Development And Application Of A Novel Bionic Gastrointestinal Tract Bioreactor

The gastrointestinal bioreactor is designed for studying food digestion by simulating the physiological conditions of the human gastrointestinal tract in vitro,such as temperature,dynamic p H,digestive enzyme secretion,food residence time,flow mixing and gastrointestinal wall peristalsis.Besides,it can sort out numerous substances,including dietary ingredients,pathogens,active pharmaceutical ingredients,and toxic or radioactive compounds,with the aim of evaluating the way in which they change the gastrointestinal environment via freeing the sampling process of ethical constraints.However,compared with advanced foreign equipment,the domestic gastrointestinal reactor is still in its infancy,and it is difficult to realize the simulation of the actual gastrointestinal digestion process,which poses obstacles during the interdisciplinary research on the digestion of foods in China.In this paper,the bionics technology was employed to simulate the geometry and internal structure of the gastrointestinal tract,and the bionic silica gel stomach,small intestine and large intestine were prepared;the internal environment simulation technology was applied to the control of temperature,p H,peristaltic frequency and endocrine parameters;a colonization model of intestinal microorganisms with high efficiency was established through the fermentation engineering technology.On this basis,an intestinal gas array sensor was integrated with the intelligent control system,and a bionic gastric,small intestine and large intestine bioreactor were developed.The reactor was gradually put into an application through Akkermansia muciniphila dynamic fermentation culture,in vitro fecal colonization culture,in vitro digestion of highly resistant starch rice,the effect of resistant starch on fecal fermentation,and the effect of dietary fatty acids on intestinal gas distribution.The main research results in this paper are as follows:(1)The development of a biomimetic stomach and small intestine combined bioreactor and research on its in vitro digestionA biomimetic stomach and small intestine combined bioreactor with up to 9 chambers was developed.In terms of the geometry of the gastrointestinal tract,this bioreactor can simulate the gastric fundus,gastric body,gastric antrum,duodenum,jejunum,and ileum compartment reactors.It was convenient to disassemble and sterilize those compartments,which,as a result,can be used independently or in series.In terms of the control system,the offline control system and online cloud platform control system were developed,which can realize the detection of peristaltic frequency,secretion rate,p H,etc.,and the control of historical data export and operation status alarm.In terms of the simulation of the internal structure of the gastrointestinal tract,the smooth silica gel stomach and the silica gel small intestine,the pleated silica gel stomach and the fluffy silica gel small intestine were prepared respectively,which increased the surface area of the intestine,changed the rheological entropy of the chyme,and promoted the food fragmentation.In terms of the mixing effect,the bioreactor can achieve a good mixing effect for both Newtonian fluid and non-Newtonian fluid,with better performance than the traditional kettle-type stirred reactor under the same conditions.In terms of the gastric pressure,the bioreactor can realize the peristaltic contraction of the stomach through the control of basic motion mode and strong motion mode,under which the contraction strength can reach 18-22 mm Hg and 120-220 mm Hg respectively.In terms of the crushing force,the maximum crushing force of the bioreactor was greater than 0.72 N,which was enough to simulate the crushing function of the stomach.In terms of the control of p H,the bioreactor can adjust p H dynamically according to the digestion process of food,which can reproduce the dynamic adjustment of enzyme activity in the stomach and small intestine.In terms of the emptying rate,there was no significant difference compared with the emptying of piglet's stomach that has been published.In terms of application,wheat flour,potato flour,corn flour,sweet potato flour,lotus root flour and rice flour were employed to simulate the dynamic digestion of starch and protein in the bionic stomach and small intestine reactors,and it was found that there were obvious differences in the digestion stages of the stomach and small intestine.(2)Research on the development of biomimetic large intestine bioreactor and its fecal colonizationA biomimetic large intestine bioreactor was developed in this study,with up to 6 chambers.Besides,the offline control system and online cloud platform intelligent control system were integrated,which can effectively control the key parameters of the fermentation process(peristaltic frequency,secretion rate,absorption rate,p H real-time curve,etc.).In terms of the structure of the large intestine,smooth and wrinkled silica gel large intestines were developed.In terms of the mixing effect,this bioreactor was superior to the tank-type stirred reactor under the same conditions.In terms of the intestinal pressure,two kinds of peristaltic frequencies,namely the low and high frequencies,were simulated,with the internal contraction strength reaching 60-90 mm Hg and 100-150 mm Hg respectively.In terms of the simulated absorption,the bioreactor can keep the short-chain fatty acids in the fermentation broth within the normal physiological concentration,so that the growth of microorganisms would not be inhibited.In terms of the sterility testing,long-term run bioreactor is not easy to be infected afterwards,ensuring the accuracy of the experiment.In terms of the control of p H,the bioreactor had a good ability to adjust the acid-base balance,maintain the environment in the large intestine,and ensure the normal growth of microorganisms.In terms of colonizing fecal microorganisms,the similarity rate of microbial communities was greater than 85.17%,which showed a relatively stable colonization effect.(3)The growth and metabolism of Akkermansia muciniphila in the biomimetic large intestine bioreactorBased on the developed biomimetic large intestine bioreactor,the brain heart infusion broth(BHI)medium,porcine mucin(PM)medium,human mucin(PM)medium,BHI +PM(BPM)medium and BHI+HM(BHM)medium were adopted for the dynamic fermentation culture of A.muciniphila,and for a comparison with the traditional static culture.It was found in this study that: in terms of biomass,the biomass of BHI dynamic culture was 1.92 g·L-1,which was 44.36% higher than that of static culture;while the biomass would further increase under the circumstances of HM dynamic culture,reaching 2.89 g·L-1.In terms of metabolites,PM and HM were applied for the dynamic cultivation.The main metabolites were short-chain fatty acids(acetic acids and butyric acids),while the other three media had produced a considerable amount of branched-chain fatty acids(isobutyric acids and isovaleric acids).In terms of appearance and morphology,under the circumstances of HM dynamic culture,the diameter of cells reached 999 nm,and the outer membrane protein concentration reached the highest value of 26.26 ?g·mg-1.The results showed that the nutrient composition and culture conditions of the medium can directly affect the biomass,outer membrane protein concentration and thickness and cell diameter of A.muciniphila in the biomimetic large intestine culture.(4)Research on the effect of different processing methods of highly resistant starch rice on in vitro digestion and intestinal floraThe high-resistant starch rice was selected as raw material and processed into rice,rice milk,rice cakes and popcorn through steaming,crushing,fermentation and high temperature and high-pressure treatment.The in vitro digestion and fecal microbial fermentation processes were analyzed respectively.It was found in this study that the starch digestibility of all those four foods in the stomach and small intestine reactors conformed to the first-order two-phase equation,and the resistant starch content in rice cakes was the highest(11.98%).In the process of biomimetic large intestine fermentation,undigested rice cakes were slower than inulin in the fermentation speed,and the concentration of butyric acid increased by 67.85%.The richness of Prevotellaceae that promoted the synthesis of short-chain fatty acids and Faecalibacterium,which had anti-inflammatory functions,increased.The richness of group imbalance markers Proteobacteria and Megamonas decreased.The results showed that the high-resistant starch rice can regulate the fermentation metabolites and ecological composition of the intestinal microbiota,which conduced to providing a reference for the design of functional foods for patients with diabetes and obesity.(5)Research on the impacts of dietary fatty acids on the distribution of intestinal gasBased on the developed biomimetic large intestine bioreactor,the intestinal gas array sensor was employed as a tool for the real-time monitoring of intestinal gas,the basic medium and dietary fatty acid medium were applied as the nutrient substrate,and the human fecal microorganisms were used as fermentation strains for fecal fermentation in vitro.The effects of basic nutrients and fatty acids on intestinal gas composition,concentration and volume were analyzed,and the effects of dietary fatty acids on the distribution of intestinal gas distribution kinetics.In this study,it was found that the gas components produced by fecal microbes using dietary fatty acids mainly included CO₂,H₂,H₂S and VOC,of which the content of CO₂ ranked first;fatty acids can regulate microbial fermentation to increase the concentration and content of H₂,H₂S and VOC.The results showed that dietary fatty acids can promote an increase in the concentration of H₂S and VOC in the intestine,which provided a reference for the increase in the onset of intestinal diseases caused by a high-fat diet,and provided dietary guidance for the reduction in the concentration of H₂S and VOC in the intestine.