Amino acid composition and use thereof

Abstract

The application discloses an amino acid composition and application thereof, and belongs to the technical field of biological medicines. The amino acid composition is composed of L-leucine, L-isoleucine and L-histidine at a mass ratio of 100:30:50. The amino acid formula is designed based on SIAI specificity metabolic spectrum for the first time, and amino acid imbalance is precisely corrected, so that the amino acid composition can be used for preparing medicines or nutritional preparations for improving SIAI related muscle atrophy, and has significant clinical application value.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, and in particular relates to an amino acid composition and its application. Background Technology

[0002] Severe intra-abdominal infection (SIAI) is a common critical illness in clinical practice, characterized by high morbidity and mortality. Following the onset of SIAI, the body exhibits a significant hypercatabolic state, with amino acid metabolism disorders being the most prominent. Studies have shown that the levels of various amino acids, including branched-chain amino acids (BCAAs), glutamine, and arginine, are significantly altered in SIAI patients. This metabolic disorder further exacerbates protein breakdown and negative nitrogen balance. Particularly during the first week of critical illness, patients can experience a muscle loss of up to 15%, a process known as "critical illness-related muscle atrophy." Muscle loss not only leads to decreased immune function, respiratory muscle weakness, and delayed wound healing, but is also closely related to prolonged hospital stays and increased mortality. Therefore, correcting amino acid metabolism disorders and slowing muscle atrophy are crucial for improving the prognosis of SIAI patients.

[0003] In the field of critical care nutritional support, protein and amino acid supplementation is considered a core strategy for improving clinical outcomes, and its role is even believed to surpass that of simple calorie supplementation. However, how to scientifically and precisely supplement amino acids to meet the specific metabolic needs of SIAI patients remains a pressing clinical challenge.

[0004] Currently, most amino acid formulations used in clinical practice for critically ill patients are standardized products with a "one-size-fits-all" approach. Their component design is mainly based on the average nutritional needs of healthy individuals or general critically ill patients, lacking customized adjustments for the metabolic characteristics of specific diseases (such as SIAI). Specific defects include: (1) Ignoring SIAI-specific metabolic profile changes: Existing formulations fail to incorporate research findings based on serum amino acid metabolomics of SIAI patients, and cannot effectively correct the amino acid imbalance patterns specific to this disease state (such as the relative deficiency of certain essential amino acids or the abnormal accumulation of non-essential amino acids). (2) Lack of consideration for individual differences: The intensity of inflammatory response, liver and kidney function status, and catabolism rate vary significantly among different patients, but existing products cannot achieve precise matching. (3) Poor clinical efficacy: Multiple studies have shown that the application of standard amino acid formulations in SIAI patients has failed to adequately improve adverse prognoses such as muscle loss and immunosuppression, and may even exacerbate metabolic burden due to inappropriate amino acid load.

[0005] The aforementioned deficiencies directly lead to the inability of existing nutritional support methods to effectively reverse amino acid metabolism disorders in SIAI patients, nor to significantly slow down the process of muscle atrophy, thus limiting the potential for improvement in prognosis. Summary of the Invention

[0006] Given the lack of existing compound amino acid formulations targeting specific amino acid metabolic disorders in patients with severe intra-abdominal infections (SIAI), this invention provides a compound amino acid formulation designed based on serum amino acid metabolomics for SIAI patients, aiming to precisely correct their amino acid metabolic imbalance, slow muscle atrophy, reduce septic inflammatory response, and improve clinical prognosis.

[0007] To achieve the above-mentioned technical objectives, the present invention adopts the following technical solution: One objective of this invention is to provide an amino acid composition comprising L-leucine, L-isoleucine and L-histidine, wherein the mass ratio of L-leucine, L-isoleucine and L-histidine is 100 : (20-40) : (40-60).

[0008] Preferably, the mass ratio of L-leucine, L-isoleucine and L-histidine is 100:30:50.

[0009] Preferably, the amino acid composition is used to improve muscle atrophy associated with severe intra-abdominal infection.

[0010] More preferably, the improvement in muscle atrophy associated with severe intra-abdominal infection includes at least one of the following molecular mechanisms: (a) Inhibit the expression of genes related to muscle atrophy; (b) Promotes the expression of muscle-building-related genes; (c) Reduce the expression levels of inflammatory factors; (d) Activation of the autophagy pathway.

[0011] In a further preferred embodiment, the muscle atrophy-related genes include FBXO32 and / or TRIM63, the muscle generation-related genes include MYOD and / or MYOG, the inflammatory factors include TNF-α and / or IL-6, and the autophagy pathway-related proteins include LC3B and / or P62.

[0012] Preferably, the amino acid composition can achieve at least one of the following effects in an animal model of severe intraperitoneal infection induced by cecal ligation and perforation: (a) Improve survival rate; (b) Increase body weight and / or muscle mass index; (c) Improve grip strength.

[0013] A second objective of this invention is to provide the use of the amino acid composition in the preparation of a medicament or nutritional formulation for improving muscle atrophy associated with severe intra-abdominal infection.

[0014] A third objective of this invention is to provide a drug or nutritional preparation for improving muscle atrophy associated with severe intra-abdominal infection, comprising the amino acid composition described above.

[0015] Compared with the prior art, the present invention has the following beneficial effects: I. Precision Advantage – Targeted Design Based on SIAI Real Metabolic Profiles (1) For the first time, the design was based on serum amino acid metabolomics data of SIAI patients, rather than the average nutritional requirements of healthy individuals or general critically ill patients. The serum amino acid profiles of SIAI patients and controls with non-infectious appendicitis and mild trauma were detected by UPLC-Q-TOF / MS platform. The three amino acids that were most significantly reduced in SIAI patients—leucine, isoleucine, and histidine—were screened out, so that the formulation design had a clear pathophysiological basis.

[0016] (2) Precisely correct the unique amino acid imbalance pattern of SIAI. This invention specifically supplements leucine and histidine, which are significantly lacking in SIAI patients, while moderately supplementing isoleucine. This effectively corrects the characteristic pattern of "low leucine, low histidine, and relatively normal isoleucine" in this disease state, avoiding the problem of insufficient supplementation of some amino acids and excessive supplementation of other amino acids caused by the "one-size-fits-all" approach of existing standard formulas.

[0017] II. Functional Effects – Comprehensive Improvement of SIAI-Related Muscle Atrophy Across Multiple Indicators The following effects were experimentally verified in the CLP-induced SIAI mouse model and the LPS-induced C2C12 myoblast atrophy model: (1) Significantly inhibits the expression of muscle atrophy genes. At the mRNA and protein levels, the composition of the present invention can significantly downregulate the expression of muscle-specific E3 ubiquitin ligases FBXO32 and TRIM63 (the expression level in the CLP mouse model was reduced to about 0.3 times that of the control group, P<0.01), blocking the ubiquitin-proteasome pathway-mediated muscle protein degradation from the molecular source.

[0018] (2) Activation of myogenic gene expression. The composition of the present invention can significantly upregulate the mRNA expression levels of myogenic regulatory factors MYOD and MYOG (more than twice that of the control group in the CLP mouse model, P<0.01), promote the differentiation of muscle satellite cells and myotube formation, and drive muscle regeneration and repair.

[0019] (3) Activate autophagy pathway and maintain protein homeostasis. Western blot results showed that the composition of the present invention can increase the LC3B-II / I ratio and downregulate P62 protein expression, indicating that it can effectively activate autophagic flux, promote the clearance of damaged proteins and organelles, and maintain protein homeostasis in skeletal muscle cells.

[0020] (4) Inhibition of inflammatory response. The composition of the present invention can significantly reduce the mRNA expression levels of TNF-α and IL-6, and alleviate the secondary damage to skeletal muscle caused by systemic inflammatory response in SIAI state.

[0021] (5) Improved overall prognostic indicators. In the CLP mouse model, the composition of the present invention can significantly improve the 96-hour survival rate (from 55% to 80%), while increasing the body weight of the model animals, improving forelimb grip strength, and increasing muscle mass index (gastrocnemius muscle / body weight ratio), comprehensively reflecting the overall improvement of the body's functional status.

[0022] III. Safety Advantages – Avoiding the metabolic burden caused by ineffective or excessive components (1) Elimination of ineffective components. This invention has demonstrated through single amino acid intervention experiments that isoleucine at doses of 50-200 mg / kg / day has no significant effect on improving body weight, grip strength, muscle index, and FBXO32 / TRIM63 / MYOD / MYOG expression in SIAI mice. Therefore, this invention controls isoleucine at a low level (30 mg / kg / day) to avoid amino acid loads that are ineffective in improving muscle atrophy.

[0023] (2) Synergistic effect brought about by ratio optimization. Through comparative experiments between MixedAA1 (Leu:Ile:His = 100:30:50) and MixedAA2 (Leu:Ile:His = 100:100:100), it was confirmed that MixedAA1 was significantly better than MixedAA2 in all indicators such as survival rate, grip strength, muscle index, PCR and Western blot (P<0.05). This proves that the specific ratio screened in this invention is not a simple superposition, but has the optimal synergistic effect, and avoids the metabolic risks such as azotemia that may be caused by high doses of amino acids.

[0024] IV. Clinical Translation Potential This invention provides a conversion basis from mouse doses to human equivalent doses (based on body surface area conversion; daily dose for a 60 kg adult: approximately 8.1 g L-leucine, approximately 2.4 g L-isoleucine, and approximately 4.1 g L-histidine), and gives preparation methods for two formulations: lyophilized powder for injection and enteral nutrition solution, demonstrating clear feasibility for clinical translation. Furthermore, it allows for dose adjustments of ±20% based on patient weight and disease severity, reserving space for individualized clinical application.

[0025] In summary, this invention has advantages such as precise design basis, clear molecular regulation, comprehensive functional improvement, superior safety, and clear transformation potential. It effectively solves the clinical problem that existing amino acid formulations cannot correct SIAI-specific metabolic disorders and cannot effectively alleviate muscle atrophy. Attached Figure Description

[0026] Figure 1 This is a heatmap of differentially metabolized serum metabolites in SIAI vs. appendicitis in this invention.

[0027] Figure 2 This is a heatmap of differentially metabolized serum metabolites in SIAI vs. mild trauma in this invention.

[0028] Figure 3 This is a Venn diagram showing the top 10 amino acids that were significantly decreased in SIAI vs. appendicitis and SIAI vs. mild trauma in this invention.

[0029] Figure 4 The figure shows the effect of different doses of a single amino acid on the body weight of CLP mice in this invention.

[0030] Figure 5 The figure shows the effect of different doses of a single amino acid on the forelimb grip strength of CLP mice in this invention.

[0031] Figure 6 The figure shows the effect of different doses of a single amino acid on the muscle mass index (MQI) of CLP mice in this invention.

[0032] Figure 7 The figure shows the effect of different doses of a single amino acid on the expression of FBXO32, TRIM63, MYOD and MYOG mRNA in the gastrocnemius muscle of CLP mice in this invention.

[0033] Figure 8 This is a diagram illustrating the construction process of the CLP mouse in this invention.

[0034] Figure 9 The figure shows the effects of the optimal concentrations of leucine and histidine and the mixed amino acid formulations MixedAA1 and MixedAA2 in this invention on the survival rate, body weight, forelimb grip strength, and gastrocnemius muscle mass index of CLP mice.

[0035] Figure 10 The figure shows the effect of the optimal concentrations of leucine and histidine and the mixed amino acid formulations MixedAA1 and MixedAA2 on the expression of FBXO32, TRIM63, MYOD, and MYOG mRNA in the gastrocnemius muscle of CLP mice.

[0036] Figure 11The figure shows the effect of the optimal concentrations of leucine and histidine and the mixed amino acid formulations MixedAA1 and MixedAA2 on the expression of TNF-α and IL-6 mRNA in the gastrocnemius muscle of CLP mice.

[0037] Figure 12 The results of HE staining and muscle fiber quantification of mouse gastrocnemius muscle in this invention are shown.

[0038] Figure 13 The results of the CCK8 screening experiment in this invention are shown.

[0039] Figure 14 The figure shows the effect of the mixed amino acid formulation MixedAA1 in this invention on the expression of LPS-induced LC3B, FBXO32, TRIM63, LC3B-II / I and P62 proteins in C2C12 myotube cells.

[0040] Figure 15 The figure shows the effect of the mixed amino acid formulation in this invention on the expression of LPS-induced C2C12 myotubes FBXO32, TRIM63, MYOD and MYOG mRNA.

[0041] Figure 16 The figure shows the effect of the mixed amino acid formulation in this invention on LPS-induced expression of TNF-α and IL-6 mRNA in C2C12 myotubes.

[0042] Figure 17 This is a flowchart illustrating the preparation process of the single amino acid injection solution in this invention. Detailed Implementation

[0043] The following examples are used to illustrate the present invention, but are not intended to limit the scope of the invention. Any modifications or substitutions made to the methods, steps, or conditions of the present invention without departing from the spirit and essence of the invention are within the scope of the invention. The reagents, products, and instruments used in the following examples are all commercially available, and the methods used in the examples, unless otherwise specified, are consistent with conventionally used methods.

[0044] The technical solution of the present invention will be further described in detail below with reference to the embodiments.

[0045] Example 1: Metabolomics screening of differentially expressed amino acids in serum of SIAI patients 1. Research Subjects Infection group: 10 patients with intra-abdominal infection. Inclusion criteria: ① abdominal infection symptoms and signs; ② abnormal infection-related inflammatory markers; ③ confirmed intra-abdominal infection focus, pus or abscess by imaging or intraoperative exploration; ④ exclusion of non-infectious peritonitis and abdominal pain caused by other reasons; ⑤ APACHE II score > 15.

[0046] Non-infectious control group: 10 patients with appendicitis and 10 patients with minor trauma. Appendicitis inclusion criteria: ① Simple appendicitis without complications; ② At least two typical symptoms (migratory right lower quadrant pain, fixed right lower quadrant tenderness, rebound tenderness); ③ Laboratory tests: white blood cell count >10×10⁻⁶. 9 / L, neutrophil percentage >75%, CRP >10 mg / L; ④ Imaging examination meets at least one of the following criteria. Inclusion criteria for mild trauma: ① No concurrent infection; ② Stable basic vital signs; ③ Injury Severity Score (ISS) ≤8 points, Glasgow Coma Scale (GCS) = 15 points; ④ Limited extent of injury; ⑤ Functional assessment: Activities of Daily Living (ADL) score ≥75 points.

[0047] 2. Detection Method The amino acids and amine metabolites in the human serum of the above-mentioned subjects were quantitatively detected using an ultra-high performance liquid chromatography-high resolution tandem quadrupole mass spectrometry (UPLC-Q-TOF / MS) platform.

[0048] 3. Screening strategy and results The top 10 amino acids with the most significant decreases were compared between the "abdominal infection group vs. appendicitis group" and the "abdominal infection group vs. mild trauma group". The amino acids with a common significant decrease were identified by taking the intersection of Venn diagrams. The results are as follows: Figures 1-3 As shown, the three amino acids with the most significant differences were finally identified as: leucine (Leu), isoleucine (Ile), and histidine (His).

[0049] Example 2: Construction of CLP-induced SIAI mouse model and screening of optimal dosage of single amino acid 1. Laboratory animals C57BL / 6 or BALB / c mice, aged 8-12 weeks and weighing 20-25 g, were selected.

[0050] 2. Main Reagents and Instruments Reagents: 1.25% tribromoethanol (for anesthesia), povidone-iodine, 75% ethanol, 5-0 sterile silk sutures, 4-0 sterile sutures, sterile saline (preheated to 37°C).

[0051] Instruments and consumables: fine surgical scissors, ophthalmic forceps (straight / curved), hemostats, needle holders, 21G sterile syringe needles, heating pads, operating tables, razors, sterile gauze, and sterile gloves.

[0052] 3. Procedure for cecal ligation and perforation (CLP) Preoperative fasting for 6 hours, with free access to water. Anesthesia was administered via intraperitoneal injection of 1.25% tribromoethanol (0.2 ml / 10 g). After the righting reflex disappeared, the mouse was fixed prone on the operating table. Abdominal hair was shaved, and disinfection was performed using iodine-ethanol-iodine. A 1.5-2 cm incision was made along the midline of the abdomen, and the muscle layer was bluntly dissected. The peritoneum was incised along the linea alba to enter the abdominal cavity. The cecum was gently lifted to locate the ileocecal valve and the distal end of the cecum. The distal half of the cecum (in a moderate model) was ligated with 5-0 silk sutures. A 21G needle was used to puncture the midpoint of the ligated segment 1-2 times, gently pressing the cecum to expel a small amount of fecal matter to ensure patency. The cecum was gently returned to the abdominal cavity. The peritoneum and muscle layer were continuously sutured with 4-0 sutures, and the skin was intermittently sutured. The incision was disinfected with iodine, and pre-warmed saline (0.2 ml) was injected intraperitoneally for rehydration. The mouse was placed on a 37°C warming pad for resuscitation and returned to its cage after fully waking. The process of constructing CLP mice is as follows: Figure 8 As shown.

[0053] 4. Postoperative observation and group medication Monitor the body temperature, weight, activity level, and fur condition of mice.

[0054] CLP model mice were randomly divided into 10 groups (n=6 / group): CLP + saline control group, CLP + L-Leu 50 / 100 / 200 mg / kg / day group, CLP + L-Ile 50 / 100 / 200 mg / kg / day group, CLP + L-His 50 / 100 / 200 mg / kg / day group, and a sham-operated group (Sham) as a baseline control. Intraperitoneal injection of CLP was started 6 hours after CLP, once daily, for 96 hours until sacrifice.

[0055] 5. Detection Indicators and Methods (1) Weight change: Weigh yourself at a fixed time every day (9-11 am) using a precision electronic balance.

[0056] (2) Forelimb grip strength: Using a mouse grip strength tester, gently guide the mouse's forelimbs to grasp the probe, gently pull the mouse's tail, record the peak pull force, repeat 3-5 times and take the average value.

[0057] (3) Muscle mass index (MQI): After the mice were euthanized, the gastrocnemius muscle was quickly dissected and separated. The residual blood was removed by rinsing with pre-cooled physiological saline. The surface moisture was absorbed by filter paper and the wet weight was measured. The ratio of gastrocnemius muscle mass to mouse body weight was calculated.

[0058] (4) PCR detection of muscle atrophy genes (FBXO32, TRIM63) and myogenic genes (MYOD, MYOG): Approximately 50 mg of mouse gastrocnemius muscle tissue was used to extract total RNA using TRIzol reagent. The concentration and purity were determined by NanoDrop (A260 / A280 ratio 1.8-2.0). 2 μg of total RNA was used to synthesize cDNA using a reverse transcription kit. Amplification was performed using SYBR Green qPCR premix on a real-time quantitative PCR instrument. β-actin was used as an internal reference gene, and the relative expression levels of each gene were calculated using the 2^(-ΔΔCt) method. Primer sequences are shown in Table 1.

[0059] Table 1. Details of qPCR primer sequences for internal control, muscle atrophy, and myogenic genes.

[0060] 6. Screening Results The results showed that the L-Leu 100 mg / kg / day group significantly improved body weight, forelimb grip strength, and MQI (P<0.05), significantly decreased FBXO32 / TRIM63 expression, and significantly increased MYOD / MYOG expression (P<0.05). The L-His 100 mg / kg / day group also significantly improved the above indicators (P<0.05). None of the L-Ile dosage groups showed significant improvement in body weight, grip strength, or MQI, and PCR results showed no significant changes (P>0.05). Therefore, the single effective doses were determined to be L-Leu 100 mg / kg / day and L-His 100 mg / kg / day. Figures 4-7 ).

[0061] Example 3: Design and in vivo animal validation of a mixed amino acid formulation 1. Formula design Based on the metabolic differences of Leu, Ile, and His in the serum of SIAI patients, two mixed formulations were designed (Table 2, unit: mg / kg / day, intraperitoneal injection): Table 2 Formulation Design Details

[0062] 2. Animal grouping and administration CLP mice were randomly divided into 4 groups (n=6 / group): Sham group, CLP+saline group, CLP+MixedAA1 group, and CLP+MixedAA2 group. Intraperitoneal injection of the drug was started 6 hours after CLP, once a day, and mice were observed or euthanized after 96 hours of drug administration.

[0063] 3. Detection indicators In addition to body weight, grip strength, MQI, and PCR (FBXO32, TRIM63, MYOD, MYOG) as described in Example 2, the following tests were added: (1) Survival rate: The survival status of mice in each group was recorded after 96 hours of continuous observation.

[0064] (2) PCR detection of inflammatory factors: mRNA expression of TNF-α and IL-6 (HPRT was used as an internal reference for inflammatory factors), primer sequences are shown in Table 3: Table 3. Details of qPCR primer sequences for inflammatory factor genes

[0065] (3) Western blot detection: Total protein was extracted from gastrocnemius muscle tissue, and after SDS-PAGE electrophoresis, membrane transfer and blocking, anti-FBXO32, anti-TRIM63, anti-LC3B, anti-P62 and internal control antibody (β-actin) were added and incubated overnight. After secondary antibody incubation, ECL color development was performed, and the imaging system was used for scanning and quantitative analysis.

[0066] 4. Results (1) The survival rate, body weight, grip strength, and MQI of the MixedAA1 group at each time point were significantly better than those of the MixedAA2 group and the saline group (P<0.05). Figure 9 ).

[0067] (2) The expression levels of FBXO32 and TRIM63 mRNA in the MixedAA1 group were 0.32±0.05 and 0.28±0.04 times that in the CLP+saline group, respectively, which were significantly lower than those in the MixedAA2 group (0.67±0.08 and 0.61±0.07 times, respectively) (P<0.01); the expression levels of MYOD and MYOG were 2.45±0.21 and 2.13±0.19 times that in the CLP+saline group, respectively, which were significantly higher than those in the MixedAA2 group (1.52±0.18 and 1.48±0.15 times, respectively) (P<0.01). Figure 10 ).

[0068] (3) The expression of TNF-α and IL-6 mRNA in the MixedAA1 group was significantly lower than that in the MixedAA2 group and the saline group (P<0.05). Figure 11 ).

[0069] (4) The FBXO32 and TRIM63 protein bands in the MixedAA1 group were significantly lighter; the LC3B-II / I ratio was significantly increased, and the expression of P62 protein was significantly downregulated, indicating that autophagic flux was activated. Figure 14 ).

[0070] Example 4: LPS-induced C2C12 myoblast atrophy model and in vitro validation 1. Cell Culture and Differentiation C2C12 mouse myoblasts were proliferated to 80%-90% confluence in DMEM (growth medium) containing 10% FBS, and then cultured in DMEM (differentiation medium) containing 2% horse serum for 4-6 days to differentiate into mature myotubes.

[0071] 2. Construction of LPS-induced muscle atrophy model and screening of optimal drug concentration Mature myotubes were transferred to differentiation medium containing LPS (500 ng / mL) and treated for 24-48 hours. The optimal amino acid concentrations were screened using the CCK-8 assay: Leu, Ile, and His were administered at concentration gradients of 0, 0.5, 1, 2, 4, and 8 μmol / L, respectively. Results showed that Leu and His exhibited the best cell proliferation at 2 μmol / L, while Ile showed a slightly weaker promoting effect. Figure 13 ).

[0072] 3. Single amino acid intervention LPS-induced muscle atrophy models were treated with 2 μmol / L Leu, Ile, and His, respectively. The mRNA expression of FBXO32, TRIM63, MYOD, and MYOG was detected by qPCR. The results were consistent with animal experiments: FBXO32 and TRIM63 expression was significantly decreased, while MYOD and MYOG expression was significantly increased in the Leu and His groups (P<0.05); no significant changes were observed in any genes in the Ile group (P>0.05).

[0073] 4. Mixed amino acid intervention Based on the above results, the concentrations of the in vitro mixed formulations were set (Table 4, unit: μmol / L): Table 4 In vitro mixed formulation

[0074] LPS-induced myotubes were treated with MixedAA1 or MixedAA2, respectively. qPCR was used to detect the mRNA expression of FBXO32, TRIM63, MYOD, MYOG, and TNF-α and IL-6. Results are as follows: Figure 15 and Figure 16 As shown, the expression levels of FBXO32, TRIM63 mRNA, TNF-α, and IL-6 mRNA in the MixedAA1 group were significantly lower than those in the MixedAA2 group (P<0.01), while the expression levels of MYOD and MYOG were significantly higher in the MixedAA1 group (P<0.01). MixedAA1 was significantly superior to MixedAA2 in all indicators.

[0075] Example 5: Preparation of the preferred formulation and human equivalent dosage 1. Optimal formula composition The preferred formulation of MixedAA1 of this invention has the following composition (based on mouse dosage): L-leucine 100 mg / kg / day, L-isoleucine 30 mg / kg / day, and L-histidine 50 mg / kg / day.

[0076] 2. Preparation method (applicable to injectable formulations) (1) Weighing: Weigh L-leucine, L-isoleucine and L-histidine powders according to the formula amount; (2) Mixing: Mix the three amino acid powders evenly under sterile conditions; (3) Dissolving: Add sterile physiological saline (10 mL / mouse dose) and stir until completely dissolved; (4) Filtration and sterilization: Filter using a 0.22 μm filter membrane and collect the filtrate; (5) Quality inspection: Check the clarity and sterility of the solution; (6) Packaging: Dispense into sterile containers, seal, and store at 4°C.

[0077] This formula can also be prepared as a lyophilized powder for injection (reconstituted with physiological saline before use) or a solution / gel for enteral nutrition.

[0078] 3. Human equivalent dose conversion Based on body surface area conversion, the above mouse dosages are converted to the following daily doses for a 60 kg adult: approximately 8.1 g / day of L-leucine, approximately 2.4 g / day of L-isoleucine, and approximately 4.1 g / day of L-histidine. In clinical use, the dosage can be adjusted by ±20% according to the patient's weight and the severity of their condition.

[0079] Comparison of Example 1 with commercially available standard amino acid formulations A commercially available critical care nutritional formula contains a Leu, Ile, and His ratio of approximately 1:0.5:0.4, which has not been adjusted for SIAI. Based on the metabolomics data of this invention, it is estimated that the commercially available formula has a relatively high Ile content and a relatively low His content, which cannot effectively correct the characteristic spectrum of SIAI patients of "low Leu, low His, and relatively normal Ile," and may exacerbate the risk of azotemia due to excessive Ile.

[0080] Comparison of Example 2 and MixedAA2 Comparing MixedAA1 and MixedAA2 (Leu:Ile:His = 100:100:100) of the present invention, MixedAA1 was significantly superior to MixedAA2 in all indicators, including survival rate, grip strength, muscle index, PCR and Western blot in CLP mouse models (P<0.05). This proves that the Leu:Ile:His = 100:30:50 ratio selected by the present invention has the optimal synergistic effect, and that more amino acid types and dosages are not necessarily better.

[0081] 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. An amino acid composition, characterized in that, It is composed of L-leucine, L-isoleucine and L-histidine, wherein the mass ratio of L-leucine, L-isoleucine and L-histidine is 100 : (20-40) : (40-60).

2. The amino acid composition according to claim 1, characterized in that, The mass ratio of L-leucine, L-isoleucine and L-histidine is 100:30:

50.

3. The amino acid composition according to claim 1, characterized in that, The amino acid composition is used to improve muscle atrophy associated with severe intra-abdominal infection.

4. The amino acid composition according to claim 3, characterized in that, The improvement in muscle atrophy associated with severe intra-abdominal infection includes at least one of the following molecular mechanisms: (a) Inhibit the expression of genes related to muscle atrophy; (b) Promotes the expression of muscle-building-related genes; (c) Reduce the expression levels of inflammatory factors; (d) Activation of the autophagy pathway.

5. The amino acid composition according to claim 4, characterized in that, The muscle atrophy-related genes include FBXO32 and / or TRIM63, the muscle growth-related genes include MYOD and / or MYOG, the inflammatory factors include TNF-α and / or IL-6, and the autophagy pathway-related proteins include LC3B and / or P62.

6. The amino acid composition according to claim 1, characterized in that, The amino acid composition can achieve at least one of the following effects in an animal model of severe intraperitoneal infection induced by cecal ligation and perforation: (a) Improve survival rate; (b) Increase body weight and / or muscle mass index; (c) Improve grip strength.

7. Use of the amino acid composition according to any one of claims 1-6 in the preparation of a medicament or nutritional preparation for improving muscle atrophy associated with severe intra-abdominal infection.

8. A pharmaceutical or nutritional preparation for improving muscle atrophy associated with severe intra-abdominal infection, characterized in that, It contains the amino acid composition according to any one of claims 1-6.

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