Low-molecular hyaluronic acid fragment oral preparation and application thereof

Abstract

The invention discloses a low-molecular hyaluronic acid fragment oral preparation. The low-molecular hyaluronic acid fragment has an average molecular weight of 35kDa, gt; the distribution range of the hyaluronic acid fragments is between 10 kDa and 70 kDa; or has an average molecular weight of 70 kDa, gt; the distribution range of the hyaluronic acid fragments is between 40 kDa and 100 kDa; the traditional Chinese medicine composition is directly used as an oral preparation and is a high-dose gel oral preparation, and the oral dosage is 5-10g / day and is 1-2 times per day. The oral preparation can be rapidly absorbed into blood through intestinal lymph nodes and spleen after being orally taken. The oral preparation produces an effect within 2 hours after being orally taken, and is a quick-acting oral preparation. The invention breaks through the barrier of oral druggability of the low-molecular hyaluronic acid fragment in the prior art, and finds for the first time that the low-molecular hyaluronic acid fragment in a high-dose gel form is directly used as an oral preparation and can be rapidly absorbed into blood through intestinal lymph nodes and spleen after being orally taken; the health-care wine has various effects of treating various pains, subcutaneous fat, beautifying skin, reducing blood pressure, relieving spicy mouth feel after drinking, protecting gastrointestinal tracts and the like.

Description

Technical Field

[0001] The present invention relates to the field of biomedicine, and in particular to an oral preparation of low-molecular hyaluronic acid fragments and application thereof. Background Art

[0002] Patent applications (WO / 2021 / 180252) and published articles (1, 2) indicate that low-molecular-weight hyaluronic acid fragment injections with an average molecular weight of 35 kDa (>90% distribution range 10-70 kDa) with good human tissue permeability are effective in treating chronic non-healing wounds and various pains by injecting 0.1 g of subcutaneous fat once a day. Studies in Nature Communication have shown that the analgesic effect of the above low-molecular-weight hyaluronic acid fragment injections is through the inhibition of the pain-related calcium channel TPPV1 (3, 4).

[0003] It is well known to those skilled in the art that more than 90% of injections cannot be converted into oral preparations because the two dosage forms have different absorption pathways, which may lead to different mechanisms of action and fail to produce corresponding therapeutic effects.

[0004] As for hyaluronic acid fragments, current research literature shows that oral hyaluronic acid is generally degraded into particularly small hyaluronic acid fragments in the intestine and absorbed into the blood (5), which does not support the oral drugability of low-molecular hyaluronic acid fragments with an average molecular weight of 35 kDa. Therefore, if oral preparations are to be made, those skilled in the art generally think of adopting a preparation method of encapsulating low-molecular hyaluronic acid fragments with an average molecular weight of 35 kDa to reduce gastric acid degradation, various trypsin degradation in the duodenum and upper small intestine (jejunum), and bacterial degradation in the terminal small intestine and large intestine, such as using capsule preparations or dry powder oral preparations with a large amount of encapsulated excipients, but this will reduce the absorption area of ​​the upper gastrointestinal tract of the low-molecular hyaluronic acid fragments with an average molecular weight of 35 kDa, reduce the absorption of the low-molecular hyaluronic acid fragments with an average molecular weight of 35 kDa, and fail to reach a daily absorption dose of >0.1 gram of low-molecular hyaluronic acid fragments of 35 kDa, nor does it support the oral drugability of low-molecular hyaluronic acid fragments with an average molecular weight of 35 kDa.

[0005] As can be seen from the above, hyaluronic acid fragments, especially low molecular weight hyaluronic acid fragments with an average molecular weight of 35 kDa (>90% distribution range 10-70 kDa), are currently mainly used as injections to effectively treat chronic non-healing wounds and various pains by injecting 0.1 gram of subcutaneous fat each time per day.

[0006] References:

[0007] 1.Feng He Xu, Dylan Treger, Xinyue Ma, Xiao Xiao Jia, Joshua H Shofaro, Taivanbat Ganbaatar, Munkh-Amgalan Gantumur, Jessica Hui, Mizhou Hui, Chao Meng(2024)Local injection of a freshly manufactured 35kDa hyaluronan fragmentreduces neuropathic and inflammatory pain:A clinical study,European Journalof InflammationVolume 22,May 2024,https: / / doi.org / 10.1177 / 1721727X241258268.

[0008] 2.Treger D,Zhang L,Jia X,et al.A clinical study of the localinjection of a freshly manufactured 35kDa hyaluronan fragment for treatingchronic wounds.Int Wound J 2024:21(5):e14906.

[0009] 3.Caires,R.Luis,E.Taberner,FJ.Fernandez-Ballester,G.Ferrer-Montiel,A.Balazs,EA.Gomis,A.Belmonte,C.Dela E.Hyaluronan modulates TRPV1 channel opening, reducing peripheral nociceptor activity and pain.NatCommun 2015:6:8095.

[0010] 4.Bujak JK,Kosmala D,Szopa IM,Majchrzak K,Bednarczyk P.Inflammation, Cancer and Immunity-Implication of TRPV1 Channel.Front Oncol.2019 Oct 16;9:1087.doi:10.3389 / fonc.2019.01087.PMID:31681615;PMCID:PMC6805766.

[0011] 5. Kimura M, Maeshima T, Kubota T, Kurihara H, Masuda Y, Nomura Y. Absorption of Orally Administered Hyaluronan. J Med Food. 2016 Dec; 19(12):1172-1179.doi:10.1089 / jmf.2016.3725.PMID:27982756. Summary of the invention

[0012] The first technical problem to be solved by the present invention is to provide a novel oral preparation of low-molecular-weight hyaluronic acid fragments, so as to explore more potential clinical application values ​​of low-molecular-weight hyaluronic acid fragments.

[0013] The second technical problem to be solved by the present invention is to provide an application of an oral preparation of a low-molecular hyaluronic acid fragment.

[0014] In order to solve the above technical problems, the present invention adopts the following technical solutions:

[0015] A low-molecular-weight hyaluronic acid fragment oral preparation, wherein the low-molecular-weight hyaluronic acid fragment has an average molecular weight of 35 kDa and >90% of the distribution range is between 10-70 kDa; or the low-molecular-weight hyaluronic acid fragment has an average molecular weight of 70 kDa and >90% of the distribution range is between 40-100 kDa; the low-molecular-weight hyaluronic acid fragment is directly used as an oral preparation, which is a high-dose gel oral preparation, and its oral dosage is 5-10 g / day, 1-2 times a day.

[0016] After oral administration, the oral preparation can be rapidly absorbed into the blood through the intestinal lymph nodes and spleen.

[0017] The oral preparation is a fast-acting oral preparation.

[0018] The use of the aforementioned low-molecular hyaluronic acid fragment oral preparation as a drug for treating pain or in the preparation of a drug for treating pain can produce an effect within 2 hours after oral administration.

[0019] The pain described is: inflammatory pain in the shoulder / neck / back, neuropathic pain from herpes zoster / diabetic foot, pain from bone metastasis of advanced cancer, pain from radiotherapy for advanced cancer, pain after dental implant surgery, pain after vaginal egg retrieval, muscle strain, muscle stiffness and discomfort, sports injury pain, or pain from diabetic foot wounds.

[0020] The shoulder / neck / back inflammatory pain is shoulder / neck / back inflammatory pain caused by fasciitis.

[0021] The low-molecular-weight hyaluronic acid fragment oral preparation is used as a subcutaneous lipolysis or skin beauty drug or in the preparation of a subcutaneous lipolysis or skin beauty drug.

[0022] The skin beauty treatment includes hydrating the skin, reducing skin inflammation, improving skin elasticity, improving skin gloss or tightening the skin; the subcutaneous lipolysis includes reducing subcutaneous fat on the face, reducing subcutaneous fat on the neck, reducing subcutaneous fat on the abdomen or reducing subcutaneous fat on the legs.

[0023] The invention relates to the use of the aforementioned low-molecular hyaluronic acid fragment oral preparation as a drug for lowering blood pressure or in the preparation of a drug for lowering blood pressure.

[0024] The invention relates to an application of the aforementioned oral preparation of low-molecular-weight hyaluronic acid fragments as a drug for protecting the gastrointestinal tract for relieving a sore and spicy feeling in the mouth, discomfort after drinking or symptoms of drunkenness, or improving abdominal distension and digestive discomfort after drinking, or in the preparation of a drug for protecting the gastrointestinal tract for relieving a sore and spicy feeling in the mouth, discomfort after drinking or symptoms of drunkenness, or improving abdominal distension and digestive discomfort after drinking.

[0025] The present invention breaks through the barrier of oral drugability of low molecular weight hyaluronic acid fragments in the prior art, and discovers for the first time that high doses of hyaluronic acid fragments in gel form can be directly used as oral preparations, which can be rapidly absorbed into the blood through the intestinal lymph nodes and spleen after oral administration, and have multiple functions such as treating various pains (with effects occurring within 2 hours after oral administration), subcutaneous fat storage, skin beauty, lowering blood pressure, relieving spicy taste in the mouth after drinking, and protecting the gastrointestinal tract, etc., and explores more potential clinical application values ​​of low molecular weight hyaluronic acid fragments. DETAILED DESCRIPTION

[0026] The present invention is described below by means of specific embodiments. The embodiments described herein are only used to illustrate and explain the present invention and are not used to limit the present invention.

[0027] As described in the background of the present invention, the prior art does not support the use of low molecular weight hyaluronic acid fragments with an average molecular weight of 35 kDa (>90% of the distribution range is 10-70 kDa) as oral preparations to treat a variety of diseases that can be successfully treated by injection.

[0028] It is assumed that the use of low-molecular-weight hyaluronic acid fragments taken orally and absorbed through the gastrointestinal tract can also treat the various diseases successfully treated by injection. The intestine is the main place for the absorption of various nutrients in humans, but the low-molecular-weight hyaluronic acid fragments with an average molecular weight of 35kDa are first degraded by gastric acid, then by various trypsin degradation in the duodenum and upper small intestine (jejunum), and then by bacteria in the terminal small intestine and large intestine. The possibility of direct intestinal absorption of 0.1 grams per day should also be low. Therefore, the above theoretical assumption cannot be established. Current research literature also shows that orally taken hyaluronic acid is generally degraded into very small hyaluronic acid fragments in the intestine and absorbed into the blood (5), which does not support the oral drugability of low-molecular-weight hyaluronic acid fragments with an average molecular weight of 35kDa.

[0029] However, the gastrointestinal tract is an organ rich in lymphatic tissue, for example, mesenteric lymph nodes are very abundant. The applicant's research shows that the low-molecular hyaluronic acid fragment with an average molecular weight of 35kDa rapidly combines with inflammatory lymphocytes after subcutaneous injection, diffuses into lymph and spleen backflow into the blood, and prompts that the low-molecular hyaluronic acid fragment with an average molecular weight of 35kDa produces the effect of anti-inflammatory treatment of chronic inflammatory swelling by "diffusion into lymph and spleen backflow into the blood". Therefore, the present invention hypothesis: 1. The low-molecular hyaluronic acid fragment with an average molecular weight of 35kDa for oral administration is different from other oral foods and drugs. After oral administration, part of it can enter the gastrointestinal lymphatic tissue by "affinity" with lymph and rapid "absorption"; 2. If the low-molecular hyaluronic acid fragment with an average molecular weight of 35kDa really mainly enters the gastrointestinal lymphatic tissue first, it will rapidly combine with inflammatory lymphocytes, diffuse into lymph and spleen backflow into the blood, thereby producing the effect of treating chronic inflammatory swelling and pain of other tissues in the intestine and human body. The present invention also verifies through research that after small intestinal perfusion or oral administration, it can be effectively absorbed through the mesenteric lymphatic system and transported into the blood through the spleen. The oral administration provides an alternative and new indication for the HA35 injection route.

[0030] However, further research of the present invention has found that oral products with the same dosage as injection or similar small dosage cannot produce corresponding therapeutic effects, which further hinders its application as an oral preparation.

[0031] The present invention discovered through an unexpected study that when a high dose (tens of times higher than the conventional dose) is used, it unexpectedly has a very good therapeutic effect. Based on this, a new oral preparation and application of low-molecular hyaluronic acid fragments have been developed. At the same time, corresponding studies on other low-molecular hyaluronic acid fragments have also been conducted, proving that the use of low-molecular hyaluronic acid fragments with an average molecular weight of 70kDaHA (>90% of the distribution range is 40-100kDa) can also achieve basically the same effect.

[0032] Regarding the dosage of hyaluronic acid fragments, taking food as an example, my country stipulates that the amount added to food shall not exceed 160 mg per day, and the conventional dosage is 0.16 g per day; while the maximum dosage abroad is 0.26 g per day. Although the present invention is a medicine, it is generally difficult for those skilled in the art to think of a dosage that exceeds that of food by dozens of times. However, the present invention has found that when a high dose (exceeding the conventional dose by dozens of times) is used, it unexpectedly has a good therapeutic effect.

[0033] It should be noted that the low molecular weight hyaluronic acid fragments with an average molecular weight of 35 kDa (>90% of the distribution range is 10-70 kDa) and an average molecular weight of 70 kDa HA (>90% of the distribution range is 40-100 kDa) used in the present invention are in the form of gel as a whole, and the oral preparation directly uses the easily absorbed gel fragments.

[0034] The following is a description of the embodiments:

[0035] Example 1

[0036] Objective: To investigate the absorption pathway of low molecular weight hyaluronic acid fragment HA35 with an average molecular weight of 35 kDa (>90% distribution range 10-70 kDa) after oral administration, especially its transport through the intestinal lymphatic system and the dynamic changes of absorption in the spleen, lymph nodes and portal vein.

[0037] Background: Previous studies have shown that orally administered hyaluronic acid (HA) is degraded into small molecular fragments by bacteria in the intestine, which are then absorbed into the blood through the intestine and resynthesized into large molecular HA in the body [5]. It is well known that the intestinal lymphatic system is not only involved in immune response, but also responsible for the absorption and transport of some water-soluble molecules. This example study hypothesized that orally administered HA35k can quickly enter the body through the intestinal lymphatic system. To verify this hypothesis, this study used a rat model to study the absorption pathway of HA35 through the intestinal lymphatic system and portal vein after oral administration, and evaluated its absorption rate and concentration distribution in the spleen, lymph nodes, portal vein and liver.

[0038] method:

[0039] 1. Experimental animals: Male Wistar rats, 5-6 weeks old, weighing 160-180 g, fasted for 14-16 hours before the experiment.

[0040] 2. Experimental design: Each rat was orally administered 1 g of HA35. The following three different collection methods were used in the study: (1) Peripheral blood collection: After the rats were orally administered with the HA35 preparation, peripheral blood samples were collected and plasma samples were obtained by centrifugation; (2) Portal vein blood collection: After anesthesia, the rats underwent a midline abdominal incision to expose the portal vein, and blood was collected with a 30G needle (about 300 μL each time) for 120 minutes; (3) Lymph fluid collection: The rats were used for lymph fluid collection by making a midline incision in the abdomen, inserting a polyethylene cannula into the thoracic lymphatic duct, and collecting lymph fluid from 0 to 24 hours every 2 hours. Sample storage and testing: All samples were stored at -20°C until testing. The concentration of hyaluronic acid HA in blood and lymph was determined using a hyaluronic acid quantification kit (COSMO Bio Co., Ltd., Tokyo, Japan). The limit of quantification was 3 ng / mL HA. HA35 fragments were provided by Qingdao Huinod Biotechnology Co., Ltd.

[0041] result:

[0042] 1. Changes in HA concentration in plasma: After oral administration of HA35, the HA concentration in peripheral plasma increased significantly within 1 to 8 hours after administration, with a peak concentration of approximately 120 ng / mL; 24 hours after administration, the concentration increased to 150 ng / mL. The HA concentration in portal vein plasma increased significantly 2 hours after administration, reaching a peak of 573.50±133.79 ng / mL at 120 minutes.

[0043] 2. Changes in HA concentration in lymph: After oral administration of 35kDa HA, the HA concentration in lymph increased significantly, reaching a peak of 4293.47±689.30ng / mL. By calculating the cumulative volume of lymph, the total amount of 35kDa HA flowing into the lymph within 24 hours was approximately 69.54±16.78μg.

[0044] Discussion and Conclusion:

[0045] This study verified that oral HA35 can enter the body through the intestinal lymphatic system and portal vein. The lymphatic system is not only an important component of immune function, but also plays a key role in the transport of highly water-soluble molecules. The experimental results showed that HA35 can be rapidly absorbed by the intestinal epithelium and transported to the blood and lymph through the portal vein and lymphatic system. In particular, the high concentration of HA35 in the lymph further supports the important role of the intestinal lymphatic system in the absorption of HA35.

[0046] Studies have shown that HA35 can be effectively absorbed through the oral route, mainly entering the body through the intestinal lymphatic system and portal vein. At different time points after oral administration, the plasma and lymph concentrations of HA35 increased significantly, which provides a basis for the development of HA35 in oral applications. The intestinal lymphatic system is a key HA35 absorption and transport pathway. The absorption and transport of HA35 may be through the affinity of hyaluronic acid receptors CD44 and LYVE-1, which can become a key direction for further research and optimization of small molecule hyaluronic acid formulas in the future.

[0047] Example 2

[0048] Objective: To study the absorption pathway of a fluorescently labeled low-molecular-weight hyaluronic acid fragment of 35 kDa (>90% of the distribution range is 10-70 kDa) after perfusion through the upper small intestine, with a focus on its absorption and distribution in the blood, lymph nodes, and spleen.

[0049] Background: Hyaluronic acid (HA) is partially degraded by artificial gastric juice, but due to the limited effect of gastric acid on its molecular weight, HA can still maintain high integrity and reach the upper small intestine. This study hypothesized that direct perfusion of HA35 into the upper small intestine can better represent the intestinal absorption pathway of oral administration, and its absorption pathway includes the intestinal lymphatic system and portal vein. This experiment is the first to analyze the lymphatic absorption pathway of HA35 in vivo by fluorescently labeling HA35 and evaluate its amount and efficiency of transport to the spleen and lymph nodes.

[0050] method:

[0051] 1. Preparation of fluorescently labeled HA35: (1) Take 0.5 ml (20 mg / mL) HA35; (2) Add 25 μl Cy5.5 amine (33 mg / ml) to the HA35 solution; (3) Weigh 11 mg NHS and 5 mg EDC respectively. Dissolve NHS / EDC powder in 100 μl DMSO, then add to the mixed solution of HA35 and Cy5.5, and vortex briefly and vigorously; (4) Add 5 μl TEA (1% v / v) to the mixed solution to catalyze the reaction. Then incubate the solution overnight on a shaker at room temperature at 1000 rpm; (5) The reaction product is purified three times using a PD-10 column (GE Healthcare Life Sciences, PA, USA); (6) Collect the Cy5.5-HA35 conjugate fraction and lyophilize it to obtain a bright blue dry product.

[0052] 2. Animal experiment preparation: Select male Wistar rats, 5-6 weeks old, weighing 160-180g. Rats fasted for 14-16 hours before the experiment. Rats were divided into an experimental group (Cy5.5-HA35) and a control group (normal saline). Cy5.5-HA35 (at a dose of 0.5-1g / kg body weight) was delivered into the gastrointestinal tract by oral gavage to observe its absorption and distribution in the body.

[0053] 3. Collection and processing of tissue samples: Intestinal tissue, spleen and lymph nodes were collected at different time points (such as 4h, 8h, 24h). The samples were fixed in 4% paraformaldehyde solution and thin slices were made using a freezing microtome or a paraffin embedding microtome. The slices can be stained with DAPI to mark the cell nuclei. The distribution of Cy5.5-HA in the slices was observed under a fluorescence microscope or a confocal microscope. The distribution and intensity of fluorescent HA in the spleen, lymph nodes and intestines were analyzed using ImageJ and other software. The absorption efficiency of HA at different time points and in different tissues was evaluated through statistical analysis.

[0054] result:

[0055] Through fluorescence microscopy, Cy5.5-labeled HA35 was rapidly transferred to the blood and lymphatic system after being absorbed by the intestinal epithelium. The presence of fluorescent HA in the spleen, lymph nodes and liver can be detected at different time points of 4h, 8h and 24h, and the distribution is time-dependent. At 4h, HA is mainly concentrated in intestinal epithelial cells, and over time, fluorescent HA gradually accumulates in lymph nodes and spleen. After 24 hours, the highest fluorescence intensity is in the lymphatic system, including the spleen and mesenteric lymph nodes.

[0056] Discussion and Conclusion:

[0057] This study used fluorescent labeling technology to reveal for the first time that HA35 is absorbed and transported not only through the portal vein and liver, but also through the intestinal lymph nodes and spleen. The lymphatic system plays an important role in the absorption of hyaluronic acid, especially after perfusion of the upper small intestine of rats, HA35 quickly enters the lymph nodes and is further distributed to tissues such as the spleen.

[0058] These findings reveal a dual absorption mechanism for HA35, namely, transport to the liver through the portal vein and to the spleen through the lymphatic system. This provides new insights into the oral application of small molecule hyaluronic acid and its targeted delivery in different tissues, which will help further develop formulations for oral small molecule hyaluronic acid. Future studies can further refine the mechanism of action of hyaluronic acid in different immune organs and optimize its absorption pathway to improve bioavailability.

[0059] Example 3

[0060] Objective: To study the mesenteric lymph absorption pathway of 35kDa (>90% distribution range 10-70kDa) low molecular weight hyaluronic acid fragment HA35 after perfusion in the upper small intestine, and to explore the mechanism by which it enters the mesenteric lymph nodes and spleen after intestinal absorption.

[0061] Background: HA35 has been shown in clinical studies to have significant therapeutic effects on inflammatory wounds that are difficult to heal or do not heal. PET-CT imaging results show that 125 I-labeled HA35 can quickly enter the inguinal and popliteal lymph nodes and spleen within 5 minutes through subcutaneous injection at the limb terminals. In vitro studies have shown that HA35 can significantly promote the migration and spread of inflammatory cells such as neutrophils, lymphocytes and glial cells. However, the absorption pathway of HA35 through oral administration or small intestinal perfusion has not been clearly determined. This study used PET-CT technology with the help of 125 HA35 was labeled with I, and its absorption process in the mesenteric lymph nodes and spleen after perfusion through the upper small intestine was systematically explored, and a comparative study was conducted with high molecular weight hyaluronic acid HA.

[0062] method:

[0063] 1. 125 Preparation of I-labeled HA35: Prepare by the same iodination method as the original method. 125 I-labeled HA35. The hyaluronic acid fragment (HA35) was labeled with I-125, and the specific steps included CNBr activation of HA35, iodination by tyramide (TA), and radiochemical purity (RCP) detection. 125 I-HA35 was used for subsequent experiments.

[0064] 2. Small intestinal perfusion experimental design: Healthy C57BL / 6j mice, 6-8 weeks old, 5 mice per group. The experimental group was perfused with 20-25 μCi 125 I-HA35 and high molecular weight hyaluronic acid (HA) were used as controls. The small intestine perfusion method involved surgically exposing the upper small intestine of mice and using a microcatheter to 125 I-HA35 was slowly injected into the small intestine to simulate the oral absorption route.

[0065] 3. PET-CT imaging: Perform whole-body PET-CT scans of mice 2 hours, 4 hours, and 24 hours after perfusion to record 125 The distribution of I-HA35 in mice, focusing on the accumulation of radioactivity in organs such as mesenteric lymph nodes, spleen, and liver. After imaging, mice were euthanized, and tissues such as mesenteric lymph nodes, spleen, and liver were collected to measure the percent absorbed dose (%ID / g) of each tissue.

[0066] 4. Tissue analysis: Gamma-ray imaging was performed on tissue samples to confirm the absorption and distribution of radiolabeled HA35. ROI analysis was used to compare the radioactivity intensity of each lymph node and organ to evaluate the absorption pathway and distribution of HA35 after small intestinal perfusion.

[0067] result:

[0068] After small bowel perfusion, PET-CT imaging showed that 125 I-HA35 was rapidly absorbed and entered the lymphatic system within 2 hours of perfusion, and first appeared in the mesenteric lymph nodes. 125 I-HA35 accumulated significantly in the spleen and liver, and the intensity of radioactivity in the mesenteric lymph nodes and spleen was higher than that in the liver and soft tissues. 125 I-HA35 accumulation reached its peak in the mesenteric lymph nodes and spleen. ROI analysis showed that the radioactivity distribution intensity in the mesenteric lymph nodes and spleen was significantly higher than that in the liver and other major organs, indicating that HA35 was mainly absorbed through the mesenteric lymphatic system.

[0069] discuss:

[0070] This study is the first to clarify the mesenteric lymph absorption pathway of HA35 through the small intestinal perfusion method. Compared with subcutaneous injection, small intestinal perfusion can also effectively deliver HA35 to the lymphatic system, especially in the mesenteric lymph nodes, where there is significant accumulation. This is consistent with the results of subcutaneous injection studies in which HA35 quickly enters the lymph nodes, indicating that HA35 has good mesenteric lymphatic system transmission potential. The high radioactivity accumulation in the spleen also verifies the distribution characteristics of HA35 in lymphatic organs.

[0071] Compared with high molecular weight hyaluronic acid, HA35 is more easily absorbed through the small intestinal epithelium and enters the mesenteric lymphatic system due to its smaller molecular weight. This provides a theoretical basis for the clinical application of HA35 via oral administration or small intestinal perfusion, which may help develop new treatment methods and new indications.

[0072] in conclusion:

[0073] This study demonstrated 125 I-labeled HA35 can be effectively absorbed through the mesenteric lymphatic system after intestinal perfusion or oral administration, and transported into the blood through the spleen. The oral route provides an alternative and new indication for HA35 injection.

[0074] In addition, studies have found that the absorption pathway of the 70kDa (>90% of the distribution range is 40-100kDa) low molecular weight hyaluronic acid fragment HA70 is similar to that of the above-mentioned 35kDa (>90% of the distribution range is 10-70kDa) low molecular weight hyaluronic acid fragment HA35, which will not be repeated here.

[0075] Example 4

[0076] Objective: To orally treat back pain or myofasciitis with a low molecular weight hyaluronic acid fragment HA35 of 35 kDa (>90% of the distribution range 10-70 kDa).

[0077] Background: Back pain is mainly caused by myofasciitis. The main clinical feature of myofasciitis is tender trigger points. Clinical studies before this study have shown that local injection of 0.2g HA35 is effective in treating back pain or myofasciitis. This example studies the effect of oral administration of different doses of HA35 in treating back pain or myofasciitis.

[0078] Method: This example studied 20 patients with back pain, divided into two dosage groups. The first group had 5 males and 5 females, aged between 40-56 years old, and took HA35 orally at a dosage of 5 grams / time / day for 21 days; the second group had 5 males and 5 females, aged between 42-60 years old, and took HA35 at a dosage of 0.2 grams / time / day. This example also evaluated the degree of facial fat, facial skin and mental state of the patients. The numerical pain score (NPRS) was used to evaluate the pain and facial state of the patients, with a pain rating of 0-10 points, with 0 indicating no pain, no fat, the whitest skin and the most energetic, and 10 indicating the greatest degree of pain, the most fat, the darkest skin and the most listless.

[0079] Results: The results in Table 1 show that HA35 can effectively treat back pain by 50%-60% 2 hours after oral administration; 80%-90% after 2 days of oral treatment; and completely relieve the patient's back pain after 21 days of oral treatment. In contrast, oral administration of 0.2g of HA35 has no effect on pain relief. The results in Table 2 indicate that after 21 days of treatment, the fat on the face, the whiteness of the skin, and the spirit of vitality have all been significantly improved. After treatment, the patient's face became thinner by more than 50%, the skin became whiter by more than 50%, and the spirit of vitality increased by 50%.

[0080] Table 1. Pain scores before and after treatment of back pain

[0081]

[0082] Table 2. Facial fat degree and facial skin condition

[0083] Evaluation Metrics Before treatment After 21 days of treatment Facial fat level 6.0±2.0 2.5±1.0 Facial skin whiteness 5.5±2.5 2.0±1.5 Facial mental state 5.0±2.0 2.5±1.5

[0084] Note: The dosage of HA35 in Table 2 is 5 g / 1 time / day.

[0085] Discussion and conclusion: The results of this study showed that oral administration of HA35 can relieve back pain caused by myofasciitis in a dose-dependent manner. After 21 days of treatment, the pain was completely relieved by taking 5g of HA35 orally per day, while no significant effect was observed after 21 days of treatment by taking HA35 at a dose of 0.2g per day. By analyzing the time of pain relief, it can be seen that the pain was effectively relieved after 2 days or even 2h, indicating that HA35 can quickly and effectively treat back pain. The 21-day treatment cycle is for maintenance treatment of patients. Although the relief of back pain is rapid, the inflammation that causes back pain requires time to treat and heal. Since HA35 is dose-dependent, combined with our unpublished data, oral administration of 2.5g also has a therapeutic effect in people with light weight. In addition to the effect on back pain, after 21 days of treatment, we also observed changes in the patient's facial mental state, which was manifested as a reduction in facial fat, whitening of facial skin, and a good facial mental state, indicating that oral HA35 has the effects of face slimming, beauty, and whitening in addition to pain relief. Therefore, in summary, oral administration of 2.5g-5g of HA35 can quickly relieve and treat back pain caused by myofasciitis.

[0086] Example 5

[0087] Objective: This study aimed to evaluate the therapeutic effect of oral low-molecular-weight hyaluronic acid fragment HA35 (35 kDa, >90% distribution range 10-70 kDa) on shoulder pain, especially shoulder pain caused by shoulder myofasciitis.

[0088] Background: Shoulder pain is common in myofasciitis of the shoulder. The main clinical feature of myofasciitis is the presence of tender trigger points, which lead to pain and limited movement. Previous studies have shown that local injection of 0.2g HA35 can effectively relieve back pain and myofasciitis symptoms. However, it is not clear whether oral HA35 has the same therapeutic effect on shoulder pain. This study compares the effects of different doses of oral HA35 in the treatment of shoulder pain and explores its potential role in relieving shoulder pain.

[0089] Methods: Twenty patients with shoulder pain were selected for this study and divided into two groups according to the dose. The first group was a high-dose group, with 10 patients (5 males and 5 females), aged 40-56 years old, who took HA35 5 g / day orally for 21 days; the second group was a low-dose group, with 10 patients (5 males and 5 females), aged 42-60 years old, who took HA35 0.2 g / day orally for 21 days. Pain was assessed by the Numerical Pain Rating Scale (NPRS), which ranges from 0 to 10 points, with 0 indicating no pain and 10 indicating severe pain. The patients were evaluated before treatment, 2 hours after treatment, 2 days after treatment, and 21 days after treatment.

[0090] Results: Patients in the high-dose HA35 group (5 g / day) experienced 50%-60% pain relief 2 hours after oral administration; 80%-90% pain relief 2 days later; and shoulder pain completely disappeared after 21 days of treatment. In contrast, the shoulder pain scores of patients in the low-dose HA35 group (0.2 g / day) were almost unchanged, with no significant improvement, as shown in Table 3:

[0091] Table 3. Pain scores of patients with shoulder pain before and after treatment

[0092] HA35 dosage Before treatment 2 hours after treatment 2 days after treatment 21 days after treatment 5 g / 1 time / day 5.0±1.5 2.0±1.0 0.5±0.5 0.0±0.0 0.2 g / 1 time / day 5.5±2.0 5.0±2.5 5.5±1.5 5.0±2.0

[0093] Discussion and Conclusion:

[0094] The results of the study showed that oral HA35 can significantly relieve pain caused by shoulder myofasciitis in a dose-dependent manner. After 21 days of treatment with 5 grams of HA35 taken orally daily, shoulder pain completely disappeared, while patients who took 0.2 grams of HA35 daily did not experience pain relief. Patients in the high-dose group showed significant pain relief within 2 hours, showing that HA35 has a rapid analgesic effect, indicating that the drug may have advantages in acute pain management. Although pain relief is faster, the treatment and healing of inflammation takes longer, so a treatment cycle of at least 21 days is recommended.

[0095] This study also showed that the oral effect of HA35 is dose-related, and a lower dose (0.2 g / day) has no significant effect, suggesting that the oral dose needs to reach a certain level far beyond the norm (such as 5 g / day) to exert a significant analgesic effect. Comprehensive analysis shows that this study supports that daily oral administration of 5 g of HA35 can effectively relieve shoulder pain, and it has good safety and is suitable as an adjuvant therapy for the treatment of shoulder pain.

[0096] Example 6

[0097] Objective: This study aimed to evaluate the efficacy of oral administration of a low-molecular-weight hyaluronic acid fragment, HA35 (35 kDa, >90% distribution range 10-70 kDa), for the treatment of neck pain, particularly pain due to cervical myofasciitis.

[0098] Background: Neck pain is common in cervical myofasciitis, and patients often feel local pain, stiffness, and limited movement. The main feature of myofasciitis is tender trigger points, which seriously affect the quality of life of patients. Previous studies have shown that local injection of 0.2g HA35 can effectively relieve back pain and myofasciitis symptoms. However, it is unclear whether oral HA35 can produce similar therapeutic effects on neck pain. This study compared the effects of different doses of oral HA35 in the treatment of neck pain and explored its potential application in relieving neck pain.

[0099] Methods: Twenty patients with neck pain were enrolled in this study and divided into two groups according to the dose. The first group was a high-dose group, with 10 patients (5 males and 5 females), aged 40-56 years old, who took HA35 5 g / time / day orally for 21 consecutive days; the second group was a low-dose group, with 10 patients (5 males and 5 females), aged 42-60 years old, who took HA35 0.2 g / time / day orally for 21 consecutive days. Pain was assessed using the Numerical Pain Scale (NPRS), with a score range of 0-10, with 0 indicating no pain and 10 indicating severe pain. The assessment time points were before treatment, 2 hours after treatment, 2 days after treatment, and 21 days after treatment.

[0100] Results: Patients in the high-dose HA35 group (5 g / day) experienced 50%-60% pain relief within 2 hours after oral administration; 80%-90% pain relief after 2 days; and neck pain completely disappeared after 21 days of treatment. In contrast, the pain scores of patients in the low-dose HA35 group (0.2 g / day) did not change significantly at each time point, and no significant pain relief effect was observed. See Table 4 for details:

[0101] Table 4. Pain scores of patients with neck pain before and after treatment

[0102] HA35 dosage Before treatment 2 hours after treatment 2 days after treatment 21 days after treatment 5 g / 1 time / day 5.0±1.5 2.0±1.0 0.5±0.5 0.0±0.0 0.2 g / 1 time / day 5.5±2.0 5.0±2.5 5.0±2.0 5.0±2.0

[0103] Discussion and Conclusion:

[0104] The results of the study showed that oral HA35 can significantly relieve neck pain caused by myofasciitis, and showed a clear dose-dependency. After 21 days of treatment with 5 grams of HA35 per day, the patient's neck pain completely disappeared; while oral administration of 0.2 grams of HA35 did not result in effective pain relief. The high-dose HA35 group showed rapid pain relief within 2 hours, indicating that HA35 has significant therapeutic potential for acute pain. Although pain relief is rapid, it takes longer to treat the inflammatory process associated with neck pain, and a treatment cycle of at least 21 days is recommended to ensure full recovery.

[0105] This study also showed that oral HA35 has good safety, good patient tolerance, and no obvious side effects. Compared with low doses, high doses (5 grams / day) of HA35 can more effectively relieve neck pain and are suitable for the management of long-term neck pain. Comprehensive analysis shows that oral administration of 5 grams of HA35 can be used as an effective adjuvant therapy for the treatment of neck pain, with the dual effects of rapid analgesia and long-term relief.

[0106] Example 7

[0107] Objective: This study aimed to evaluate the therapeutic effect of oral administration of a low-molecular-weight hyaluronic acid fragment, HA35 (35 kDa, >90% distribution range 10-70 kDa), on postherpetic neuralgia.

[0108] Background: Postherpetic neuralgia (PHN) is the most common chronic complication after herpes zoster infection, especially in elderly patients. PHN is characterized by persistent or intermittent severe pain, which seriously affects the quality of life of patients. Currently, there are limited treatments for PHN. Conventional analgesics such as antidepressants, antiepileptic drugs and opioids can relieve pain in some patients, but they are accompanied by side effects and have limited effects. In recent years, the analgesic effect of hyaluronic acid HA35 in various neuropathic pain has attracted attention, especially its analgesic mechanism related to TRPV1 channels may be effective for PHN patients. This study evaluated the efficacy of different doses of oral HA35 in PHN patients and explored its potential as a new treatment.

[0109] Methods: A total of 10 patients with postherpetic neuralgia were included in this study and divided into two groups according to the dosage. The first group was the high-dose group, with 5 patients aged 40-70 years old, who took HA35 5 g / time / day orally for 21 consecutive days; the second group was the low-dose group, with 5 patients aged 42-68 years old, who took HA35 0.2 g / time / day orally for 21 consecutive days. The pain level of the patients was assessed using the Numerical Pain Scale (NPRS), with a score range of 0-10, with 0 indicating no pain and 10 indicating severe pain. The assessment time points were before treatment, 1 day after treatment, 2 days after treatment, and 21 days after treatment.

[0110] result

[0111] Patients in the high-dose HA35 group (5 g / day) experienced 30%-40% pain relief within 1 day of oral administration; 60%-70% pain relief after 2 days; and the pain almost completely disappeared after 21 days of treatment. In contrast, the pain scores of patients in the low-dose HA35 group (0.2 g / day) did not change much at each time point, and no significant relief effect was observed. See Table 5 for details:

[0112] Table 5. Pain scores of patients with postherpetic neuralgia before and after treatment

[0113] HA35 dosage Before treatment 1 day after treatment 2 days after treatment 21 days after treatment 5 g / 1 time / day 7.0±1.2 4.0±1.0 3.0±0.8 0.5±0.5 0.2 g / 1 time / day 6.5±1.5 6.5±1.2 6.0±1.3 6.0±1.2

[0114] Discussion and Conclusion:

[0115] The results of the study showed that oral HA35 can significantly relieve postherpetic neuralgia (PHN) in a dose-dependent manner. Patients who took 5 grams of HA35 orally daily experienced significant pain relief within 1 day of treatment, and the pain was further reduced after 2 days. At the end of the 21-day treatment, the patient's pain almost completely disappeared. This shows that HA35 has a rapid and sustained analgesic effect and is particularly suitable for the treatment of chronic and intractable neuralgia such as PHN. In contrast, patients who took 0.2 grams of HA35 orally did not experience significant pain relief, showing the ineffectiveness of lower doses.

[0116] The rapid analgesic effect of HA35 may be related to its mechanism of action on the TRPV1 channel, which is closely related to neuropathic pain. Since the pain of PHN patients is long-term and difficult to relieve, the effects of existing treatment options are often limited, and HA35 provides an alternative without obvious side effects, showing better tolerability and safety. Especially for patients who have long-term use of opioids or other analgesics and have dependence or side effects, oral HA35 provides a safe option.

[0117] In summary, oral administration of 5 grams of HA35 has a significant therapeutic effect on postherpetic neuralgia, can quickly and effectively relieve pain, and provide patients with long-term analgesic effects. It is a promising auxiliary treatment method.

[0118] Example 8

[0119] Objective: This study aimed to evaluate the efficacy of oral administration of a low-molecular-weight hyaluronic acid fragment, HA35 (35 kDa, >90% distribution range 10-70 kDa), on the relief of bone pain in patients with advanced anorectal cancer, especially persistent pain caused by bone metastasis.

[0120] Background: Patients with advanced anorectal cancer often have bone metastases, which lead to persistent and unbearable bone pain. Cancer-related bone pain is a complex pain sensation that is often difficult to fully control with conventional analgesics. Patients usually need to rely on opioids for a long time to relieve pain, but these drugs may cause serious side effects such as drug dependence, constipation, and nausea. In addition, traditional nonsteroidal anti-inflammatory drugs (NSAIDs) are also limited in their safety for long-term use. In recent years, the low-molecular hyaluronic acid fragment HA35 has shown potential in the treatment of neuropathic pain and cancer-related pain. It provides effective analgesic effects by interacting with TRPV1 channels. This study aims to explore the pain-relieving effect of oral HA35 on patients with advanced bone pain in anorectal cancer.

[0121] Methods: This study included 10 patients with advanced anorectal cancer, all of whom had clear bone metastases and ranged in age from 50 to 75 years. The patients were divided into two groups according to the dose. The first group was the high-dose group, with 5 patients taking HA35 5 g / time / day orally for 21 consecutive days; the second group was the low-dose group, with 5 patients taking HA35 0.2 g / time / day orally for 21 consecutive days. The pain level was assessed by the Numerical Pain Scale (NPRS), with a score range of 0-10 points, 0 for no pain and 10 for severe pain. The assessment time points included before treatment, 1 day after treatment, 2 days after treatment, and 21 days after treatment.

[0122] Results: The patients in the high-dose HA35 group (5 g / day) experienced 20%-30% pain relief within 1 day after oral administration, and further 40%-50% pain relief after 2 days; after 21 days, the pain stabilized at a 30%-45% relief level. Although the pain did not completely disappear, the patients reported a significant reduction in pain and a reduced impact on their daily lives. The pain scores of the patients in the low-dose group (0.2 g / day) did not change much, and no significant relief was observed. See Table 6 for details:

[0123] Table 6. Pain scores before and after treatment in patients with advanced anorectal cancer

[0124] HA35 dosage Before treatment 4 hours after treatment 2 days after treatment 21 days after treatment 5 g / 1 time / day 8.5±1.0 3.0±0.8 2.0±0.7 1.0±0.8 0.2 g / 1 time / day 8.0±1.2 7.8±1.0 7.5±1.1 7.4±1.0

[0125] Discussion and Conclusion:

[0126] The results of the study showed that oral HA35 can significantly relieve bone pain in advanced anorectal cancer, especially in patients in the high-dose group (5 g / day), where pain was reduced in a short period of time and maintained at a controllable level after 21 days of treatment. Although the pain did not completely disappear, the patients' pain intensity was significantly reduced compared to baseline, which improved their quality of life and reduced their dependence on opioids. In contrast, no significant analgesic effect was observed in the low-dose HA35 group (0.2 g / day).

[0127] The analgesic effect of HA35 may be related to its mechanism of alleviating neuropathic pain through the TRPV1 channel. In cancer-related bone pain, the activity of the TRPV1 channel is closely related to the perception of pain, and HA35, as a non-hormonal substance, provides effective analgesia without significant side effects, indicating that its safety and tolerability in long-term treatment are better than traditional analgesics.

[0128] In summary, oral administration of 5 grams of HA35 has a significant analgesic effect in patients with advanced anorectal cancer bone pain. It can maintain the pain at a controllable level without completely eliminating it, and reduce dependence on conventional analgesics. This provides a new and safe adjuvant treatment option for patients with advanced cancer bone pain.

[0129] Example 9

[0130] Objective: This study aimed to evaluate the efficacy of oral administration of a low-molecular-weight hyaluronic acid fragment, HA35 (35 kDa, >90% distribution range 10-70 kDa), on pain relief in patients with advanced lung cancer.

[0131] Background: Lung cancer is one of the malignant tumors with the highest cancer-related mortality worldwide. Especially in advanced patients, cancer cells often metastasize to bones, causing severe bone pain. This bone pain is a complex cancer-related pain. Although conventional opioids can relieve some pain, their side effects such as drug dependence, nausea, and constipation limit their long-term use. In recent years, the analgesic effect of hyaluronic acid fragment HA35 in various neuropathic and cancer-related pain has been preliminarily verified due to its interaction with TRPV1 pain receptor. This study aims to evaluate the pain relief effect of oral HA35 in patients with advanced lung cancer bone metastasis.

[0132] Methods: A total of 10 patients with advanced lung cancer and bone metastasis were included in this study, with an age range of 50 to 75 years. The patients were divided into two groups: the first group was a high-dose group, with 5 patients taking HA35 5 g / time / day orally for 21 consecutive days; the second group was a low-dose group, with 5 patients taking HA35 0.2 g / time / day orally for 21 consecutive days. The pain level was assessed using the Numerical Pain Scale (NPRS), with a score range of 0-10, with 0 indicating no pain and 10 indicating extremely severe pain. The assessment time points included before treatment, 1 day after treatment, 2 days after treatment, and 21 days after treatment.

[0133] Results: The patients in the high-dose HA35 group (5 g / day) had pain relief of about 20%-30% within 1 day after taking it; the pain relief reached 40%-50% after 2 days; after 21 days of continuous treatment, the pain was maintained at a relief level of 30%-40%. Although it did not completely disappear, the patients reported that the pain was significantly reduced and they were able to carry out daily activities better. The pain scores of patients in the low-dose group (0.2 g / day) remained almost unchanged during the treatment period. See Table 7 for details:

[0134] Table 7. Pain scores of patients with advanced lung cancer before and after treatment

[0135] HA35 dosage Before treatment 1 day after treatment 2 days after treatment 21 days after treatment 5 g / 1 time / day 6.0±1.0 3.0±0.8 2.5±0.7 0.0±0.8 0.2 g / 1 time / day 5.5±1.2 5.2±1.0 5.0±1.1 5.0±1.0

[0136] Discussion and Conclusion:

[0137] Studies have shown that oral HA35 can effectively relieve pain in patients with advanced lung cancer bone metastasis, especially in the high-dose group (5 g / day). The pain was significantly reduced within 1 day of treatment, significantly relieved within 2 days, and the pain was under long-term control after 21 days. Although the pain cannot be completely eliminated, the patient's pain level is maintained at a low level, reducing dependence on opioids and improving the quality of life. However, the pain scores of patients in the low-dose group (0.2 g / day) changed little, and no significant relief effect was observed.

[0138] The analgesic mechanism of HA35 may be related to its effect on TRPV1 pain receptors, which play an important role in cancer-related bone pain. Compared with conventional opioid analgesics, HA35 has no obvious drug dependence or other serious side effects, so it shows better safety and tolerability in long-term treatment.

[0139] The results of this study showed that daily oral administration of 5 grams of HA35 has a significant effect on relieving pain caused by bone metastasis of advanced lung cancer without significant side effects. Although the pain cannot be completely eliminated, HA35 can control the pain within a tolerable range and reduce the patient's need for other analgesics, providing a new and potentially effective adjuvant treatment option for patients with bone pain in the advanced stage of lung cancer.

[0140] Example 10

[0141] Objective: This study aimed to evaluate the efficacy of oral administration of a low-molecular-weight hyaluronic acid fragment, HA35 (35 kDa, >90% distribution range 10-70 kDa), on pain relief after radiotherapy in patients with different cancers, especially cancer-related pain associated with bone metastasis.

[0142] Background: Radiotherapy is an important method in the treatment of various cancers. It is often used in a variety of tumors such as lung cancer, breast cancer, and prostate cancer, especially in patients with advanced cancer and concomitant bone metastases. However, radiotherapy often causes radiation-related pain, including bone pain, neuropathic pain, and local tissue inflammation. Conventional analgesic methods such as opioids and nonsteroidal anti-inflammatory drugs (NSAIDs), although effective in the short term, can lead to a series of side effects such as drug dependence and gastrointestinal complications with long-term use. Therefore, exploring safer and more effective analgesic alternatives is of great clinical significance. Low-molecular hyaluronic acid fragment HA35 has shown an alleviating effect in neuropathic pain and cancer pain by interacting with TRPV1 pain receptors without obvious side effects. This study investigated the efficacy of HA35 in post-radiotherapy pain in patients with different cancers.

[0143] Methods: A total of 10 patients with different types of cancer after radiotherapy were included in this study. All patients had varying degrees of bone metastasis, including lung cancer, breast cancer, and prostate cancer patients. The patients were randomly divided into two groups: a high-dose group and a low-dose group. Five patients in the high-dose group took HA35 5 g / time / day orally for 21 consecutive days; five patients in the low-dose group took HA35 0.2 g / time / day orally for 21 consecutive days. The pain level was assessed by the Numerical Pain Scale (NPRS), with a score range of 0-10, with 0 indicating no pain and 10 indicating severe pain. The assessment time points included before treatment, 1 day after treatment, 2 days after treatment, and 21 days after treatment.

[0144] Results: The pain of patients in the high-dose HA35 group (5 g / day) was reduced by about 15%-20% within 4 hours after treatment; the pain was relieved by about 30%-40% after 2 days; the pain score was maintained at a relief level of 40%-50% after 21 days. Although the pain did not completely disappear, the patients reported that the pain was significantly reduced and the quality of life was improved. The pain score of patients in the low-dose group (0.2 g / day) hardly changed during the treatment period, and no significant relief was observed. The specific results are shown in Table 8 below:

[0145] Table 8. Pain scores of patients with cancer bone metastasis pain before and after treatment

[0146]

[0147] Discussion and Conclusion:

[0148] The results of this study showed that oral HA35 can effectively relieve pain after radiotherapy in patients with different types of cancer, especially long-term pain caused by bone metastasis. The pain of patients in the high-dose group (5 g / day) was relieved within 1 day after treatment, and the pain continued to decrease after 2 days. After 21 days of treatment, the pain remained at a low level, showing the sustained analgesic effect of HA35. No obvious analgesic effect was observed in the low-dose group (0.2 g / day), indicating that the analgesic effect of HA35 was dose-dependent.

[0149] HA35 may play an important role in alleviating cancer-related neuropathic pain and bone pain by interacting with the TRPV1 channel. Compared with traditional opioids and nonsteroidal anti-inflammatory drugs, HA35 has no side effects such as drug dependence and gastrointestinal discomfort, making it suitable as an adjuvant therapy for long-term pain management. In addition, the use of HA35 has no obvious side effects and does not affect the patient's other treatment options.

[0150] In summary, oral high-dose HA35 (5 g / day) has a good effect on relieving pain after radiotherapy in patients with different types of cancer, especially for pain control in patients with bone metastasis. As a safe and effective analgesic alternative, HA35 provides a new approach to pain management after cancer radiotherapy.

[0151] Embodiment 11

[0152] Objective: To evaluate the efficacy of oral HA35 (5 g / 2 times / day) on pain relief in patients before and after dental implant surgery.

[0153] Background: After dental implant surgery, patients often face a large postoperative pain problem, especially elderly patients, who may experience a stronger pain response due to poor periodontal health and decreased body recovery ability. Traditional analgesics, such as nonsteroidal anti-inflammatory drugs (NSAIDs), are effective, but long-term use may cause side effects such as gastrointestinal discomfort. HA35 has been shown to reduce Ca2+ influx by inhibiting TRPV1 channels and relieve pain. Clinically, it has been shown that injection of HA35 can relieve inflammatory pain in the shoulder, neck, back and temporal regions, neuropathic pain of herpes zoster, cancer pain and bone metastasis pain. HA35 has a significant analgesic effect on open organs. Therefore, in order to study the analgesic effect of oral HA35. We conducted a study on the treatment of postoperative pain after dental implant surgery to explore whether oral hyaluronic acid can be used as an alternative or adjuvant treatment to relieve postoperative pain after dental implant surgery and reduce the use of traditional analgesics.

[0154] method:

[0155] 1. Study subjects: Two 55-year-old male patients who were scheduled for dental implant surgery. Both patients had no other serious underlying diseases and were similar in periodontal health and general health status.

[0156] 2. Experimental design: Each patient undergoing dental implant surgery was divided into two groups. One group took HA35 orally at a dose of 5g twice a day, starting from 1 day before surgery and continuing until 3 days after surgery. The other group took acetaminophen according to the dosage after surgery. Both groups took antibiotics to prevent inflammatory infection.

[0157] 3. Pain assessment method: The pain perception of patients on the 1st, 2nd and 3rd day after the two surgeries was recorded using the National Pain Rating Scale (NPRS). The NPRS score ranged from 0 (no pain) to 10 (severe pain).

[0158] 4. Statistical analysis: Paired t-test was used to compare the pain scores of patients taking HA35 at different time points, and the significance level was set at p < 0.05.

[0159] Results: Pain score (VAS): The pain scores of patients taking HA were significantly lower than those of the control group on the first, second and third days after surgery. On the third day, the pain scores of both groups were 0, with no significant difference. NSAIDs use: The dose of NSAIDs used within 24 hours after surgery in patients taking HA35 was significantly lower than that in the control group. The average dose of acetaminophen used within 24 hours after surgery in the HA35 group was 0.3 g, while that in the control group was 0.9 g; from the first to the third day after surgery, the average number of acetaminophen used in the HA35 group was 3 times, while that in the control group was 15 times (p<0.01); from the third day onwards, patients in the HA35 group stopped using acetaminophen.

[0160] Table 9. Pain scores of patients undergoing dental implant surgery before and after treatment

[0161]

[0162] Discussion and Conclusion:

[0163] This study shows that oral HA35 (5g / 2 times / day) can effectively reduce the pain patients feel after dental implant surgery and significantly reduce the need for NSAIDs after surgery. On the one hand, HA35 promotes lymphatic return and reduces inflammatory response by promoting the diffusion of lymphocytes; on the other hand, it relieves pain by regulating the activation of TRPV1 channels. At the same time, HA35 promotes postoperative tissue healing and reduces the duration of pain. HA35 is highly safe for clinical use and well tolerated by patients. Adverse reactions may include lowered blood pressure and itching. This provides clinical support for oral hyaluronic acid as an auxiliary pain management solution, especially for patients who cannot take NSAIDs for a long time, such as elderly patients with gastrointestinal discomfort or cardiovascular risks. HA35 may provide a more ideal alternative treatment method.

[0164] Oral administration of HA35 (5 g / 2 times / day) can effectively relieve pain after dental implant surgery and reduce the use of NSAIDs after surgery, with significant pain management effects. As a safe and effective adjuvant treatment, hyaluronic acid has the potential for application in oral surgery. Future research should be devoted to exploring its wider clinical application to improve the postoperative rehabilitation experience of elderly patients.

[0165] Example 12

[0166] Objective: To evaluate the efficacy of oral HA35 (5 g / time / day) in relieving postoperative pain in female patients before and after transvaginal oocyte retrieval (TVOR) surgery.

[0167] Background: Although TVOR is the least invasive procedure for assisted reproductive technology (ART) such as in vitro fertilization (IVF), complications caused by the procedure are not uncommon. Pain is the most common complication, so adequate analgesia is required before and after the operation. The difference between anesthesia for TVOR surgery and most surgeries is that the effect on oocyte quality needs to be considered. Therefore, conscious sedation and rectal administration of diclofenac sodium suppositories are preferred with minimal side effects and no difference in pregnancy rate. However, the difference in analgesic effect and gastrointestinal side effects also hinder the use of some patients. Clinically, HA35 has been used as an analgesic substance to relieve inflammatory pain in the shoulder, neck, back and temporal regions, neuropathic pain associated with herpes zoster, cancer pain, and bone metastasis pain. It has significant effects, no side effects, and can reduce the side effects of hormonal drugs. Analysis of its effective analgesic mechanism may be related to the capsaicin pain receptor TRPV1 channel.

[0168] method:

[0169] 1. Research subject: The research subject was a 40-year-old female patient who underwent two egg retrieval operations. The patient was in good general health and had no obvious underlying diseases. The two egg retrieval operations were separated by ten months.

[0170] 2. Experimental design: Before the first egg retrieval surgery, the patient was given diclofenac sodium suppository for anesthesia and antibiotics. Before the second egg retrieval surgery, the patient took HA35 (5g / time / day) orally every day from 2 days before the surgery, and continued to take it after the surgery until 3 days after the surgery. The second surgery patient was also given diclofenac sodium suppository for anesthesia and antibiotics.

[0171] Results: Pain scores (NPRS) As shown in Table 10, after the second surgery, patients in the oral HA35 group had significantly lower postoperative pain scores than those in the first control surgery.

[0172] Table 10. Pain scores of patients undergoing egg retrieval before and after the two surgeries

[0173] Operation 0h after surgery 2 hours after surgery 3 hours after surgery 6 hours after surgery First surgery 6 4 3 2 Second surgery 4 1 1 0

[0174] Discussion and Conclusion:

[0175] Studies have shown that the most common complication after transvaginal oocyte retrieval (TVOR) is pain, and some patients experience severe or even extreme pain after surgery. Anesthesia is indispensable to relieve pain caused by surgery, but traditional anesthetics may affect the quality of oocytes, which in turn has an adverse effect on fertilization and implantation rates.

[0176] In this study, the patient used local anesthesia during both egg retrieval operations due to low oocyte quality and quantity, and used diclofenac sodium suppositories to relieve pain. However, after using diclofenac sodium suppositories alone, the patient still experienced gastrointestinal discomfort and significant pain. Therefore, in the second operation, HA35 was combined with diclofenac sodium suppositories to achieve better analgesic effect. HA35 is a low molecular weight hyaluronic acid with significant analgesic and anti-inflammatory effects, and its mechanism may be related to the capsaicin receptor TRPV1 channel.

[0177] The results of the study showed that the combined use of HA35 can significantly reduce postoperative pain scores, accelerate wound healing, and reduce patient discomfort. Compared with the single use of traditional analgesics, HA35 has higher safety and no obvious side effects. This study provides new ideas for the application of HA35 in TVOR surgery, showing its potential as an auxiliary analgesic drug, which can effectively improve postoperative recovery and improve the quality of life of patients.

[0178] Embodiment 13

[0179] Objective: This study aimed to evaluate the therapeutic effect of oral administration of a low-molecular-weight hyaluronic acid fragment, HA35 (35 kDa, >90% distribution range 10-70 kDa), on muscle strain, muscle stiffness, and physical weakness caused by maintaining the same posture in men aged 55-70 years.

[0180] Background: Hyaluronic acid (HA) as an oral supplement has been developed and tested in clinical trials by several Japanese, American and European companies. Studies have shown that patients with moderate knee osteoarthritis treated with HA have significant improvements in the short term (1-4 months), especially in VAS, WOMAC, JKOM and SF-36v2 scores. However, there is still controversy about the absorption of oral HA. Some studies have found that the high molecular weight HA of 900kDa cannot be directly absorbed by the human body, but is degraded into oligosaccharides by intestinal bacteria, which are then absorbed through the large intestine and distributed to tissues throughout the body. The 2016 study further demonstrated that oligosaccharide HA can be effectively distributed to the skin and other parts.

[0181] As they age, older men often suffer from muscle strain and stiffness due to maintaining the same posture for a long time, which leads to limited movement, decreased physical strength and shortened walking time. In addition, problems such as high blood pressure are also common in the elderly, further affecting their health. HA has shown efficacy in the repair of various soft tissue injuries due to its anti-inflammatory and lubricating effects. In particular, the low molecular weight HA35 fragment may provide an effective treatment option for the elderly by relieving muscle stiffness and improving mobility.

[0182] Methods: This study included 4 male subjects aged between 55 and 70 years old, all of whom had symptoms such as muscle strain, stiffness, and inflexible hands and feet caused by maintaining the same posture. Two subjects were included in the experimental group, taking HA 355g orally twice a day for 2 days, then switching to 5g once a day for 19 days. The remaining 2 subjects were included in the control group, taking 0.1g orally twice a day for 21 consecutive days. The evaluation indicators included muscle stiffness, flexibility of hands and feet, agility of bending over to pick up objects, prolonged walking time, and improvement in physical strength. The evaluation time points were before treatment, 2 days, 7 days, and 21 days after treatment.

[0183] Results: After taking HA35 orally for 2 days, the subjects reported that muscle stiffness was significantly relieved, the hands and feet were more flexible, and the agility of bending down to pick up objects was improved. The results in Table 11 show that the walking time was extended by an average of 15%, and the physical strength was significantly improved. In addition, the high blood pressure (systolic blood pressure) of all subjects decreased by about 10mmHg, from 145±10mmHg before treatment to 135±8mmHg. After taking it for 21 days, muscle stiffness was further relieved, the walking time was extended by an average of 25%, and the physical strength was significantly improved.

[0184] However, some subjects reported mild side effects, especially abdominal skin itching after taking high doses. In addition, subjects with low blood pressure experienced a further drop in blood pressure, such as a drop from 100 mmHg to 90 mmHg, accompanied by a mild feeling of weakness.

[0185] Table 11. Scores of elderly patients with limited activity in the experimental group before and after treatment

[0186]

[0187] Discussion and Conclusion:

[0188] This study shows that oral HA35 has a significant effect on relieving muscle strain and stiffness caused by maintaining the same posture in men aged 55-70. After taking HA35 for 2 days, the subjects' flexibility of hands and feet, agility of bending over to pick up objects were significantly improved, walking time was prolonged, and physical strength increased, indicating that HA35 can effectively relieve the discomfort symptoms related to muscle strain in a short period of time. In addition, HA35 can also effectively reduce the systolic blood pressure of hypertensive subjects, with an average reduction of 10mmHg, which has a certain auxiliary therapeutic effect for patients with hypertension.

[0189] However, potential side effects of HA35 were also found in the study. When taken at high doses, some subjects reported itchy abdominal skin. For subjects with lower blood pressure, blood pressure dropped further after taking HA35, accompanied by a mild sense of weakness. This suggests that special attention should be paid to patients with low blood pressure when using HA35, and the dosage should be adjusted appropriately.

[0190] In summary, oral HA35 (5 g / 2 times for 2 days, 5 g / time for 19 days) can effectively relieve muscle stiffness and mobility problems caused by maintaining the same posture in elderly men, enhance physical strength, and has a certain blood pressure lowering effect. For patients with low blood pressure, HA35 should be used with caution to avoid further drop in blood pressure.

[0191] Embodiment 14

[0192] Objective: This study aimed to evaluate the effect of oral administration of a low-molecular-weight hyaluronic acid fragment, HA35 (35 kDa, >90% distribution range 10-70 kDa), in relieving oral irritation, alcohol discomfort, and drunkenness symptoms after hangover, and its effect on improving abdominal distension and digestive discomfort after continuous administration for 21 days.

[0193] Background: Drinking alcohol is often accompanied by oral burning, spicy pain and stomach discomfort, which is closely related to alcohol irritation to the mucosa and gastrointestinal reactions. In addition, symptoms such as dizziness, nausea, and abdominal distension may occur after being drunk, and long-term and frequent drinking will increase the burden on the gastrointestinal tract. Existing alcohol sobering products mainly focus on reducing the degree of drunkenness, but have limited effects on the spicy pain in the mouth and abdominal distension after sobering up. Low-molecular-weight hyaluronic acid fragment HA35 has been studied in the regulation of inflammation and neuropathic pain, showing that it relieves pain and mucosal discomfort through mechanisms such as TRPV1 channels. This study will evaluate its role in sobering up and symptoms after drinking.

[0194] Methods: A total of 20 healthy subjects aged between 25 and 50 years were included in this study. All subjects had experienced oral pain and abdominal distension after drinking. All subjects took HA35 (5 g / time, 21 times / day) for 21 days. The subjects were evaluated for relief of oral pain immediately after drinking and improvement of drunkenness symptoms. In addition, changes in post-drinking symptoms such as abdominal distension and digestive discomfort were evaluated after taking it for 21 consecutive days. The evaluation indicators included the pain score, drunkenness discomfort score and abdominal distension score, which were evaluated using a numerical rating scale (NRS), with 0 points indicating no discomfort and 10 points indicating the most severe discomfort.

[0195] Results: After taking HA35, the 20 subjects were relieved from oral irritation and discomfort after drinking. The results in Table 12 show that high-dose oral HA35 5g / time can significantly reduce oral irritation within 1-5 minutes after taking it, and the relief rate of irritation is 60%. The score of discomfort from drunkenness is also reduced, with a relief rate of 50%. After taking HA35 for 21 consecutive days, the subjects' abdominal distension score on the second day after drinking was significantly improved, with an improvement rate of 55%, and the discomfort of abdominal distension continued to be relieved.

[0196] Table 12. Scores of indicators related to hangover

[0197] Evaluation Metrics Before treatment 5 minutes after taking 2 days after taking 21 days after taking Pain Rating 7.5±1.2 3.0±1.0 - - Intoxication Discomfort Score 8.0±1.0 3.0±0.9 - - Abdominal distension score 5.5±1.3 - 3.0±1.2 2.0±1.0

[0198] Discussion and Conclusion:

[0199] The results of this study showed that oral HA35 has a significant effect on relieving the immediate oral irritation and discomfort of drunkenness after drinking. After taking 5 grams of HA35, the subjects' oral irritation and drunkenness symptoms were significantly alleviated within 1-5 minutes, showing a rapid analgesic and hangover-relieving effect. The subjects who took it for 21 consecutive days had a significant improvement in abdominal distension and digestive discomfort on the second day after drinking, indicating that HA35 also has a significant effect in regulating the gastrointestinal mucosa and relieving abdominal distension.

[0200] HA35 may play a role in relieving alcohol irritation to the oral cavity and gastrointestinal tract by interacting with the TRPV1 channel. Compared with traditional hangover methods, HA35 can not only quickly relieve oral irritation and pain, but also significantly improve abdominal distension and discomfort after drinking, showing good safety and effectiveness. Therefore, HA35 has the potential to be used as a hangover and post-drinking relief product, especially for patients with oral mucosal discomfort and abdominal distension caused by frequent drinking.

[0201] Embodiment 15

[0202] Objective: This study aimed to evaluate the effects of oral administration of a low-molecular-weight hyaluronic acid fragment, HA35 (35 kDa, >90% distribution range 10-70 kDa), on the resolution of skin problems.

[0203] Background: With aging, the skin gradually loses its elasticity and radiance and is prone to various inflammatory skin problems, such as erythema, yellow spots, and tiny inflammatory wounds. These phenomena are closely related to the weakening of the skin barrier function and the enhancement of the inflammatory response. Hyaluronic acid (HA) is an important moisturizing component of the skin, which can maintain the skin barrier function and regulate the inflammatory response. Low-molecular-weight hyaluronic acid HA35 has been shown to have significant effects in reducing inflammation and promoting tissue repair. Therefore, this study explored the effects of oral HA35 on the skin health of middle-aged women and evaluated its efficacy in improving skin appearance and relieving skin inflammation.

[0204] Methods: This study included 20 female subjects aged between 45 and 55 years old, all of whom had varying degrees of skin inflammation, such as erythema, yellow spots, and tiny inflammatory wounds. All subjects took HA35 5 grams orally every day for 21 consecutive days. Evaluation indicators included skin moisture content, elasticity, glossiness, and improvement of inflammatory symptoms. Moisture and elasticity were measured using a professional skin measurement instrument, and the number and severity of inflammatory red and yellow spots and wounds were evaluated using a numerical rating system (NRS), with a score range of 0-10 points, 0 indicating no inflammation or wounds, and 10 indicating the most severe inflammation. The evaluation time points were before treatment, 7 days after taking it, and 21 days after taking it.

[0205] Results: After taking HA35 orally for 7 days, the subjects' skin moisture, elasticity and glossiness were improved, and inflammatory symptoms such as red and yellow spots and tiny inflammatory wounds were significantly reduced. The results in Table 13 show that the skin inflammation score decreased from 6.5±1.2 before treatment to 4.0±1.0 after 7 days, and further decreased to 2.0±0.8 after 21 days, indicating that HA35 has a significant effect in reducing skin inflammation. The skin elasticity and glossiness scores increased by about 20% after 7 days and by about 40% after 21 days.

[0206] Table 13. Ratings of patients with skin problems before and after treatment

[0207] Evaluation Metrics Before treatment 7 days after treatment 21 days after treatment Skin inflammation score 6.5±1.2 4.0±1.0 2.0±0.8 Skin elasticity improvement rate 0％ 20％ 40％ Skin gloss improvement rate 0％ 18％ 38％

[0208] Discussion and Conclusion:

[0209] The results of this study showed that oral HA35 has a significant cosmetic effect on the skin of women aged 45-55, especially in reducing skin inflammation. After taking it for 7 days, the subjects' skin inflammation symptoms (such as erythema, yellow spots and micro-wounds) were significantly reduced, and skin moisture and elasticity were also improved. After taking it for 21 days, skin inflammation was further alleviated, and skin elasticity and glossiness were more significantly improved. The subjects generally reported that their skin condition had improved significantly.

[0210] The anti-inflammatory mechanism of HA35 may be related to its regulation of skin immune response and repair of barrier function. Unlike topical hyaluronic acid products, oral HA35 can act more comprehensively on the whole body, combining internal and external factors to improve skin health. In addition, oral HA35 has no obvious side effects, and the subjects had no adverse reactions during the entire treatment process, demonstrating its safety as a beauty product.

[0211] Therefore, oral administration of HA35 (5 g / time for 21 consecutive days) can not only effectively reduce skin inflammation, but also improve the skin's moisture content, elasticity and glossiness, and has broad potential for beauty and anti-aging applications.

[0212] Example 16

[0213] Objective: This study aimed to evaluate the effect of oral administration of a low-molecular-weight hyaluronic acid fragment, HA35 (35 kDa, >90% distribution range 10-70 kDa), on dissolving subcutaneous fat, slimming the face, improving neck skin laxity, and reducing abdominal and leg fat in women aged 45-55 years within 14 to 21 days.

[0214] Background: With aging, especially in middle age, women's skin gradually becomes loose, and fat accumulation in the face, neck, and other parts of the body increases significantly, affecting appearance and self-confidence. Increased subcutaneous fat is often accompanied by loose skin, especially in the face, neck, and abdomen. Although traditional lipolysis methods such as surgery and topical products are effective, they are invasive and have side effects. In recent years, hyaluronic acid has not only been favored for its moisturizing and anti-inflammatory effects, but also has shown potential in fat metabolism and skin tightening. This study investigated the efficacy of oral HA35 in improving subcutaneous fat accumulation, skin laxity, and face slimming.

[0215] Methods: Twenty female subjects aged 45-55 years were included in this study. All subjects had varying degrees of fat accumulation and skin laxity in the face, neck, abdomen and legs. The subjects took HA35 5 grams orally every day for 21 consecutive days. The fat thickness of the face, neck, abdomen and legs was assessed by a fat measuring instrument, and the skin laxity was assessed using a numerical rating system (NRS) with a score range of 0-10, with 0 indicating the tightest skin and 10 indicating the loosest skin. The evaluation time points were before treatment, 14 days after taking the medicine, and 21 days after taking it.

[0216] Results: After taking HA35 orally for 14 days, the subcutaneous fat thickness of the subjects' face, neck, abdomen and legs decreased, and the skin laxity was significantly improved. The results in Table 14 show that after 14 days, the subjects' facial fat decreased by an average of about 30%, and the abdominal and leg fat decreased by about 25%. After 21 days, facial fat further decreased to 45%, neck skin laxity improved by 35%, and abdominal and leg fat decreased by 40%. The subjects generally reported that the facial contour became tighter, the sagging of the neck skin was significantly improved, the abdominal fat thickness was significantly reduced, and the leg lines were slimmer.

[0217] Table 14. Improvement rate of facial skin problems

[0218] Evaluation Metrics Before treatment 14 days after treatment 21 days after treatment Facial fat reduction rate 0％ 20％ 35％ Improvement rate of neck skin sagging 0％ 20％ 35％ Abdominal fat reduction rate 0％ 15％ 20％ Leg fat reduction rate 0％ 15％ 20％

[0219] Discussion and Conclusion

[0220] The results of the study showed that oral HA35 can significantly reduce subcutaneous fat accumulation in female subjects aged 45-55, especially in the face, neck, abdomen and legs. After taking it for 14 days, the subjects' facial fat was significantly reduced and the facial contour was firmer. In addition, the sagging condition of the neck skin was also significantly improved, with the skin becoming firmer, and the fat thickness of the abdomen and legs was also significantly reduced.

[0221] HA35 achieves this effect through its deep repair and anti-inflammatory properties on the skin structure, possibly by promoting fat metabolism and regulation of the extracellular matrix. Compared with invasive cosmetic methods, HA35 provides a gentler and side-effect-free way to help women restore firm skin and shape their ideal body shape. This study provides new evidence for the application of HA35 in the field of fat dissolving and skin tightening.

[0222] Therefore, oral administration of HA35 (5 g / time, for 21 consecutive times) can be used as a safe and effective cosmetic method to help reduce subcutaneous fat, especially showing significant effects in improving the face, neck, abdomen and legs, and has wide potential for cosmetic and body shaping applications.

[0223] Embodiment 17

[0224] Objective: This study aimed to evaluate the therapeutic effect of oral administration of a low-molecular-weight hyaluronic acid fragment, HA35 (35 kDa, >90% distribution range 10-70 kDa), on sports injury pain.

[0225] Background: Sports injuries are a common health problem among sports enthusiasts and athletes. Injuries are usually accompanied by pain, swelling and inflammation, which seriously affect sports performance and daily life. Traditional treatments include rest, ice packs, physical therapy and medication, but these methods often have certain side effects or require a long recovery period. Hyaluronic acid (HA), a naturally occurring component in human tissue, has the functions of lubricating joints, relieving inflammation and promoting tissue repair. Low-molecular-weight hyaluronic acid fragment HA35 has shown its potential in relieving pain and promoting soft tissue repair in clinical studies. Therefore, this study aims to explore its therapeutic effect on sports injury pain.

[0226] Methods: This study included 20 subjects aged 25-45 years with pain caused by sports injuries. The subjects took HA35 5 grams orally every day for 21 consecutive days. The pain score was scored using a numerical rating system (NRS) ranging from 0 to 10 points, with 0 indicating no pain and 10 indicating the most severe pain. The evaluation time points were before treatment, 3 days, 7 days, 14 days and 21 days after treatment, and the evaluation indicators included the degree of pain relief and functional recovery of the injured part.

[0227] Results: The results in Table 15 show that 3 days after oral administration of HA35, the subjects generally felt that the pain was relieved. The average pain score dropped from 6.5±1.0 before treatment to 5.0±1.0 after 3 days, and further dropped to 3.0±0.8 after 7 days. The pain score was 2.0±0.5 after 14 days, and dropped to 1.5±0.4 on day 21. The functional recovery of the subjects at the injured part was also improved, and the range of motion and activity were gradually restored.

[0228] Table 15. Pain scores of patients with sports injuries before and after treatment

[0229]

[0230] Discussion and Conclusion:

[0231] The results of this study showed that oral HA35 has a significant effect on relieving pain caused by sports injuries. On the third day after treatment, the subjects' pain was significantly relieved, and the pain relief effect was more significant on the seventh day, and the pain was further relieved on the 14th and 21st days. This shows that HA35 can take effect quickly and continuously relieve pain associated with sports injuries. In addition to the reduction of pain, the subjects' motor ability and functional recovery of the injured area were also significantly improved, indicating that HA35 may play its role by regulating inflammatory response and promoting soft tissue repair.

[0232] Compared with traditional analgesics and physical therapy, oral HA35 has no obvious side effects, can effectively relieve pain and promote tissue repair, and is suitable as an adjuvant therapy for sports injury pain management. The mechanism of action of HA35 may be related to its inhibition of inflammation and promotion of tissue repair.

[0233] In summary, oral administration of HA35 (5 g / time, for 21 consecutive days) has a significant therapeutic effect on pain caused by sports injuries. Significant pain relief can be felt from the third day after treatment, and it has good potential for clinical application.

[0234] Embodiment 18

[0235] Objective: This study aimed to evaluate the therapeutic effect of oral administration of low-molecular-weight hyaluronic acid fragment HA35 (35 kDa, >90% distribution range 10-70 kDa) on diabetic foot wound pain.

[0236] Background: Diabetic foot is a common and serious complication in patients with diabetes, often accompanied by chronic wounds, pain, and difficulty in healing. Due to poor blood circulation and nerve damage, wounds in patients with diabetic foot heal slowly and pain increases, which may even lead to infection, gangrene, or amputation in severe cases. Traditional treatments include wound care, antibiotics, and analgesics, but these methods cannot quickly promote wound healing or effectively relieve long-term pain. Hyaluronic acid (HA) plays an important role in tissue repair, anti-inflammation, and moisturizing. The low molecular weight hyaluronic acid HA35 fragment is believed to accelerate wound repair and relieve pain. Therefore, this study aims to explore the therapeutic effect of oral HA35 on diabetic foot wound pain.

[0237] Methods: Twenty subjects aged 50-70 years with diabetic foot wound pain were included in this study. The subjects took HA35 5 g orally daily for 21 consecutive days. The degree of wound pain was assessed using a numerical rating system (NRS) ranging from 0 to 10 points, with 0 indicating no pain and 10 indicating the most severe pain. The assessment time points were before treatment and 7, 14, and 21 days after treatment. In addition, the healing of the wound was evaluated by clinical examination, including the reduction of wound area and inflammation.

[0238] Results: The results in Table 16 show that after 7 days of oral HA35 treatment, the patient's pain began to significantly alleviate, and the pain score dropped from 7.0±1.2 before treatment to 5.5±1.0. After 14 days, the pain score further dropped to 3.5±0.8, and after 21 days, the pain score dropped to 2.5±0.5. In addition, the healing of the patient's wound also improved significantly, with the wound area reduced by an average of 30% to 40%, and inflammatory symptoms such as redness, swelling and exudation were significantly reduced.

[0239] Table 16. Pain scores and wound area reduction rates in patients with diabetic foot before and after treatment

[0240] Evaluation Metrics Before treatment 7 days after treatment 14 days after treatment 21 days after treatment Pain score 7.0±1.2 2.5±1.0 1.0±0.8 0.0±0.0 Wound area reduction rate 0％ 5％ 10％ 15％

[0241] Discussion and Conclusion:

[0242] The results of this study showed that oral HA35 has a significant effect on relieving pain in diabetic foot wounds. After taking HA35 orally for 7 days, the pain of the subjects began to ease, and the pain was further relieved on the 14th and 21st days. This shows that HA35 has a faster onset time in relieving chronic pain in patients with diabetic foot. In addition, wound healing also showed a good improvement trend. Oral HA35 can promote wound shrinkage and reduce local inflammation, which may be achieved through its anti-inflammatory, cell regeneration and accelerated tissue repair mechanisms.

[0243] The pain of diabetic foot is usually difficult to control, and HA35, as a natural substance with no side effects, shows advantages over traditional analgesics, not only relieving pain but also accelerating wound healing. Compared with other treatments, the oral route of HA35 is simple and convenient, and has good patient compliance.

[0244] In summary, oral administration of HA35 (5 g / time, for 21 consecutive days) has a significant therapeutic effect on diabetic foot wound pain. Patients can feel the pain relief after 7 days of treatment, and the wound healing and pain relief effects become more obvious as the treatment progresses. HA35 provides a safe and effective treatment option for patients with diabetic foot.

[0245] The above Examples 11-18 also show that a high dose of a low-molecular-weight hyaluronic acid oral preparation in the form of a gel has a good therapeutic effect. Similar to the above Examples 4-10, a low dose of a low-molecular-weight hyaluronic acid oral preparation cannot produce a corresponding effect.

[0246] Embodiment 19

[0247] Objective: To orally treat back pain or myofasciitis with a low molecular weight hyaluronic acid fragment HA70 of 70 kDa (>90% of the distribution range 40-100 kDa).

[0248] Background: Back pain is mainly caused by myofasciitis. The main clinical feature of myofasciitis is tender trigger points. Studies have shown that local injection of 0.2g HA70 is effective in treating back pain or myofasciitis. This example studies the effect of oral administration of different doses of HA70 in treating back pain or myofasciitis.

[0249] Method: This example studied 20 patients with back pain, divided into two dosage groups. The first group had 5 males and 5 females, aged between 42 and 60 years old, and took HA70 orally at a dosage of 5 grams / time / day for 21 days; the second group had 5 males and 5 females, aged between 40 and 65 years old, and took HA70 at a dosage of 0.2 grams / time / day. This example also evaluated the degree of facial fat, facial skin and mental state of the patients. The numerical pain score (NPRS) was used to evaluate the pain and facial state of the patients, with a pain rating of 0-10 points, with 0 indicating no pain, no fat, the whitest skin and the most energetic, and 10 indicating the greatest degree of pain, the most fat, the darkest skin and the most listless.

[0250] Results: The results in Table 1 show that HA70 can effectively treat back pain by 40%-50% 2 hours after oral administration; 90%-95% after 2 days of oral treatment; and completely relieve the patient's back pain after 21 days of oral treatment. In contrast, oral administration of 0.2g of HA70 has no effect on pain relief. The results in Table 2 show that after 21 days of treatment, the fat on the face, the whiteness of the skin, and the spirit of vitality have all been significantly improved. After treatment, the patient's face became thinner by more than 48%, the skin became whiter by more than 51%, and the spirit of vitality increased by 51%.

[0251] Table 17. Pain scores before and after treatment of back pain

[0252]

[0253] Table 18. Facial fat level and facial skin condition

[0254] Evaluation Metrics Before treatment After 21 days of treatment Facial fat level 5.6±2.0 2.4±1.0 Facial skin whiteness 5.8±2.0 2.1±1.3 Facial mental state 5.4±2.2 2.5±1.4

[0255] Note: The dosage of HA70 in Table 18 is 5 g / 1 time / day.

[0256] Conclusion: The results of this study suggest that oral administration of HA70, like HA35, can relieve back pain caused by myofasciitis.

[0257] Embodiment 20

[0258] Objective: This study aimed to evaluate the therapeutic effect of oral administration of a low molecular weight hyaluronic acid fragment HA70 with a size of 70 kDa (>90% of the distribution range 40-100 kDa) on postherpetic neuralgia.

[0259] Background: Postherpetic neuralgia (PHN) is the most common chronic complication after herpes zoster infection, especially in elderly patients. PHN is characterized by persistent or intermittent severe pain, which seriously affects the quality of life of patients. Currently, there are limited treatments for PHN. Conventional analgesics such as antidepressants, antiepileptic drugs and opioids can relieve pain in some patients, but they are accompanied by side effects and have limited effects.

[0260] Methods: A total of 10 patients with postherpetic neuralgia were included in this study and divided into two groups according to the dosage. The first group was the high-dose group, with 5 patients aged 42-64 years old, who took HA70 5 g / time / day orally for 21 consecutive days; the second group was the low-dose group, with 5 patients aged 44-68 years old, who took HA70 0.2 g / time / day orally for 21 consecutive days. The pain level of the patients was assessed using the Numerical Pain Scale (NPRS), with a score range of 0-10, with 0 indicating no pain and 10 indicating severe pain. The assessment time points were before treatment, 1 day after treatment, 2 days after treatment, and 21 days after treatment.

[0261] result

[0262] Patients in the high-dose HA70 group (5 g / day) experienced more than 35% pain relief within one day of oral administration; more than 65% pain relief after two days; and the pain almost completely disappeared after 21 days of treatment. In contrast, the pain scores of patients in the low-dose HA70 group (0.2 g / day) did not change much at each time point, and no significant relief effect was observed. See Table 19 for details:

[0263] Table 19. Pain scores of patients with postherpetic neuralgia before and after treatment

[0264] HA70 dosage Before treatment 1 day after treatment 2 days after treatment 21 days after treatment 5 g / 1 time / day 6.5±1.2 4.0±1.2 3.0±0.6 0.5±0.3 0.2 g / 1 time / day 6.6±1.6 6.6±1.2 6.6±1.3 6.4±1.2

[0265] in conclusion:

[0266] The results of the study showed that oral administration of HA70 could significantly relieve postherpetic neuralgia (PHN).

[0267] Embodiment 21

[0268] Objective: This study aimed to evaluate the efficacy of oral administration of a low-molecular-weight hyaluronic acid fragment, HA70 (70 kDa, >90% distribution range 40-100 kDa), on the relief of bone pain in patients with advanced anorectal cancer, especially persistent pain caused by bone metastasis.

[0269] Background: Patients with advanced anorectal cancer often have bone metastases, resulting in persistent and unbearable lower limb bone pain.

[0270] Methods: This study included 10 patients with advanced anorectal cancer, all of whom had clear bone metastases and an age range of 60 to 65 years. The patients were divided into two groups according to the dose. The first group was the high-dose group, with 5 patients taking HA70 10 g / time / day orally for 21 consecutive days; the second group was the low-dose group, with 5 patients taking HA70 0.2 g / time / day orally for 21 consecutive days. The pain level was assessed by the Numerical Pain Scale (NPRS), with a score range of 0-10 points, 0 for no pain and 10 for severe pain. The evaluation time points included before treatment, 1 day after treatment, 2 days after treatment, and 21 days after treatment.

[0271] Results: The pain of patients in the high-dose HA70 group (10 g / day) was relieved by more than 30% within 1 day after oral administration, and the pain was further relieved by more than 50% after 2 days; after 21 days, the pain stabilized at a relief level of about 30%. Although the pain did not completely disappear, the patients reported that the pain was significantly relieved and the impact on daily life was reduced. The pain scores of patients in the low-dose group (0.2 g / day) did not change much, and no significant relief was observed. See Table 20 for details:

[0272] Table 20. Pain scores before and after treatment in patients with advanced anorectal cancer

[0273] HA70 dosage Before treatment 4 hours after treatment 2 days after treatment 21 days after treatment 10 g / 1 time / day 7.5±1.2 3.0±0.4 2.0±0.5 1.0±0.4 0.2 g / 1 time / day 8.0±1.2 7.8±1.0 7.8±1.0 7.8±1.0

[0274] in conclusion:

[0275] The results of the study showed that oral administration of HA70 could significantly relieve bone pain in advanced anorectal cancer, especially in the high-dose group (10 g / day), where the pain was reduced in a short period of time and maintained at a controllable level after 21 days of treatment.

[0276] Embodiment 22

[0277] Objective: To evaluate the efficacy of oral HA70 (10 g / 2 times / day) on pain relief in patients before and after dental implant surgery.

[0278] Background: After dental implant surgery, patients often face significant postoperative pain problems, especially elderly patients, who may experience stronger pain reactions due to poor periodontal health and reduced body recovery ability.

[0279] method:

[0280] 1. Study subjects: Two 65-year-old male patients who were scheduled for dental implant surgery. Both patients had no other serious underlying diseases and were similar in periodontal health and general health status.

[0281] 2. Experimental design: Each patient undergoing dental implant surgery was divided into two groups. One group was the oral HA70 group, which took HA70 orally at a dose of 10g twice a day, starting from 1 day before surgery and continuing until 3 days after surgery. The other group took acetaminophen according to the dosage after surgery. Both groups took antibiotics to prevent inflammatory infection.

[0282] 3. Pain assessment method: The pain perception of patients on the 1st, 2nd and 3rd day after the two surgeries was recorded using the National Pain Rating Scale (NPRS). The NPRS score ranged from 0 (no pain) to 10 (severe pain).

[0283] 4. Statistical analysis: Paired t-test was used to compare the pain scores of patients taking HA70 at different time points, and the significance level was set at p < 0.05.

[0284] Results: Pain score (VAS): The pain scores of patients taking hyaluronic acid on the first, second and third days after surgery were significantly lower than those of the control group. On the third day, the pain scores of both groups were 0, with no significant difference. NSAIDs use: The dose of NSA IDs used within 24 hours after surgery in patients taking hyaluronic acid was significantly lower than that in the control group. The average dose of acetaminophen used within 24 hours after surgery in the HA35 group was 0.3 g, while that in the control group was 0.9 g; from the first to the third day after surgery, the average number of acetaminophen used in the HA70 group was 3 times, while that in the control group was 15 times (p<0.01); from the third day onwards, patients in the HA70 group stopped using acetaminophen.

[0285] Table 21. Pain scores of patients undergoing dental implant surgery before and after treatment

[0286]

[0287] in conclusion:

[0288] This study showed that oral administration of HA70 (10 g / 2 times / day) can effectively reduce the pain experienced by patients after dental implant surgery and significantly reduce the need for NSAIDs after surgery.

[0289] Embodiment 23

[0290] Objective: This study aimed to evaluate the effect of oral administration of a low-molecular-weight hyaluronic acid fragment, HA70 (70 kDa, >90% distribution range 40-100 kDa), on dissolving subcutaneous fat, slimming the face, improving neck skin laxity, and reducing abdominal and leg fat in women aged 42-54 years within 14 to 21 days.

[0291] Methods: This study included 20 female subjects aged 42-54 years, all of whom had varying degrees of fat accumulation and skin laxity in the face, neck, abdomen, and legs. The subjects took HA70 10 grams orally every day for 21 consecutive days. The fat thickness of the face, neck, abdomen, and legs was assessed by a fat measuring instrument, and the skin laxity was assessed using a numerical rating system (NRS) with a score range of 0-10, with 0 indicating the tightest skin and 10 indicating the loosest skin. The evaluation time points were before treatment, 14 days after taking the medicine, and 21 days after taking it.

[0292] Results: After taking HA70 orally for 14 days, the subcutaneous fat thickness of the subjects' face, neck, abdomen and legs decreased, and the skin laxity was significantly improved. The results in Table 22 show that after 14 days, the subjects' facial fat decreased by an average of about 30%, and the abdominal and leg fat decreased by about 30%. After 21 days, facial fat was further reduced to 50%, neck skin laxity improved by 40%, and abdominal and leg fat decreased by 40%. The subjects generally reported that the facial contour became tighter, the sagging of the neck skin was significantly improved, the abdominal fat thickness was significantly reduced, and the leg lines were slimmer.

[0293] Table 22. Improvement rate of facial skin problems

[0294] Evaluation Metrics Before treatment 14 days after treatment 21 days after treatment Facial fat reduction rate 0％ 22％ 40％ Improvement rate of neck skin sagging 0％ 24％ 40％ Abdominal fat reduction rate 0％ 20％ 25％ Leg fat reduction rate 0％ 20％ 25％

[0295] Discussion and Conclusion

[0296] The results of the study showed that oral HA70 can significantly reduce subcutaneous fat accumulation in female subjects, especially in the face, neck, abdomen and legs. After taking it for 14 days, the subjects' facial fat was significantly reduced and the facial contour was firmer. In addition, the sagging condition of the neck skin was also significantly improved, with the skin becoming firmer, and the fat thickness of the abdomen and legs was also significantly reduced.

[0297] Embodiment 24

[0298] Objective: This study aimed to evaluate the effects of oral administration of a low-molecular-weight hyaluronic acid fragment, HA70 (70 kDa, >90% distribution range 40-100 kDa), on the resolution of skin problems.

[0299] Methods: This study included 20 female subjects aged between 40 and 50 years old, all of whom had varying degrees of skin inflammation, such as erythema, yellow spots, and tiny inflammatory wounds. All subjects took 10 grams of HA70 orally every day for 21 consecutive days. Evaluation indicators included skin moisture content, elasticity, glossiness, and improvement of inflammatory symptoms. Moisture and elasticity were measured using a professional skin measurement instrument, and the number and severity of inflammatory red and yellow spots and wounds were evaluated using a numerical rating system (NRS), with a score range of 0-10 points, 0 indicating no inflammation or wounds, and 10 indicating the most severe inflammation. The evaluation time points were before treatment, 7 days after taking the medicine, and 21 days after taking it.

[0300] Results: After taking HA70 orally for 7 days, the subjects' skin moisture, elasticity and glossiness were improved, and inflammatory symptoms such as red and yellow spots and tiny inflammatory wounds were significantly reduced.

[0301] Table 23. Ratings of patients with skin problems before and after treatment

[0302]

[0303]

[0304] in conclusion:

[0305] The results of this study indicate that oral administration of HA70 has significant cosmetic effects on female skin, especially in reducing skin inflammation.

[0306] The above Examples 22-24 also show that a high dose of a low-molecular-weight hyaluronic acid oral preparation in the form of a gel has a good therapeutic effect. Similar to the aforementioned Examples 19-21, a low dose of a low-molecular-weight hyaluronic acid oral preparation cannot produce a corresponding effect.

[0307] It should also be noted that the oral dosage of the oral preparation of low molecular weight hyaluronic acid fragments in the present application is 5-10 g / day, 1-2 times a day. In addition to the preferred methods listed in the above embodiments, the oral dosage can also be 6 g / day, 1-2 times a day; 7 g / day, 1-2 times a day, 8 g / day, 1-2 times a day; 9 g / day, 1-2 times a day, which can also achieve similar effects, and will not be repeated here.

[0308] The above lists the application of oral preparations of 35kDa (>90% distribution range 10-70kDa). At the same time, the present invention also conducted corresponding research on other low-molecular hyaluronic acid fragments, proving that the use of low-molecular hyaluronic acid fragments with an average molecular weight of 70kDaHA (>90% distribution range 40-100kDa) can also achieve basically similar effects. However, due to the limitation of the number of embodiments, only some of the more preferred embodiments are given above. Since the action mechanisms of the two are similar, they have similar effects and will not be repeated here.

[0309] The above description is only a preferred embodiment of the present invention and does not limit the present invention in any form. Those skilled in the art may make some simple modifications, equivalent changes or modifications using the technical contents disclosed above, which all fall within the protection scope of the present invention.

Claims

1. An oral preparation of low molecular weight hyaluronic acid fragments, characterized in that: The low-molecular-weight hyaluronic acid fragments are low-molecular-weight hyaluronic acid fragments with an average molecular weight of 35 kDa and a distribution range of >90% between 10-70 kDa; or the low-molecular-weight hyaluronic acid fragments are low-molecular-weight hyaluronic acid fragments with an average molecular weight of 70 kDa and a distribution range of >90% between 40-100 kDa; the low-molecular-weight hyaluronic acid fragments are directly used as an oral preparation, which is a high-dose gel oral preparation, and its oral dosage is 5-10 g / day, 1-2 times a day.

2. The oral preparation of low-molecular hyaluronic acid fragments according to claim 1, characterized in that After oral administration, the oral preparation can be rapidly absorbed into the blood through the intestinal lymph nodes and spleen.

3. The oral low-molecular hyaluronic acid fragment preparation according to any one of claims 1 or 2, characterized in that: The oral preparation is a fast-acting oral preparation.

4. An application of the oral preparation of low molecular weight hyaluronic acid fragments according to any one of claims 1 to 3, characterized in that: Application as a medicine for treating pain or in the preparation of a medicine for treating pain.

5. The use of the oral preparation of low molecular weight hyaluronic acid fragments according to claim 4, characterized in that: The pain described is: inflammatory pain in the shoulder / neck / back, neuropathic pain from herpes zoster / diabetic foot, pain from bone metastasis of advanced cancer, pain from radiotherapy for advanced cancer, pain after dental implant surgery, pain after vaginal egg retrieval, muscle strain, muscle stiffness and discomfort, sports injury pain, or pain from diabetic foot wounds.

6. The use of the oral preparation of low molecular weight hyaluronic acid fragments according to claim 5, characterized in that: The shoulder / neck / back inflammatory pain is shoulder / neck / back inflammatory pain caused by fasciitis.

7. Use of the oral preparation of low molecular weight hyaluronic acid fragments according to any one of claims 1 to 3, characterized in that: Application as a subcutaneous lipolysis or skin beauty drug or in the preparation of a subcutaneous lipolysis or skin beauty drug.

8. The use of the oral preparation of low-molecular hyaluronic acid fragments according to claim 7, characterized in that: The skin beauty treatment is to moisturize the skin, reduce skin inflammation, improve skin elasticity, improve skin gloss or tighten the skin; The subcutaneous lipolysis is facial fat reduction, neck subcutaneous fat reduction, abdominal subcutaneous fat reduction or leg subcutaneous fat reduction.

9. Use of the oral preparation of low molecular weight hyaluronic acid fragments according to any one of claims 1 to 3, characterized in that: Application as a drug for lowering blood pressure or in the preparation of a drug for lowering blood pressure.

10. Use of the oral preparation of low molecular weight hyaluronic acid fragments according to any one of claims 1 to 3, characterized in that: The invention is used in a gastrointestinal tract protection drug for relieving the spicy and painful feeling in the mouth, the discomfort after drinking or the symptoms of drunkenness, or improving abdominal distension and digestive discomfort after drinking, or in the preparation of a gastrointestinal tract protection drug for relieving the spicy and painful feeling in the mouth, the discomfort after drinking or the symptoms of drunkenness, or improving abdominal distension and digestive discomfort after drinking.

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