Plant fermentation product for reducing infection after surgical operation and preparation method thereof

Abstract

According to the plant fermentation product and the preparation method thereof, plant raw materials including aronia melanocarpa, acai, inula nervosa stems and leaves, horseradish tree leaves, andrographis paniculata leaves and ginkgo leaves are fermented through probiotics, and andrographolide, quinic acid and shikimic acid are obtained. The fermentation product shows anti-infection and death rate reducing effects on a rat sepsis model, provides a new thought for improving prognosis of surgical patients and reducing postoperative infection probability, and has a good market application prospect.

Description

Technical Field

[0001] The invention relates to a plant fermentation product, and more specifically, to a plant fermentation product capable of reducing postoperative infection and improving the prognosis of surgical operations. Background Art

[0002] Pseudomonas aeruginosa (PA) is a common Gram-negative bacterium, also known as Pseudomonas aeruginosa, which is widely found in soil, water and air and has strong environmental adaptability. Pseudomonas aeruginosa is a conditional pathogen. When skin tissue is damaged or the body's immune function is low, the invasion of this pathogen can cause fatal infection of tissues and organs. It is reported that hospital-acquired infections caused by this bacterium account for about 17.27% in China, ranking second, and 10-31.5% abroad, making it one of the important culprits of postoperative infection.

[0003] Surgery will inevitably cause damage to the skin and tissues at the surgical site. When the microbial contamination of the surgical incision reaches a certain level, the patient is prone to postoperative infection. Postoperative surgical infection includes incision infection and infection of the organs or cavities involved in the operation. The risk factors for surgical site infection include patient and surgical factors. The main factors on the patient side are: age, nutritional status, immune function, health status, etc. The main surgical factors are: preoperative hospitalization time, skin preparation method and time, surgical site skin disinfection, operating room environment, sterilization of surgical instruments, aseptic operation during the operation, surgical technique, duration of the operation, and use of preventive antibiotics.

[0004] Patent CN111671726A discloses a composition for preventing postoperative infection of hemorrhoids, which is prepared by mixing antibacterial agents, healing agents, moisturizers, film formers, astringent carbamide peroxide, cooling agent WS-23, buffer sodium gluconate and preservative octenidine, and is sprayed on the wound surface to form a protective film to prevent secondary infection. The composition can only be used to reduce surgical incision infection, so it is suitable for postoperative hemorrhoid surgery, but not for more complex cardiac, neurological and brain surgeries involving more organs and cavities.

[0005] According to the China Health Statistics Yearbook, in 2019, more than 69 million inpatient surgeries were performed nationwide. With the aging of society and the continuous extension of life expectancy, it is foreseeable that the number of surgical operations will continue to rise for a long time, which puts higher demands on surgical postoperative care. There is an urgent need for a product that can be widely used in the prevention of postoperative infections in different surgical operations, reduce the probability of postoperative infection in patients, enable patients to recover smoothly after surgery, return to their families and society, and enable surgical operations to truly achieve the purpose of curing diseases and saving lives. Summary of the invention

[0006] The invention relates to a plant fermentation product capable of reducing postoperative infection and improving the prognosis of surgical operations and a preparation method thereof.

[0007] Technical solution A: S1 preferably 5 kg of black fruit glandular rib aronia, 5 kg of acai, and 3 kg of stems and leaves of Inula villosa, which are quickly washed at low temperature, and the black fruit glandular rib aronia and acai are removed from the stems and mixed with the stems and leaves of Inula villosa, and 65 mg / L SO2 and 25 mg / L pectinase are added during the crushing, and the mixture is allowed to stand at 4°C for 20 hours after being fully mixed, and then 2 kg of saturated sterilized sucrose solution is added, and 120 cfu / mL of brewer's yeast is inoculated and fermented at 26±2°C for 5-7 days, and the skin residue is separated to obtain the first stage fermentation liquid. S2 takes 2 kg of Moringa leaves, 4 kg of Andrographis paniculata leaves, and 4 kg of Ginkgo biloba leaves, which are washed at low temperature, centrifuged and dried, chopped, and sterilized, and then poured into the fermentation liquid of step S1, pectinase is added, and one or more of activated lactic acid bacteria, Lactobacillus plantarum, Lactobacillus rhamnosus, and Leuconostoc mesentericus are inoculated for fermentation, and fermented at 26-44°C for 38-50 hours to obtain plant fermentation products.

[0008] Technical solution B: S1 preferably 5 kg of black fruit glandular rib aronia, 5 kg of acai, and 3 kg of stems and leaves of Inula villosa, which are quickly washed at low temperature, and the black fruit glandular rib aronia and acai are removed from the stems and mixed with the stems and leaves of Inula villosa, and 65 mg / L SO2 and 25 mg / L pectinase are added during the crushing, and the mixture is allowed to stand at 4°C for 20 hours after being fully mixed, and then 2 kg of saturated sterilized sucrose solution is added, and 120 cfu / mL of brewer's yeast is inoculated and fermented at 26±2°C for 5-7 days, and the skin residue is separated to obtain the first stage fermentation liquid. S2 takes 2 kg of Moringa leaves, 4 kg of Andrographis paniculata leaves, and 4 kg of Ginkgo biloba leaves, which are washed at low temperature, centrifuged and dried, chopped and sterilized, and poured into the fermentation liquid of step S1, pectinase is added, and one or more of activated lactic acid bacteria, Lactobacillus plantarum, Lactobacillus rhamnosus, and Leuconostoc mesentericus are inoculated for fermentation, and fermented at 26-44°C for 38-50 hours to obtain plant fermentation products. DETAILED DESCRIPTION

[0009] The object of the present invention is to provide a plant fermentation product for reducing postoperative infection in patients.

[0010] In order to achieve the above object, the present invention adopts the following technical solutions:

[0011] In a technical solution A, two fermentation stages are included:

[0012] S1 fermentation stage: preferably 5 kg of Aronia nigra, 5 kg of Acai, and 3 kg of Inula villosa stems and leaves, quickly wash at low temperature, remove the stems of Aronia nigra and Acai, mix and crush them with the stems and leaves of Inula villosa, and add 65 mg / L SO2 and 25 mg / L pectinase during crushing. After fully mixing, let stand at 4°C for 20 hours, then add 2 kg of saturated sterilized sucrose solution, inoculate 120 cfu / mL of brewer's yeast, ferment at 26±2°C for 5-7 days, separate the skin and residue, and obtain the first stage fermentation liquid.

[0013] S2 fermentation stage: take 2kg of Moringa leaves, 4KG of Andrographis paniculata leaves and 4KG of Ginkgo biloba leaves, wash them at low temperature, centrifuge and dry them, chop them and sterilize them, then pour them into the fermentation liquid of step S1, add pectinase and inoculate activated lactic acid bacteria, Lactobacillus plantarum, Lactobacillus rhamnosus and Leuconostoc mesenteroides for fermentation, ferment at 26-44°C for 38-50 hours, filter and sterilize to obtain plant fermentation products.

[0014] In another technical solution B, three fermentation stages are included:

[0015] S1 fermentation stage: preferably 5 kg of Aronia nigra, 5 kg of Acai, and 3 kg of Inula villosa stems and leaves, quickly wash at low temperature, remove the stems of Aronia nigra and Acai, mix and crush them with the stems and leaves of Inula villosa, and add 65 mg / L SO2 and 25 mg / L pectinase during crushing. After fully mixing, let stand at 4°C for 20 hours, then add 2 kg of saturated sterilized sucrose solution, inoculate 120 cfu / mL of brewer's yeast, ferment at 26±2°C for 5-7 days, separate the skin and residue, and obtain the first stage fermentation liquid.

[0016] S2 fermentation stage: Take 1kg of Moringa leaves, 2KG of Andrographis paniculata leaves and 2KG of Ginkgo biloba leaves, wash them at low temperature, centrifuge and dry them, chop them and sterilize them, then pour them into the fermentation liquid of step S1, add pectinase and inoculate activated lactic acid bacteria, Lactobacillus plantarum, Lactobacillus rhamnosus and Leuconostoc mesenteroides for fermentation, and ferment at 26-44℃ for 38-40 hours.

[0017] S3 fermentation stage: 1 kg of Moringa leaves, 2 kg of Andrographis paniculata leaves and 2 kg of Ginkgo biloba leaves are washed at low temperature, centrifuged and dried, chopped and sterilized, and then poured into the fermentation liquid of step S2, and fermented at 26-44°C for 8-12 hours, filtered and sterilized to obtain plant fermentation products.

[0018] The plant fermentation product a obtained by technical solution A and the plant fermentation product b obtained by technical solution B were taken, and the contents of andrographolide, quinic acid and shikimic acid therein were detected as shown in the following table:

[0019]

[0020] The plant fermentation products obtained by Technical Solution A and the plant fermentation products obtained by Technical Solution B were taken and their anti-infection effects were verified through in vitro antibacterial experiments and animal experiments.

[0021] In vitro antibacterial experiment:

[0022] Materials and methods

[0023] Plant fermentation product ethanol solution: After plant fermentation product a is freeze-dried and dehydrated, weigh 6 g of dry powder and add 10 ml of 70% ethanol to fully dissolve it. Filter the supernatant to obtain the plant fermentation product a ethanol solution for use; similarly, obtain the plant fermentation product b ethanol solution.

[0024] Positive control: 200 g / L sodium benzoate was prepared with 70% ethanol solution as a positive control.

[0025] Negative control: 70% ethanol solution was used as negative control.

[0026] After activation, Pseudomonas aeruginosa was transferred to LB agar medium, placed in a 37°C biochemical incubator for 24 hours, and subcultured twice. The total number of colonies was determined by plate dilution method, and then diluted to a bacterial suspension (5×108CFU / mL) with 0.9% sterile NaCl solution. Take 0.1mL of each test bacterial suspension on the plate and spread it evenly. Place 4 sterile Oxford cups on each plate. Pipette 100μL of plant fermentation product a ethanol solution, plant fermentation product b ethanol solution, 70% ethanol solution and sodium benzoate ethanol solution into sterile Oxford cups respectively. Place the culture dish in a biochemical incubator and culture at 37°C for 24 hours. Observe the size of the inhibition zone and measure its diameter with an automatic colony counter.

[0027] The diameters of the inhibition zones are shown in the following table:

[0028]

[0029] result:

[0030] Plant fermentation products a and plant fermentation products b showed inhibitory effects on Pseudomonas aeruginosa in vitro.

[0031] Animal Experimentation:

[0032] Materials and methods

[0033] SPF SD rats, male, 150-170g. Rearing conditions: room temperature 21-25℃, relative humidity 40-60%, 12h / 12h light-dark cycle. 4 rats per cage, free access to food and water, bedding changed every other day.

[0034] After 1 week of adaptive feeding, the rats were randomly divided into a sham operation group (n=10) and a CLP group (n=80), and the intraperitoneal infection model was established as follows:

[0035] Rats were fasted for 12 hours before the experiment, but were not restricted to water. Rats were anesthetized with 2% sodium pentobarbital at 50 mg / kg intraperitoneally before the operation. After anesthesia, the rats had no righting reflex. The abdominal area was routinely disinfected with iodine tincture, and a sterile drape was laid. An incision of about 1 cm was made along the midline of the abdominal wall, and the cecum was explored and pulled out with sterile forceps to expose it.

[0036] Sham operation group (n=10): Only the cecum was pulled out of the abdominal cavity and then returned to the abdominal cavity.

[0037] CLP group (n=80): The cecum was ligated with sterile No. 4 silk thread 2 cm away from the blind end of the cecum, and the blind end of the cecum was pierced twice with a 20G trocar to avoid damaging the mesentery and cecal blood vessels. A small amount of feces was squeezed out and returned to the abdominal cavity, and the abdominal wall incision was sutured layer by layer.

[0038] After surgery, rats in each group were subcutaneously injected with 5 ml of preheated saline to replenish body fluids lost during surgery, and were placed on a heating pad until they woke up from anesthesia and were sent back to the breeding room to eat and drink freely.

[0039] The CLP group was divided into four groups: model group, CLP group-a group, CLP group-b group, and CLP-antibiotic group, with 20 mice in each group. The CLP group-a group, CLP group-b group, and CLP-antibiotic group were gavaged with plant fermentation product a, plant fermentation product b, and antibiotics (cefprozil dispersible tablets + metronidazole tablets) according to the following table:

[0040]

[0041] The model group (n=20) and the sham operation group (n=10) were given an equal amount of saline by intragastric administration once a day for 3 consecutive days. The mortality of the rats was recorded.

[0042] After the intragastric administration on the third day, the rats were fasted but allowed to drink water for 12 hours that night. The rats were anesthetized and opened the abdomen under sterile conditions, and blood samples were collected for bacterial culture.

[0043] result:

[0044] 1 Mortality

[0045] The mortality rate of rats in the model group was 60%, that in the CLP-a group was 15%, and that in the CLP-b group and the CLP-antibiotic group was 10%.

[0046]

[0047] Note: Compared with the sham operation group, a P<0.05; compared with the model group, bP<0.05

[0048] 2. Blood bacterial culture

[0049] The model group had the highest positive rate of bacterial detection, and the bacteria detected in each group are as follows:

[0050]

[0051]

[0052] Note: Compared with the sham operation group, a P<0.05; compared with the model group, b P<0.05.

[0053] Compared with the model group, plant fermentation products a and plant fermentation products b can significantly reduce the mortality rate and blood bacteria positive detection rate of CLP model rats. The data are statistically significant (P<0.05), indicating that plant fermentation products a and plant fermentation products b exhibit anti-infection effects in the rat model.

[0054] The specific implementation of the present invention described above does not constitute a limitation on the protection scope of the present invention. Any other corresponding changes and modifications made based on the technical concept of the present invention should be included in the protection scope of the claims of the present invention.

Claims

1. A plant fermentation product, characterized in that: The fermentation raw materials of the fermentation product include one or more of Aronia nigra, Acai, Inula villosa stems and leaves, Moringa oleifera leaves, Andrographis paniculata leaves and Ginkgo biloba leaves.

2. The plant fermentation product according to claim 1, characterized in that Obtained after at least two fermentations.

3. The plant fermentation product according to claim 2, characterized in that The two fermentation steps are as follows: S1 preferably uses 5Kg of Aronia nigra, 5kg of Acai, and 3kg of Inula villosa stems and leaves, which are quickly washed at low temperature. After the Aronia nigra and Acai are destemmed, they are mixed with the stems and leaves of Inula villosa, and crushed. During the crushing, 65mg / L SO2 and 25mg / L pectinase are added. After fully mixing, the mixture is allowed to stand at 4°C for 20 hours, and then 2kg of saturated sterilized sucrose solution is added. 120cfu / mL of brewer's yeast is inoculated and fermented at 26±2°C for 5-7 days. The skin and residue are separated to obtain the first stage fermentation liquid. S2: 2kg of Moringa leaves, 4kg of Andrographis paniculata leaves and 4kg of Ginkgo biloba leaves are washed at low temperature, centrifuged and dried, chopped and sterilized, and then poured into the fermentation liquid of step S1, pectinase is added and one or more of activated lactic acid bacteria, Lactobacillus plantarum, Lactobacillus rhamnosus and Leuconostoc mesenteroides are inoculated for fermentation, and fermented at 26-44°C for 38-50 hours to obtain plant fermentation products.

4. The plant fermentation product according to claim 1, characterized in that The fermentation product contains no less than 3% of andrographolide, no less than 1% of quinic acid and no less than 0.5% of shikimic acid.

5. A method for preparing a plant fermentation product, characterized in that: The fermentation raw materials of the fermentation product include one or more of Aronia melanocarpa, Acai, Inula villosa L. extract, Moringa oleifera leaf, Andrographis paniculata leaf and Ginkgo biloba leaf.

6. The method for preparing a plant fermentation product according to claim 5, characterized in that: Goes through at least two fermentation steps.

7. The method for preparing a plant fermentation product according to claim 6, characterized in that: The two fermentation steps are as follows: S1 preferably uses 5Kg of Aronia nigra, 5kg of Acai, and 3kg of Inula villosa stems and leaves, which are quickly washed at low temperature. After the Aronia nigra and Acai are destemmed, they are mixed with the stems and leaves of Inula villosa, and crushed. During the crushing, 65mg / L SO2 and 25mg / L pectinase are added. After fully mixing, the mixture is allowed to stand at 4°C for 20 hours, and then 2kg of saturated sterilized sucrose solution is added. 120cfu / mL of brewer's yeast is inoculated and fermented at 26±2°C for 5-7 days. The skin and residue are separated to obtain the first stage fermentation liquid. S2: 2kg of Moringa leaves, 4kg of Andrographis paniculata leaves and 4kg of Ginkgo biloba leaves are washed at low temperature, centrifuged and dried, chopped and sterilized, and poured into the fermentation liquid of step S1, pectinase is added, and one or more of activated lactic acid bacteria, Lactobacillus plantarum, Lactobacillus rhamnosus and Leuconostoc mesenteroides are inoculated for fermentation, and fermented at 26-44°C for 38-50 hours to obtain plant fermentation products.

8. The method for preparing a plant fermentation product according to claim 5, characterized in that: The fermentation product contains no less than 3% of andrographolide, no less than 1% of quinic acid and no less than 0.5% of shikimic acid.

9. The plant fermentation product according to claim 1, wherein the plant fermentation product is used for reducing postoperative infection.

10. The method for preparing plant fermentation products according to claim 5, and use thereof in preparing compositions and products for reducing surgical infection.

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