In-vitro fermentation synchronous dehydration device and dehydration method

Abstract

The invention relates to an in-vitro fermentation synchronous dehydration device and a dehydration method, and the device comprises a colon bioreactor main body which is provided with a sealed cavity structure; the dialysis bag is arranged in the cavity of the colon bioreactor main body and is used for accommodating fermentation liquor; the hypertonic solution is filled between the dialysis bag and the inner wall of the colon bioreactor main body, and the osmotic pressure of the hypertonic solution is higher than that of the fermentation liquor; wherein the moisture is automatically transferred from the inside of the dialysis bag to the outside by virtue of the osmotic pressure difference between the fermentation liquid and the hypertonic solution, so that synchronous simulation of dynamic viscosity change and dehydration in the fermentation process is realized. The device is simple in structure, low in cost, high in adaptability and capable of synchronously simulating the colon peristalsis and dehydration process so as to improve the physiological simulation degree of in-vitro experiments.

Description

Technical Field

[0001] This invention relates to the field of in vitro simulated fermentation technology, and in particular to an in vitro fermentation simultaneous dehydration device and dehydration method. Background Technology

[0002] In vivo colonic fermentation is a complex physiological process involving multidimensional interactions between microorganisms, substrates, and the host. It is regulated by various factors, including intestinal peristalsis, immune cells, and blood circulation, making it difficult to analyze the mechanism of action of any single variable in isolation, and it is also subject to ethical constraints. In vitro colonic fermentation technology, by precisely controlling environmental parameters such as temperature, pH, and anaerobic conditions, as well as initial variables such as microbial composition and substrate type, effectively eliminates irrelevant interferences and has become an important tool for studying the function of the gut microbiota.

[0003] The advent of in vitro colonic fermentation has, to some extent, reduced the complexity, ethical implications, and uncontrollability of in vivo colonic fermentation. The correlation between the physical parameters of in vivo and in vitro colonic fermentation has largely focused on temperature, pH, and dissolved oxygen levels, with less attention paid to the viscosity of the fermentation substrate and the human body, as well as the dehydration process during fermentation. As food residue passes through the large intestine, water is gradually absorbed, and the residue gradually becomes semi-solid. However, current in vitro fermentation devices generally lack the ability to simulate the simultaneous dehydration process, resulting in significant differences in the physicochemical properties of the fermentation products compared to real feces, thus affecting the physiological relevance of experimental results.

[0004] To address these issues, some studies have attempted to introduce dehydration devices into in vitro fermentation, often employing hollow fiber membrane technology. Hollow fiber membranes, with their hollow tubular self-supporting structure, pass through the reactor chamber and are encapsulated by the fermentation products, removing water through the pressure difference between the inside and outside of the membrane. However, this technology has significant drawbacks: firstly, it requires complex equipment such as high-pressure pumps, membrane modules, and piping systems, making operation cumbersome and costly; secondly, the hollow fiber membrane material has limited flexibility, making it prone to clogging in high-viscosity fermentation products, and its rigid structure is unsuitable for the dynamic compression environment that requires simulating intestinal peristalsis. Summary of the Invention

[0005] Therefore, the technical problem to be solved by the present invention is to overcome the shortcomings of the prior art and provide an in vitro fermentation simultaneous dehydration device and dehydration method, which has a simple structure, low cost, strong adaptability and can simultaneously simulate colon peristalsis and dehydration process, so as to improve the physiological simulation degree of in vitro experiments.

[0006] To solve the above-mentioned technical problems, the present invention provides an in vitro fermentation and simultaneous dehydration device, comprising: Colon bioreactor with a sealed cavity structure; A dialysis bag is placed inside the cavity of the main body of the colon bioreactor to contain the fermentation broth; A hypertonic solution is filled between the dialysis bag and the inner wall of the colon bioreactor body, and the osmotic pressure of the hypertonic solution is higher than that of the fermentation broth; In this process, water is automatically transferred from the inside of the dialysis bag to the outside by relying on the osmotic pressure difference between the fermentation broth and the hypertonic solution, thus simulating the dynamic changes in viscosity and dehydration during the fermentation process.

[0007] In one embodiment of the present invention, the colon bioreactor includes a flexible pipe, a power device for driving the flexible pipe to generate peristaltic squeezing, a heating device, and a detection probe, the detection probe including a pH probe, an oxygen content probe, and a temperature probe.

[0008] In one embodiment of the present invention, the tail of the dialysis bag is sealed, and the top is fixed to the body of the colon bioreactor by a rigid connector; the flexible tube is sleeved on the outside of the dialysis bag, the top of which is sealed to the rigid connector, and the bottom is provided with an openable and closable sealing clip.

[0009] In one embodiment of the present invention, the colon bioreactor is provided with a first sampling tube, a feed tube, an acid tube, and an alkali tube, and the free ends of the above four tubes all extend into the dialysis bag; the first sampling tube is used to sample the fermentation broth, the feed tube is used to replenish the fermentation broth, the acid tube is used to inject acidic liquid, and the alkali tube is used to inject alkaline liquid. A second sampling tube is also provided on the flexible pipe, and the second sampling tube communicates with the interlayer space for injecting the hypertonic solution.

[0010] In one embodiment of the present invention, a peristaltic pump assembly is further included, comprising an acid pump, an alkali pump, a feed pump, and a standby pump; the feed pipe is connected to the feed pump; the acid pipe is connected to the acid pump; the alkali pipe is connected to the alkali pump; the standby pump is a peristaltic pump and is arranged in parallel with the acid pump, alkali pump, and feed pump, for use in place of any of the acid pump, alkali pump, or feed pump in case of failure, or for connecting to a first sampling pipe for sampling.

[0011] Secondly, in order to solve the above-mentioned technical problems, the present invention provides a method for in vitro fermentation and dehydration using the in vitro fermentation and simultaneous dehydration device described in the first aspect, comprising the following steps: (1) Activate and sterilize the dialysis bags; (2) The lower end of the dialysis bag is sealed and the upper end is sealed and connected to the rigid connection part, so that the dialysis bag hangs inside the flexible tube; the upper end of the flexible tube is sealed and connected to the rigid connection part, and the lower end is sealed, forming an interlayer space between the outer wall of the dialysis bag and the inner wall of the flexible tube; a detection probe is installed in the rigid connection part and extends into the dialysis bag; a second sampling tube communicating with the interlayer space is connected. (3) Prepare fermentation culture medium to simulate artificial feces with different viscosities. Mix the culture medium with the probiotic sample to form a fermentation liquid and then inject it into the dialysis bag. (4) After removing the air from the interlayer space, inject a hypertonic solution; (5) Under the simulated in vivo fermentation environment of temperature, pH and dissolved oxygen, periodic peristaltic squeezing is applied to the flexible pipe to continuously transfer water from the dialysis bag to the interlayer space. (6) The pH value of the fermentation broth is monitored in real time by a detection probe, and acid or alkali solution is automatically added when the pH value is lower than the set threshold; (7) Periodically extract hypertonic solution and replenish with fresh hypertonic solution to maintain osmotic pressure differential; (8) After fermentation, stop squeezing, drain the hypertonic solution from the interlayer space, and take out the dehydrated fermentation product from the dialysis bag.

[0012] In one embodiment of the present invention, the dialysis bag has a molecular weight cutoff of 50-500 Da and is made of a soft material that can deform synchronously with the compression and peristalsis of the flexible tube.

[0013] In one embodiment of the present invention, the hypertonic solution is a polyethylene glycol solution.

[0014] In one embodiment of the present invention, the fermentation broth is a colon culture medium of sodium carboxymethyl cellulose, used to simulate artificial feces of different viscosities.

[0015] In one embodiment of the present invention, the colon culture medium comprises: tryptone, yeast extract, sodium bicarbonate, sodium chloride, dipotassium hydrogen phosphate, magnesium sulfate heptahydrate, calcium chloride dihydrate, L-cysteine, heme, bile salts, Tween 80, vitamin K1, resazurin solution, and sodium carboxymethyl cellulose.

[0016] Compared with the prior art, the above-described technical solution of the present invention has the following advantages: The in vitro fermentation and simultaneous dehydration device of the present invention achieves automatic water transfer through the osmotic pressure difference between the fermentation liquid and the hypertonic medium in the dialysis bag. It can simultaneously simulate the dehydration mechanism of the human colon during the fermentation process, simulating the dehydration process of real feces. At the same time, it avoids the complex high-pressure system required by hollow fiber membranes, significantly simplifies the structure, reduces costs, and is not limited by the viscosity of the fermentation liquid, making it suitable for high-viscosity semi-solid fermentation systems. Attached Figure Description

[0017] To make the content of this invention easier to understand, the invention will be further described in detail below with reference to specific embodiments and accompanying drawings.

[0018] Figure 1This is a schematic diagram of the structure of the in vitro fermentation and simultaneous dehydration device in a preferred embodiment of the present invention; Figure 2 for Figure 1 Top view of the in vitro fermentation and simultaneous dehydration device; Figure 3 This is a schematic diagram of the dehydration process of the in vitro fermentation simultaneous dehydration device of the present invention; Figure 4 The amount of water dehydrated during the fermentation process of the two probiotic products in Example 2 of this invention; Figure 5 A comparison table of pH changes of the two products in culture media of different viscosities in Example 2 of this invention; Figure 6 The variation of viable bacteria concentration in culture media of different viscosities for the two products of this invention; Figure 7 The shear viscosity changes of the two products of this invention in culture media of different viscosities. Figure 8 This invention illustrates the viscosity changes of two products in culture media of different viscosities when the shear rate is 1 s⁻¹. Figure 9 This invention illustrates the viscosity changes of two products in culture media of different viscosities when the shear rate is 10 s⁻¹. Figure 10 This invention illustrates the viscosity changes of two products in culture media of different viscosities when the shear rate is 100 s⁻¹. Explanation of reference numerals in the accompanying drawings: 1. Reactor body; 11. Flexible pipe; 12. Power unit; 13. Heating device; 16. pH probe; 17. Oxygen content probe; 18. Heating environment temperature probe; 19. Reactor internal temperature probe; 2. Dialysis bag; 3. Fermentation broth; 4. Hypertonic solution; 100. First sampling tube; 101. Feeding tube; 102. Acid tube; 103. Alkali tube; 111. Second sampling tube. Detailed Implementation

[0019] The present invention will be further described below with reference to the accompanying drawings and specific embodiments, so that those skilled in the art can better understand and implement the present invention. However, the embodiments described are not intended to limit the present invention. Example 1

[0020] Reference Figure 1 and 2 As shown, the present invention provides an in vitro fermentation and simultaneous dehydration device, comprising: The main body 1 of the colon bioreactor has a sealed cavity structure; A dialysis bag 2 is disposed within the cavity of the main body 1 of the colon bioreactor and is used to contain the fermentation broth 3; the dialysis bag 2 is made of flexible material. Hypertonic solution 4 is filled between the dialysis bag 2 and the inner wall of the colon bioreactor body 1, and the osmotic pressure of the hypertonic solution 4 is higher than that of the fermentation broth 3; In this process, water is automatically transferred from the inside of the dialysis bag 2 to the outside by relying on the osmotic pressure difference between the fermentation liquid 3 and the hypertonic solution 4, thereby simulating the dynamic change of viscosity and dehydration during the fermentation process.

[0021] The main body 1 of the colon bioreactor is a colon bioreactor used to simulate the digestive environment of the human colon. By explicitly defining the main body 1 as a colon bioreactor, the equipment is specifically designed for in vitro simulation studies of human intestinal microorganisms, improving the physiological relevance of experimental conditions and providing a more realistic experimental platform for evaluating the efficacy of probiotics and studying intestinal metabolic mechanisms.

[0022] In this embodiment, the dialysis bag 2 and the hypertonic solution 4 can constitute a detachable dehydration module. The dehydration module can be pre-installed in the main body 1 of the colon bioreactor, or it can be produced and sold as an independent component.

[0023] It should be noted that although this embodiment uses a colon bioreactor as an example, those skilled in the art will understand that the dehydration module can be installed in other types of sealed fermentation reactors to achieve the function of fermentation and dehydration at the same time.

[0024] Specifically, the colon bioreactor includes a flexible pipe 11, a power device 12 that drives the flexible pipe 11 to generate peristaltic extrusion, a heating device 13, and a detection probe. The detection probe includes a pH probe 16, an oxygen content probe 17, and a temperature probe. The temperature probe includes a heating environment temperature probe 18 and a reactor internal temperature probe 19.

[0025] The tail of the dialysis bag 3 is sealed, and its top is fixed to the main body 1 of the colon bioreactor via a rigid connector. The flexible pipe 11 is sleeved on the outside of the dialysis bag 2, with its top sealed to the rigid connector and its bottom equipped with an openable and closable sealing clip. The rigid connector is preferably a clamp, and the combination of the clamp and the sealing clip achieves coaxial sealing and fixation between the dialysis bag 2 and the flexible pipe, forming an independent dehydration chamber. This ensures that the hypertonic medium does not leak and the fermentation broth 3 is not contaminated. This structure supports rapid disassembly and assembly, facilitating sterilization, replacement of the dialysis bag 2, and cleaning and maintenance, improving experimental efficiency and reducing the risk of cross-contamination.

[0026] In addition, the colon bioreactor 1 is equipped with a first sampling tube 100, a feeding tube 101, an acid tube 102, and an alkali tube 103, and the free ends of the above four tubes extend into the dialysis bag 2; the first sampling tube 100 can sample the fermentation broth in the dialysis bag 2, the feeding tube 101 is used to feed fermentation broth 3 into the dialysis bag 2, the acid tube 102 is used to inject acidic liquid into the dialysis bag 2, and the alkali tube 103 is used to inject alkaline liquid into the dialysis bag 2; wherein, a second sampling tube 111 is also provided on the body of the flexible pipe 11, and the second sampling tube 111 is connected to the interlayer space for injecting the hypertonic solution 4.

[0027] Furthermore, it also includes a peristaltic pump assembly, which includes an acid pump, an alkali pump, a feed pump, and a standby pump; the feed pipe 101 is connected to the feed pump; the acid pipe 102 is connected to the acid pump; the alkali pipe 103 is connected to the alkali pump; the standby pump is a peristaltic pump, and is arranged in parallel with the acid pump, alkali pump, and feed pump, for use in place of any of the acid pump, alkali pump, or feed pump in case of failure, or for connecting to the first sampling pipe for sampling.

[0028] In practical use, the dialysis bag 2 is installed inside the colon bioreactor body 1; the fermentation broth 3 is inoculated into the inside of the dialysis bag 2; a hypertonic solution 4 is injected between the dialysis bag 2 and the inner wall of the colon bioreactor body 1; the reaction conditions are controlled so that the fermentation broth 3 ferments inside the colon bioreactor body, and water is automatically transferred from the fermentation broth 3 to the hypertonic solution 4. Example 2

[0029] This invention provides an in vitro fermentation dehydration method, implemented using the in vitro fermentation simultaneous dehydration device described in Example 1, specifically including the following steps: The dialysis bags are activated and sterilized. The dialysis bag 2 is sealed at its lower end and open at its upper end, and connected to a rigid connection, so that the dialysis bag 2 hangs inside the flexible tube 11; the upper end of the flexible tube 11 is sealed and connected to the rigid connection, and the lower end is closed, forming an interlayer space between the outer wall of the dialysis bag 2 and the inner wall of the flexible tube 11; a detection probe is installed at the rigid connection and extends into the dialysis bag 2; a second sampling tube communicating with the interlayer space is connected. (3) Prepare fermentation culture medium to simulate artificial feces with different viscosities. Mix the culture medium with the probiotic sample to form a fermentation liquid and then inject it into the dialysis bag. (4) After removing the air from the interlayer space, inject a hypertonic solution 4; (5) Under the simulated in vivo fermentation environment of temperature, pH and dissolved oxygen, periodic peristaltic extrusion is applied to the flexible pipe 11 to continuously transfer water from the dialysis bag 2 to the interlayer space; in this embodiment, at 37±0.5℃, periodic extrusion is applied to the flexible pipe 11 at a frequency of 10-30 times per minute to maintain the pH value of the fermentation liquid at 6.5-7.5 and the dissolved oxygen content below 0.1mg / L, so that water can continuously transfer from the dialysis bag 2 to the interlayer space through the osmotic pressure difference; (6) The pH value and viable bacteria concentration of the fermentation broth are monitored in real time by a detection probe. When the pH value is lower than the set threshold, acid and alkali solutions are automatically added. (7) Periodically extract hypertonic solution and replenish with fresh hypertonic solution to maintain osmotic pressure differential; (8) After fermentation, stop squeezing, drain the hypertonic solution 4 from the interlayer space, and take out the dehydrated fermentation product from the dialysis bag 2.

[0030] Preferably, the hypertonic solution 4 is a polyethylene glycol solution with a mass concentration of 15-25 wt%. In this embodiment, the mass concentration of the polyethylene glycol solution is 20 wt%. By using polyethylene glycol solution as the hypertonic medium, the osmotic pressure is stable and the concentration range is wide, allowing for flexible adjustment of the dehydration rate according to different fermentation stages or experimental needs. Polyethylene glycol is chemically inert and does not participate in microbial metabolism, ensuring a pure and controllable dehydration process. Furthermore, the material is economical and readily available, further reducing experimental costs.

[0031] In this embodiment, the molecular weight cutoff of the dialysis bag 2 is 50-500 Da. In this experiment, the specific molecular weight cutoff used in the dialysis bag 2 is 100 Da, and it is made of a soft material that can deform synchronously with the compression and peristalsis of the flexible tube 11. The molecular weight cutoff of 50-500 Da in the dialysis bag 2 can effectively retain microorganisms, enzymes, and metabolites, allowing only water and small molecule solutes to pass through, thus ensuring the integrity of the fermentation system. The soft material of the dialysis bag 2 allows it to deform synchronously with the flexible tube 11, avoiding additional resistance or mechanical damage, making it particularly suitable for the long-term dynamic culture of high-viscosity ferments.

[0032] In this embodiment, fermentation broth 3 is a colon culture medium containing 0-4.5 wt% sodium carboxymethyl cellulose, used to simulate artificial feces of different viscosities.

[0033] Those skilled in the art can routinely adjust the concentration of the polyethylene glycol solution and the molecular weight cutoff of the dialysis bag 2 within the parameter range disclosed in this invention, based on factors such as the composition of the fermentation broth, the target dehydration rate, and the molecular weight of the product. Such adjustments fall within the protection scope of this invention.

[0034] The colon culture medium described in this embodiment contains: tryptone, yeast extract, sodium bicarbonate, sodium chloride, dipotassium hydrogen phosphate, magnesium sulfate heptahydrate, calcium chloride dihydrate, L-cysteine, heme, bile salts, Tween 80, vitamin K1, and resazurin solution.

[0035] The specific method for preparing the culture medium is as follows: 1L of basal culture medium contains tryptone (2.0 g), yeast extract (2.0 g), sodium bicarbonate (0.4 g), sodium chloride (0.1 g), dipotassium hydrogen phosphate (0.04 g), magnesium sulfate heptahydrate (0.01 g), calcium chloride dihydrate (0.01 g), L-cysteine ​​(0.5 g), heme (0.02 g), bile salts (0.5 g), Tween 80 (2.0 mL), vitamin K1 (10.0 μL), and resazurin solution (4.0 mL, 0.025%, w / v). Add the appropriate mass of sodium carboxymethyl cellulose to adjust the pH of the colon culture medium to 7.2. When adding sodium carboxymethyl cellulose, add it while heating, in batches, at a temperature of 80-90℃.

[0036] This method can be used to prepare culture media containing different concentrations of sodium carboxymethyl cellulose, which have different viscosities and rheological properties. Taking the colonic culture medium with 0 wt% (culture medium 1) sodium carboxymethyl cellulose and the culture media with 3.0 wt% (culture medium 2) and 4.5 wt% (culture medium 3) sodium carboxymethyl cellulose as examples, these are three concentrations of "artificial feces".

[0037] Those skilled in the art can achieve the core function of simultaneous fermentation-dehydration simulation by routinely adjusting the types and contents of each component within the basic formulation framework. Moreover, the adjusted culture medium can specifically reflect the characteristics of different intestinal physiological / pathological states. These adjustments do not require creative labor and are routine optimizations by those skilled in the art based on experimental purposes, and all fall within the protection scope of this invention.

[0038] Specifically, the treatment process for dialysis bag 2 is as follows: Dialysis bag 2 with a molecular weight of 100 Da is cut to a suitable length before use; dialysis bag 2 is immersed in a sufficient amount of 2% (w / v) sodium bicarbonate and 1 mmol / L EDTA-2Na (pH = 8.0) solution and boiled continuously for 10 min to moisten the hollow fibers inside the dialysis bag 2; dialysis bag 2 is rinsed with deionized water and immersed again in a sufficient amount of 1 mmol / L EDTA-2Na (pH = 8.0) solution and boiled for 10 min to remove metal ions adhering to the dialysis bag 2 during the production process; after cooling, it is rinsed with deionized water, sterilized, and then ready for use. The specific process for inoculating the sample is as follows: the culture medium and product 1 or product 2 are mixed to obtain fermentation broth 3. The fermentation broth 3 is pumped into dialysis bag 2 using a peristaltic pump. If the fermentation broth 3 is of high viscosity, it can be poured into the dialysis bag 2 of the reactor from the top in a clean bench.

[0039] Add polyethylene glycol solution: Connect the sampling tube with a disposable syringe, extract the air between the flexible tube 11 and the dialysis bag 2, and then inject 70 mL of polyethylene glycol solution with a mass concentration of 20 wt%.

[0040] Two probiotic powder products for constipation were used (Product 1 and Product 2), both of which contain multiple types of lactobacilli and bifidobacteria.

[0041] Two products, Product 1 and Product 2, were cultured for 24 hours in colonic medium containing 0 wt% sodium carboxymethyl cellulose (medium 1), 3.0 wt% sodium carboxymethyl cellulose (medium 2), and 4.5 wt% sodium carboxymethyl cellulose (medium 3). The dehydration process was as follows: Figure 2 As shown in the figure. The water content, pH, viable lactobacillus concentration, and shear viscosity before and after 24 hours of fermentation are respectively as follows: Figure 4 , Figure 4 and Figure 6 , Figure 7 As shown, different products perform differently in culture media of different viscosities, and the dehydration effect of polyethylene glycol also varies.

[0042] Depend on Figure 4 It can be seen that the higher the concentration of the fermentation broth, the more difficult it is to dehydrate. The dehydration effect of the fermentation products in culture medium 1 is the most obvious, while the dehydration effect in culture medium 3 is the least obvious.

[0043] Depend on Figure 2 As shown, due to the concentration difference between the inside and outside of dialysis bag 2, the concentration of the 20wt% polyethylene glycol solution is relatively high, and water molecules diffuse from the fermentation product side to the polyethylene glycol solution side, thereby achieving water transfer.

[0044] Depend on Figure 4 It can be seen that lactic acid is produced by lactobacilli and bifidobacteria during fermentation, which lowers the pH value in the intestine. The lower pH value of Product 1 during fermentation may be due to the slightly higher activity of the lactobacillus strain in Product 1 compared to Product 2, or the weaker pH buffering capacity of Product 1.

[0045] Taking lactobacillus as an example, from Figure 6 It can be seen that when the two products are fermented in media of different viscosities, the growth trend of lactobacillus is more obvious in the viscous media, while it shows a decreasing trend in the media without the addition of sodium carboxymethyl cellulose.

[0046] Depend on Figure 8 As can be seen from the figure, at shear rates of 0.001-100 1 / s (logarithmic ordinate), region ③ represents the shear viscosity changes of the two products before and after 24 hours of fermentation in medium 1 (without sodium carboxymethyl cellulose) with low viscosity. It can be seen that the shear viscosity is higher after 24 hours of fermentation than at 0 hours. Table 1 shows that the two products lost 160.2g and 165.1g of water respectively during fermentation, approximately 40% of the total volume. The increase in shear viscosity is likely due to the reduction in water content. Region ② represents the shear viscosity changes of the two products before and after 24 hours of fermentation in medium 2. It can be seen that the shear viscosity of product 2 increases significantly after 24 hours of fermentation, while the change in product 1 is not significant. Meanwhile, from... Figure 4 It can be seen that the water loss of product 2 is significantly higher than that of product 1, and its shear viscosity shows a similar trend. Region ① in the figure represents the shear viscosity changes of the two products before and after 24 hours of fermentation in culture medium 3. It can be seen that the shear viscosity of product 1 increases significantly after 24 hours of fermentation, while the change in product 2 is not significant. Figure 4 It can be seen that the water loss of Product 1 after 24 hours of fermentation is significantly higher than that of Product 2, and this trend is similar to the change in its shear viscosity. Figure 7 The changes in shear viscosity of the fermentation products and the results of dehydration in Table 1 show that the amount of water loss is related to the change in shear viscosity. The more water is removed from the fermentation products, the more significant the change in shear viscosity. The lower the water content of the fermentation products, the higher the shear viscosity. This is similar to the physical changes that occur in the formation of feces in the human large intestine. As food residue passes through the large intestine, water is gradually absorbed, the food residue gradually becomes semi-solid, and the viscosity gradually increases. This experiment demonstrates that it is feasible and meaningful to use equipment that removes water while simultaneously fermenting in vitro to simulate the formation / changes of feces in the human large intestine.

[0047] Depend on Figure 8 , 9 As can be seen from Figure 10, comparing the changes in viscosity on the linear ordinate, at a shear rate of 1 s⁻¹, in the viscous culture medium 3, the viscosity of the fermentation product of product 1 increases with increasing fermentation time, while the viscosity of the fermentation product of product 2 decreases slightly with increasing fermentation time. In culture medium 2, the viscosity of the fermentation product of product 2 increases slightly with increasing fermentation time, while the viscosity of the fermentation product of product 1 does not change significantly with increasing fermentation time. In the liquid culture medium (culture medium 1), the differences in rheological properties between the products are very small with increasing fermentation time.

[0048] The results above show that the changes in pH, shear viscosity, and viable bacteria concentration during the fermentation process of different products are all affected by the product's composition. Therefore, it is of great guiding significance to use a clean "artificial feces" in vitro to simulate the viscosity and dehydration process of human feces and then predict the differences in the effects of fermentation of different product components in the human body.

[0049] Obviously, the above embodiments are merely illustrative examples for clear explanation and are not intended to limit the implementation. Those skilled in the art will recognize that other variations or modifications can be made based on the above description. It is neither necessary nor possible to exhaustively list all possible implementations here. However, obvious variations or modifications derived therefrom are still within the scope of protection of this invention.

Claims

1. An in-vitro fermentation and simultaneous dehydration device, characterized by, The application relates to a colon bioreactor, which comprises the following parts: a sealed cavity structure; a dialysis bag arranged in the cavity of the colon bioreactor body and used for containing fermentation liquor; and a high-osmotic solution filled between the dialysis bag and the inner wall of the colon bioreactor body, wherein the osmotic pressure of the high-osmotic solution is higher than that of the fermentation liquor; wherein water is automatically transferred from the inside to the outside of the dialysis bag by the osmotic pressure difference between the fermentation liquor and the high-osmotic solution, so that the dynamic viscosity change and dehydration simulation in the fermentation process are realized. The colon bioreactor comprises a flexible pipeline, a power device for driving the flexible pipeline to generate peristaltic extrusion, a heating device and a detection probe, wherein the detection probe comprises a PH probe, an oxygen content probe and a temperature probe. The tail part of the dialysis bag is sealed, and the top part is fixed to the colon bioreactor body through a rigid connecting piece; the flexible pipeline is sleeved outside the dialysis bag, the top part of the flexible pipeline is sealingly connected with the rigid connecting piece, and the bottom part is provided with a sealable sealing clamp, so that a sandwich space is formed between the outer wall of the dialysis bag and the inner wall of the flexible pipeline. The colon bioreactor is provided with a first sampling pipe, a feeding pipe, an acid liquid pipe and an alkali liquid pipe, and the free ends of the four pipe fittings are all inserted into the dialysis bag; the first sampling pipe is used for sampling the fermentation liquor, the feeding pipe is used for feeding the fermentation liquor, the acid liquid pipe is used for injecting acid liquid, and the alkali liquid pipe is used for injecting alkali liquid; a second sampling pipe is further arranged on the pipeline of the flexible pipeline and is in communication with the sandwich space and used for injecting the high-osmotic solution. The colon bioreactor further comprises a peristaltic pump set, which comprises an acid pump, an alkali pump, a feeding pump and a standby pump; the feeding pipe is connected with the feeding pump; the acid liquid pipe is connected with the acid pump; the alkali liquid pipe is connected with the alkali pump; the standby pump is a peristaltic pump and is arranged in parallel with the acid pump, the alkali pump and the feeding pump and used for replacing any one of the acid pump, the alkali pump and the feeding pump to work or connecting the first sampling pipe to sample when the pump fails.

2. The in-vitro fermentation and simultaneous dehydration device according to claim 1, characterized in that, The application further comprises the following steps:

3. The in-vitro fermentation and simultaneous dehydration device according to claim 3, characterized in that, (1) activating and sterilizing the dialysis bag; 4. The in-vitro fermentation and simultaneous dehydration device according to claim 3, characterized in that, (2) sealing the lower end of the dialysis bag, sealingly connecting the upper end of the dialysis bag to the rigid connecting part, making the dialysis bag hang in the flexible pipeline, sealingly connecting the upper end of the flexible pipeline to the rigid connecting part and sealing the lower end, forming a sandwich space between the outer wall of the dialysis bag and the inner wall of the flexible pipeline and connecting the second sampling pipe in communication with the sandwich space; 5. The in-vitro fermentation and simultaneous dehydration device according to claim 4, characterized in that (3) preparing a fermentation medium to simulate artificial feces with different viscosities, mixing the medium with a probiotic sample to form fermentation liquor and then injecting the fermentation liquor into the dialysis bag; 6. A method for in-vitro fermentation dehydration using the in-vitro fermentation simultaneous dehydration device according to any one of claims 1 to 5, characterized by, (4) extracting air in the sandwich space and then injecting a high-osmotic solution; (5) continuously transferring water from the dialysis bag to the sandwich space by periodically extruding the flexible pipeline under the conditions of temperature, pH and dissolved oxygen simulating the in-vivo fermentation environment; (6) monitoring the pH value of the fermentation liquor in real time through the detection probe and automatically adding acid liquid or alkali liquid when the pH value is lower than a set threshold value; (7) regularly extracting the high-osmotic solution and supplementing fresh high-osmotic solution to maintain the osmotic pressure difference; (8) stopping the extrusion and discharging the high-osmotic solution in the sandwich space after the fermentation is completed, and then taking out the dehydrated fermentation product from the dialysis bag. ​ ​ ​ ​ 7. The in-vitro fermentation dehydration method according to claim 6, characterized in that, The dialysis bag is made of soft material and can be deformed synchronously with the extrusion peristalsis of the flexible pipeline.

8. The in-vitro fermentation dehydration method according to claim 6, characterized in that, The high-osmotic solution is a polyethylene glycol solution.

9. The in-vitro fermentation dehydration method according to claim 6, characterized in that, The fermentation liquid is a colon culture medium containing sodium carboxymethyl cellulose, which is used for simulating artificial feces.

10. The in-vitro fermentation dehydration method according to claim 9, characterized in that, The colon culture medium comprises: tryptone, yeast extract, sodium bicarbonate, sodium chloride, dipotassium hydrogen phosphate, magnesium sulfate heptahydrate, calcium chloride dihydrate, L-cysteine, hematin, bile salt, Tween 80, vitamin K1, resazurin solution and sodium carboxymethyl cellulose.

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