optimized fat metabolism, reducing triglyceride levels in the liver and serum, further improving the overall health of the fish.

[0017] This invention provides a safe and efficient nutritional regulation scheme for sturgeon farming, which helps to improve feed utilization and farming efficiency, while avoiding ecological risks such as antibiotic abuse, and has significant economic benefits and application prospects. Attached Figure Description

[0018] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0019] Figure 1 Quadratic regression analysis of sturgeon final body weight and feed thiamine supplementation level;

[0020] Figure 2 The effects of different amounts of thiamine in the feed on liver glycogen content (A) and expression of genes related to glycogen synthesis and degradation (B) in juvenile sturgeon;

[0021] Figure 3 The effects of different amounts of thiamine in feed on serum glucose levels (A) and liver glucose transporter 2 (glut2) gene expression (B) in juvenile sturgeon;

[0022] Figure 4 The effect of different amounts of thiamine in feed on muscle glycogen content in juvenile sturgeon;

[0023] Figure 5 The effects of different amounts of thiamine in the feed on the activities of hepatic hexokinase (A), hepatic pyruvate kinase (B), and hepatic pyruvate content (C) in the liver glycolysis pathway of juvenile sturgeon;

[0024] Figure 6 The effects of different amounts of thiamine in feed on the expression of key enzyme genes in the liver glycolysis pathway of juvenile sturgeon;

[0025] Figure 7 The effects of different amounts of thiamine in feed on the expression of key enzyme genes in the hepatic gluconeogenesis pathway in juvenile sturgeon;

[0026] Figure 8 The effect of different amounts of thiamine in feed on the activity of transketolase in the liver pentose phosphate pathway of juvenile sturgeon;

[0027] Figure 9 The effects of different amounts of thiamine in feed on liver triglyceride (A) and serum triglyceride (B) levels in juvenile sturgeon;

[0028] Figure 10 The effects of different amounts of thiamine in feed on the expression of genes related to liver fat synthesis in juvenile sturgeon;

[0029] Figure 11 The effects of different amounts of thiamine in feed on the expression of genes related to liver fat breakdown in juvenile sturgeon;

[0030] Figure 12 The effects of different amounts of thiamine in feed on the serum total cholesterol (A) and liver total cholesterol (B) levels of juvenile sturgeon;

[0031] Figure 13 Images showing PAS staining of liver glycogen in juvenile sturgeon at different levels of thiamine in feed; scale bar: 100 μm.

[0032] Figure 14H&E staining images showing the effects of different thiamine contents in feed on cross-sections of juvenile sturgeon muscle; scale bar: 100 μm;

[0033] Figure 15 The effect of different amounts of thiamine in the feed on the muscle fibers of juvenile sturgeon was investigated, including (A) the average diameter of the muscle fibers; and (B) the frequency of diameters <20 μm, 20-50 μm, and >50 μm. Detailed Implementation

[0034] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0035] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Any stated value or intermediate value within a stated range, as well as each smaller range between any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0036] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0037] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be obvious to those skilled in the art. This specification and embodiments are merely exemplary.

[0038] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0039] Example

[0040] 1. Materials and Methods

[0041] 1.1 Experimental Fish

[0042] Three hundred healthy juvenile Western sturgeon (Acipenser baerii♀ × A. schrenckii♂) with an average weight of approximately 10g were purchased from Jingxing Luyuan Fishery Co., Ltd. and transported at low temperature to the aquaculture room of the College of Life Sciences, Hebei Normal University. After balancing the water temperature, they were disinfected with 25.6g / L saline solution and temporarily raised in the aquaculture system to adapt to the aquaculture environment and feeding rhythm. During the acclimatization period, they were fed commercial feed twice a day (09:00 and 16:00) until they were full. After two weeks of acclimatization, fish with an average weight of 14.63±0.8g were randomly selected for a formal feeding experiment.

[0043] 1.2 Experimental Design and Materials

[0044] Five isonitrogenous (40.56% crude protein) and isolipid (11.55% crude fat) feed formulations were prepared, and five doses (0, 0.8, 1.6, 3.2, and 6.4 mg / kg) of thiamine nitrate were added to each of the five experimental feeds. To balance the feed formulations, the microcrystalline cellulose content was varied by an equal amount, while other components remained constant. Vietnamese fish meal, chicken meal, and squid liver meal were used as animal protein sources; extruded soybean, cottonseed protein, wheat gluten, and rapeseed meal were used as plant protein sources; brewer's yeast powder was used as a single-cell protein source; pregelatinized corn starch was used as the main carbohydrate source; and soybean oil, extruded soybean, and soybean lecithin were used as the main fat sources.

[0045] Weigh all ingredients except soybean oil according to the formula, grind and mix thoroughly until the powder passes through an 80-mesh sieve. Then, thoroughly mix the soybean oil into the powder. Finally, add approximately 23% double-distilled water to the mixture and mix thoroughly using a mixer to form a soft dough. Process the dough into pellets with a diameter of 2 mm and a length of 3 mm using a laboratory feed mill (Shandong Youyi Machinery Factory). Dry the feed pellets in a 50℃ oven and store them in resealable bags at -20℃. The approximate composition of the feed formula and dry matter basis is shown in Table 1.

[0046] Table 1. Experimental feed formulation and basic nutrient composition (%, dry matter basis)

[0047]

[0048]

[0049] Note: 1 kg of premix contains the following vitamins and minerals: Vitamin A acetate, 11.2 mg; Vitamin D3, 1.68 mg; Vitamin E, 2000 mg; Nicotinamide menadione bisulfite (MSN), 982.8 mg; Riboflavin, 240 mg; Pyridoxine hydrochloride, 2400 mg; Cyanocobalamin, 2 mg; Inositol, 13444 mg; Calcium D-pantothenate, 400 mg; Niacin, 1200 mg; Folic acid, 140 mg; Biotin, 20 mg; Ascorbic acid 8000mg; CuSO4·5H2O, 1670.4mg; FeSO4·7H2O, 20000mg; MnSO4·H2O, 1647.27mg; ZnSO4·7H2O, 5317mg; CoCl2·6H2O, 240.03mg; KI, 235.3mg; Na2SeO3, 90.13mg; butylated hydroxytoluene, 25g; talc, 240g, with the remainder diluted with corn gluten powder.

[0050] 1.3 Aquaculture Management

[0051] After 14 days of acclimatization and a 24-hour fast, 225 healthy juvenile sturgeon of similar weight (14.63±0.8g) were randomly selected and randomly assigned to 15 white rectangular plastic tanks (length×width×height: 80cm×60.48cm×65cm), with 15 sturgeon per tank. Each tank was equipped with an air stone for continuous aeration and a water inlet for dechlorinated tap water. Each feed treatment group was randomly assigned to three tanks for artificial saturation feeding twice daily (09:00 and 16:00) for 8 weeks. The water was changed twice daily, with 2 / 3 of the water replaced. After each feeding, the remaining feed pellets were siphoned out and placed in a mesh screen, dried in an oven at 105℃, pulverized into powder, and the moisture content was measured to correct for the actual dry matter intake and to measure feed loss rate. During the rearing period, water temperature was monitored daily, and water quality was measured weekly. The conditions during the breeding period are: 12L / 12D light / dark, water temperature 17-21℃, pH 8.2-8.6, ammonia nitrogen <0.2mg / L, nitrite 0.01-0.05mg / L, residual chlorine 0.025mg / L, and dissolved oxygen 5-6mg / L.

[0052] 1.4 Sample Collection

[0053] At the start of the culture experiment, 15 fish were randomly netted and divided into 3 replicates of 5 fish each to determine initial body composition. After a 24-hour fast, all fish were counted and weighed to calculate growth-related parameters, including feed intake rate (FR), final body weight (FBW), weight gain rate (WGR), specific growth rate (SGR), feed conversion ratio (FCR), and protein efficiency (PER). Subsequently, all fish in each tank were captured and anesthetized with eugenol (60 mg / L). Three fish were randomly selected from each tank, gently wiped, and weighed to determine final body composition. Then, five fish were randomly selected from each tank, and their body weight and length (from snout to base of the caudal fin) were measured. Blood was drawn from the tail vein using a 1 mL syringe without heparin sodium, placed in a 2 mL centrifuge tube, stored at 4°C for 3 hours, centrifuged (4000 g, 15 min, 4°C), and the serum sample was collected and stored at -80°C for the determination of serum glucose, triglyceride, and total cholesterol levels. The livers were then dissected on an ice tray and divided into three parts. A small piece (approximately 1 cm³) was fixed in 4% paraformaldehyde for PAS staining. Another portion of the liver was flash-frozen in liquid nitrogen and then stored at -80°C to measure liver metabolites and key enzyme activities, including triglycerides, total cholesterol, glycogen, pyruvate, hexokinase, and transketolase. The last portion was used to measure the mRNA expression levels of key enzymes and transcription factors involved in liver glucose and lipid metabolism. Finally, the dorsal muscles from these five fish were dissected for muscle glycogen measurement.

[0054] 1.5 Biochemical Analysis

[0055] Nutrient composition in feed and fish was determined according to AOAC (2023) standard methods. Moisture content was determined by direct drying at 105℃; crude protein content was determined by Kjeldahl method; crude fat content was determined by petroleum ether extraction; and ash content was determined by calcination at 550℃ for 3 h in a muffle furnace. Serum glucose, triglycerides, and total cholesterol, as well as liver triglycerides, total cholesterol, glycogen, pyruvate, and hexokinase were measured using Nanjing Jiancheng reagent kits (Nanjing Jiancheng Bioengineering Research Institute Co., Ltd.), and transketolase was measured using Shanghai Guduo reagent kits (Shanghai Guduo Biotechnology Co., Ltd.).

[0056] 1.6 Liver Histological Analysis

[0057] Three fixed liver samples were selected from each treatment group and sent to Wuhan Saiwell Biotechnology Co., Ltd. for PAS staining. Liver sections were prepared according to standard technical methods and observed using a Leica 3D full-view fluorescence microscopy system (Leica DFC 9000) to observe the status of liver glycogen in liver cells.

[0058] 1.7 Total RNA extraction, reverse transcription, and qPCR

[0059] Total RNA from the liver was extracted using TransZol Up reagent. RNA purity was determined by absorbance at 260 nm using a micro-spectrophotometer, and RNA integrity was checked by agarose gel electrophoresis. Reverse transcription from RNA to cDNA was performed using the TransZol Up One-Step gDNA Removal and cDNA Synthesis SuperMix kit. The purity and concentration of cDNA were determined using a micro-spectrophotometer. All cDNA groups were diluted to the same concentration (200 ng / μL) with DEPC water, aliquoted, and stored at -80°C. Designed primers and cDNA were used for conventional PCR. The conventional PCR program was as follows: 95°C for 3 min, (95°C for 30 sec, 50–60°C for 30 sec, 72°C for 30 sec) for 35 cycles, then 72°C for 7 min. Subsequently, 1.5% agarose gel electrophoresis was performed to verify primer specificity (the target band was correctly positioned and brightly clear). Qualified specific primers were used for subsequent qPCR amplification.

[0060] qPCR was performed using the Beijing TransGen Top Green qPCR SuperMix kit. The stability of three internal controls—β-actin, EF-1α, and 18S-rRNA—was compared. EF-1α was used as the liver control to detect the mRNA expression levels of genes related to glucose and lipid metabolism. The qPCR reaction employed a two-step method: 95℃ for 30 seconds, followed by 40 cycles of 95℃ for 5 seconds and then 55℃–60℃ for 30 seconds. Melting curve analysis was then performed at 65℃–95℃. A single peak in the melting curve ensured specific amplification. -△△ct Calculate the expression level of the target gene. The sequences and primer information of the internal reference and target gene are shown in Table 2.

[0061] Table 2 Primer Information for Real-Time Quantitative PCR

[0062]

[0063]

[0064] Note: GS, glycogen synthase; GP, glycogen phosphorylase; glut2, glucose transporter 2; PFK, phosphofructokinase; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; PK, pyruvate kinase; FBPase, fructose-1,6-bisphosphatase; G6Pase, glucose-6-phosphatase; chrebp, carbohydrate response element-binding protein; FAS, fatty acid synthase; ACC, acetyl-CoA carboxylase; HSL, hormone-sensitive lipase; EF-1α, eukaryotic elongation factor.

[0065] 1.8 Calculation of relevant formulas

[0066] Survival rate (SR, %) = 100 × number of terminal tails / number of initial tails;

[0067] Weight gain rate (WGR, %) = 100 × (final weight – initial weight) / initial weight;

[0068] Feed conversion ratio (FCR) = Total feed intake (dry weight) / (Final weight – Initial weight);

[0069] Specific growth rate (SGR, % / d) = 100 × (Ln final weight – Ln initial weight) / number of days;

[0070] Average feed intake (FI, g / tail) = Average dry weight of feed / number of tails;

[0071] Protein efficiency (PER, %) = (Final weight – Initial weight) / (Average dry weight fed × Protein content in feed);

[0072] Protein deposition rate (PDR, %) = (Total dry weight of fish at the end of the experiment × Crude protein content of fish at the end of the experiment – ​​Total dry weight of fish at the beginning of the experiment × Protein content of fish at the beginning of the experiment) / (Total dry weight of feed consumed × Crude protein content of feed) × 100.

[0073] 1.9 Data Analysis

[0074] All data are expressed as mean ± standard deviation (mean ± SD). The Shapiro-Wilk test was used to determine the normality of the data, and Levene's test was used to determine the homogeneity of variance. One-way ANOVA and Student-Newman-Keuls multiple comparisons were performed using SPSS 20.0 software. A p-value < 0.05 was considered statistically significant.

[0075] 2. Results

[0076] 2.1 Growth performance, feed utilization and body composition

[0077] The effects of different thiamine contents on the growth performance and feed utilization of juvenile sturgeon are shown in Table 3. As the table shows, no juvenile sturgeon died during the entire rearing period. The final body weight and weight gain rate of the thiamine-supplemented feed group were significantly higher than those of the control group (P<0.05), and these two indicators generally showed an initial upward and then downward trend with increasing thiamine levels, reaching their maximum in group D3.2. There were no significant differences in specific growth rate, feed conversion ratio, protein deposition rate, and protein efficiency among the groups (P>0.05), but specific growth rate, protein deposition rate, and protein efficiency were highest in group D3.2, while feed conversion ratio was lowest in group D3.2. Additionally, as... Figure 1As shown, using final body weight as the evaluation index, regression analysis revealed that the optimal level of thiamine supplementation in juvenile sturgeon feed was 4.40 mg / kg.

[0078] Table 3. Effects of experimental diets with different thiamine contents on growth performance, feed utilization, and body morphology of hybrid sturgeon juveniles.

[0079]

[0080] Note: Data in the table are expressed as mean ± standard deviation. Tukey's method was used for post-hoc multiple comparisons. Different superscript letters in the same row indicate significant differences (P<0.05). The number of replicates was 3.

[0081] The effects of different thiamine contents in the diet on the body composition of juvenile sturgeon are shown in Table 4. There were no significant differences in crude protein, crude fat, and ash content among the treatment groups (P>0.05).

[0082] Table 4. Effects of experimental diets with different thiamine contents on the body composition of hybrid sturgeon juveniles.

[0083]

[0084] Note: Data in the table are expressed as mean ± standard deviation. Tukey's method was used for post-hoc multiple comparisons. Different superscript letters in the same row indicate significant differences (P<0.05). The number of replicates was 3.

[0085] 2.2 Glucose metabolism related indicators

[0086] 2.2.1 Glycogen and glucose-related indicators

[0087] Depend on Figure 2 A and Figure 13 It was found that with the increase of thiamine levels in the feed, liver glycogen content showed a trend of first decreasing and then increasing, reaching its lowest point in group D3.2. The liver glycogen content in the exogenous thiamine-supplemented groups was significantly lower than that in the control group (P<0.05). The relative transcription level of the liver glycogen synthase gene gs showed no significant difference between groups D0-D3.2, but was significantly upregulated in group D6.4 (P<0.05). Meanwhile, the relative expression level of glycogen phosphorylase gp in groups D1.6-D6.4 was significantly higher than that in the control group (P<0.05). Figure 2 (B). Therefore, the significantly lower liver glycogen levels in groups D1.6 and D3.2 compared to the control group may be due to increased glycogenolysis while glycogen synthesis remained unchanged.

[0088] Liver glycogen plays a crucial role in regulating blood glucose levels; the liver regulates blood glucose concentration by synthesizing and breaking down liver glycogen. Figure 3As shown in A, serum glucose levels reached their highest point in group D3.2 and were significantly higher than those in the control group, which was the opposite trend to the liver glycogen results (P<0.05). Simultaneously, the mRNA content of hepatic glucose transporter protein 2 (glut2) was the lowest. Figure 3 Based on B), it is speculated that group D3.2 mainly converts liver glycogen into glucose and transports it into the blood.

[0089] Muscle glycogen is also a form of glycogen storage in fish, but its overall storage content is lower than that of liver glycogen. Figure 4 It can be seen that the muscle glycogen content in group D3.2 was significantly lower than that in other treatment groups (P<0.05). In summary, the increased glycogen utilization in the liver and muscle of fish in group D3.2 is related to increased glycogenolysis, but not to glycogen synthesis.

[0090] 2.2.2 Indicators related to hepatic glycolysis and gluconeogenesis pathways

[0091] Glycolysis is a highly conserved metabolic pathway responsible for breaking down glucose into pyruvate and producing adenosine triphosphate (ATP). Hexokinase, the enzyme that catalyzes glucose phosphorylation, is the first rate-limiting enzyme in the glycolysis pathway. Pyruvate kinase, a key enzyme in glycolysis, catalyzes the reaction of phosphoenolpyruvate with adenosine diphosphate (ATP) to produce pyruvate and ATP; it is the last key enzyme in glycolysis. Pyruvate is one of the products of glycolysis, subsequently entering the mitochondria and being converted into acetyl-CoA, which then enters the tricarboxylic acid cycle. Figure 5 It was found that, at the physiological level, the liver hexokinase, pyruvate kinase activities, and pyruvate content in group D3.2 were significantly higher than those in the control group (P<0.05). Simultaneously, at the transcriptional level, the mRNA levels of phosphofructokinase (PFK), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and pyruvate kinase (PK) in group D3.2 were higher than those in the control group. Figure 6 Therefore, compared with the control group, the exogenous addition of 3.2 mg / kg thiamine significantly enhanced the hepatic glycolysis pathway.

[0092] Gluconeogenesis is the process by which simple non-carbohydrate precursors (lactic acid, glycerol, glucogenic amino acids, etc.) are converted into glucose (glucose or glycogen). Fructose-1,6-bisphosphatase (FBPase) and glucose-6-phosphatase (G6Pase) are two key enzymes in the gluconeogenesis pathway. Figure 7 It can be seen that, compared with the control group, the fbpase and g6pase of the gluconeogenesis pathway were significantly downregulated in the D0.8-D6.4 group. This shows that the exogenous addition of thiamine can enhance the glycolysis pathway and weaken the gluconeogenesis pathway.

[0093] 2.2.3 Relevant indicators of the pentose phosphate pathway

[0094] The pentose phosphate pathway is also a form of glucose oxidative breakdown, with its main products being ribose 5-phosphate (used for nucleotide synthesis) and reduced nicotinamide adenine dinucleotide phosphate (NADPH). Approximately 30% of the body's glucose enters the pentose phosphate pathway. Transketolase is a key enzyme in the pentose phosphate pathway, and thiamine pyrophosphate (TPP), the main form of thiamine, functions as a coenzyme for transketolase. Figure 8 It can be seen that the liver transketolase activity in group D3.2 was significantly higher than that in the control group (P<0.05).

[0095] 2.3 Indicators related to liver lipid metabolism

[0096] At the physiological level, by Figure 9 It was found that, compared with the control group, exogenous thiamine significantly reduced liver triglyceride levels, reaching the lowest level in group D3.2 (P<0.05). Furthermore, serum triglyceride levels in groups D1.6 and D3.2 were also significantly lower than in other treatment groups (P<0.05).

[0097] Carbohydrate response element-binding protein (chrebp) is a key transcription factor that facilitates the conversion of glucose into lipids in the body. Fatty acid synthase (fas) and acetyl-CoA carboxylase (acc) are two key enzymes in lipid synthesis. Figure 10 It can be seen that, at the molecular level, the exogenous addition of thiamine significantly downregulated the gene expression levels of the transcriptional regulator chrebp and the key lipid synthesis enzyme fas (P<0.05); compared with the control group, the gene expression level of another key lipid synthesis enzyme acc in the D3.2 group was also significantly downregulated (P<0.05).

[0098] Hormone-sensitive lipase (HSL) is a key enzyme that breaks down triglycerides into diglycerides and monoglycerides. Figure 11 It was found that, compared with the control group, the relative expression level of hsl, a key enzyme in triglycerides, was significantly upregulated in groups D1.6 and D3.2 (P<0.05). Therefore, the significant decrease in liver and blood triglycerides in group D3.2 may be due to reduced lipid synthesis and increased lipid catabolism in the liver.

[0099] Depend on Figure 12 It can be seen that the serum total cholesterol in group D3.2 was significantly lower than that in the control group, but the liver total cholesterol content was significantly higher than that in the control group (P<0.05).

[0100] 2.4 Muscle fiber related indicators

[0101] The frequency distribution of muscle fiber diameter is an indicator for assessing muscle fiber proliferation and hypertrophy, two physiological processes that jointly influence muscle growth in fish. Specifically, the frequency of muscle fibers with a diameter less than 20 μm reflects proliferation (increased number of muscle fibers), while the frequency of muscle fibers with a diameter greater than 50 μm indicates hypertrophy (increased muscle fiber volume). Figure 14 and Figure 15 It was found that the addition of thiamine did not affect the average diameter of muscle fibers, but it did affect the composition ratio of muscle fibers with different diameters. Specifically, the addition of exogenous thiamine significantly increased the frequency of muscle fibers with a diameter greater than 50 μm (P>0.05). Compared with the control group, the addition of exogenous thiamine significantly increased the frequency of muscle fibers with a diameter greater than 50 μm (P<0.05). This indicates that thiamine can promote the increase of muscle fiber volume.

[0102] In conclusion, after an 8-week aquaculture experiment, exogenous supplementation of 0.8-6.4 mg / kg thiamine significantly improved growth rate. The group supplemented with 3.2 mg / kg thiamine exhibited the highest final average weight and specific growth rate, but the lowest feed conversion ratio. Based on quadratic regression analysis using final average weight and exogenous thiamine supplementation levels, the optimal thiamine supplementation level for hybrid sturgeon juveniles was determined to be 4.4 mg / kg feed. Furthermore, the supplementation of 3.2 mg / kg thiamine significantly enhanced the fish's ability to utilize glucose, reduced fat accumulation, and promoted muscle fiber growth.

[0103] The embodiments described above are merely preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Various modifications and improvements made by those skilled in the art to the technical solutions of the present invention without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.

Claims

1. Application of thiamine in the preparation of feed additives that improve the utilization rate of sugar in sturgeon.

2. Application of thiamine as a feed additive in the preparation of feed to improve the utilization rate of sugar in sturgeon.

3. The application according to claim 2, characterized in that, The feed contains 3.2-4.4 mg / kg of thiamine.

4. The application according to claim 1 or 2, characterized in that, Thiamine enhances the utilization of sugar by promoting sugar metabolism in sturgeon.

5. The application according to claim 4, characterized in that, The promotion of glucose metabolism in sturgeon includes enhancing glycolysis, inhibiting gluconeogenesis, and promoting glycogenolysis.

6. Application of thiamine in the preparation of feed additives to improve the growth performance of sturgeon.

7. Application of thiamine as a feed additive in the preparation of feed to improve the growth performance of sturgeon.

8. The application according to claim 7, characterized in that, The feed contains 3.2-4.4 mg / kg of thiamine.

9. The application according to claim 6 or 7, characterized in that, The improvement of sturgeon growth performance includes enhancing the fish's ability to utilize sugar, reducing fat accumulation, and promoting muscle fiber growth.

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