Diarrhea control using low-methoxyl pectin-containing enteral nutrition. 

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Makoto Tanaka, Ryosuke Akiyama, Ippei Yamaoka This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-5412851/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Background Diarrhea is the most common side effect of enteral nutrition and can lead to incontinence-associated dermatitis and malabsorption, indicating an unmet medical need for patients receiving enteral nutrition. Diarrhea related to enteral nutrition often occurs in conjunction with antibiotic use, highlighting the complexity of nutritional management and infection control. Here, we report an approach to address unmet medical needs by using low-methoxyl pectin-containing liquid diets, reconciling diarrhea control and antibiotic management. Methods Dysbiosis was induced in rats through a 4-week administration of an antibiotic cocktail added to drinking water. Different liquid diets, including fiber-free, low-methoxyl pectin, and partially hydrolyzed guar gum-containing formulas, were administered for 7 days using a gastric fistula. Fecal scoring and cecal histopathological examinations were conducted. Results The group administered a low-methoxyl pectin-containing liquid diet exhibited normal feces throughout the administration period that did not deteriorate with dysbiosis; additionally, mucosal atrophy was histopathologically suppressed. In contrast, fecal appearance deteriorated chronologically in the other liquid diet groups and worsened further with dysbiosis; frequent and strong villous atrophy was also observed. Conclusions A liquid diet containing low-methoxyl pectin controlled diarrhea in rats with antibiotic-treated dysbiosis. The mechanisms behind the improved fecal characteristics were attributed to the absorption of water and the inhibitory effect of dietary fiber on mucosal atrophy through stool formation. We hope to address these unmet medical needs through the use of low-methoxyl pectin in clinical settings in the future. Diarrhea control low-methoxyl pectin enteral nutrition nutritional management Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Background Gastrointestinal complications are the most frequently occurring (52%) complications in enteral nutrition (EN), with diarrhea being the most common side effect ( 1 ) . Experiencing loose stools is inconvenient for patients as well as nursing staff ( 2 ) and can lead to incontinence-associated dermatitis (IAD) ( 3 ) . It is important to provide a holistic approach to incontinence management, considering both the financial and psychosocial impacts on the patients ( 4 ) . Taking these factors into account, diarrhea management is an unmet medical need in EN. The complexities of EN are widely recognized and typically managed symptomatically; however, intrinsic issues for patients, such as malabsorption associated with diarrhea and energy loss in stools, have not received much attention. In a previous study, approximately half of patients with diarrhea had malabsorption during enteral feeding, indicating that greater awareness regarding managing diarrhea and nourishing patients with appropriate EN is required ( 5 ) . Although diarrhea management for EN is needed, there is a gap in the selection of commercial EN for this purpose. Prioritizing nutritional content sometimes worsens diarrhea, leading to intolerance to EN continuation. Because of this, biphasic features, appropriate nutritional content, and suppressive potential for diarrhea should be implemented in EN and selected for patients. External factors in EN formulations, such as osmolality, insoluble dietary fiber, and infusion rate, are non-specific factors contributing to diarrhea ( 6 ) . Suppliers have improved EN composition, adding external factors to address diarrhea. Choosing dextrin and peptides over monosaccharides and amino acids is a basic countermeasure to reduce EN osmolarity. Additionally, viscosity-variable EN, forming a gel of low-methoxyl (LM) pectin and calcium ions in the stomach at low pH, could be an alternative to control gastrointestinal transition ( 7 ) . With advancements in formulations, external factors have become controllable; despite this, EN-related diarrhea is triggered by internal factors such as altered physiological response, use of antibiotics/medication, and enteropathogenic infections ( 8 ) . Of these intrinsic factors, the use of medication is regarded as a relatively closer factor as most patients often not only need to be nourished by EN but also have multiple diseases requiring medication. For instance, antibiotics and proton pump inhibitors (PPIs) are used for infection control and as mucosal protective agents in 61% and 28% of patients receiving enteral feeding, respectively ( 9 ) ; however, their negative side effects on mucosal homeostasis are underrated. Clostridium difficile (CD) is a major cause of antibiotic-associated diarrhea and colitis. The incidence of CD infection is increasing in hospitals worldwide owing to the widespread use of broad-spectrum antibiotics. Unnecessary antibiotic use makes the treatment of CD-associated diseases more difficult and potentially less effective ( 10 ) . Similarly, PPIs prescribed routinely in the intensive care unit (ICU) due to the high incidence of stress-related mucosal damage appear to cause an alteration in the microbiota, which may also create a niche for CD colonization. The usual acidic environment of the stomach is fatal to CD, but once the pH rises to > 5, it can survive gastric exposure ( 10 – 12 ) . Negative effects are observed in patients using these medicines alongside EN. EN-related diarrhea is significantly correlated with antibiotic or PPI use ( 11 ) . Additionally, antibiotics are recommended to be withdrawn for diarrhea lasting more than 2 days in the algorithm for clinical management of EN-associated diarrhea ( 7 ) . This complexity reflects the dilemma between EN and antibiotic use, indicating the difficulty in maintaining both effective nutritional management and infection control. Patients with diarrhea and their relatives may feel that EN product suppliers are indifferent. Commercial EN is a homogenous product that is always formulated with an optimal nutritional composition; in contrast, individual EN products that cover patient medications are rare, meaning that profitability and interpreting unmet medical needs are always conflicting themes for suppliers. In this study, we report an approach to address these unmet medical needs, reconciling diarrhea control and antibiotic management during EN use in an antibiotic-induced dysbiosis rat model using an LM pectin-containing liquid diet. Methods Ethical approval All experimental protocols and animal experiments were reviewed and approved by the Animal Ethics Committee of Otsuka Pharmaceutical Factory, Inc. (OPFCAE-2023239). All animals were treated in accordance with committee guidelines. Animals and experimental protocols Forty-two male Sprague-Dawley rats (7–8 weeks old) were purchased from Jackson Laboratory, Japan, and housed in polycarbonate cages (3 rats/cage) with ALPHA-dry bedding and free access to a standard AIN-93G diet (Oriental Yeast Co., Ltd., Japan) and tap water under a 12-hour light-dark cycle. After five days acclimation, the rats were stratified in accordance with their body weight and randomly allocated to six groups (n = 7) using the same software as used for the statistical analysis. The rats in three groups underwent dysbiotic treatment before administering the liquid diet. Following the protocol for a germ-free mouse model ( 13) , the antibiotic cocktail was formulated with metronidazole (1 g/L), neomycin sulphate (1 g/L), vancomycin hydrochloride (0.5 g/L), and ampicillin sodium salt (1 g/L) (all purchased from Fuji Film Wako Pure Chemical Corporation, Japan) in distilled water. The cocktail was dosed for 4 weeks in freely accessed drinking water to induce dysbiosis. After 3 weeks of antibiotic treatment for dysbiosis groups, all animals were individually housed in metabolic cages, and liquid diets were administered for 1 week with three liquid diets for each of the groups based on the commercial liquid diet HINEX ® - EGel (Otsuka Pharmaceutical Factory, Inc., Japan): Fiber-free (FF) with LM pectin extracted, LM pectin-containing liquid diet (LM), and a partially hydrolysed guar gum (PHGG)-based liquid diet (GG) replacing LM pectin with PHGG. The dysbiosis groups were registered as FF-D, LM-D, and GG-D, respectively. Diet compositions are summarized in Table 1. All liquid diets were dosed using gastric tubes and syringe pumps at an infusion rate and caloric intake of 5.0 mL/h and 80 kcal/day (2.5 mL/h and 40 kcal/day for the first day of administration), respectively. In total, two animals were excluded due to the catheter-related troubles from FF and GG-D groups throughout the experimental period. Analytical methods The fecal score was calculated daily throughout the dosing period. Each fecal sample was registered as either 0 (normal), 1 (loose), 2 (muddy), or 3 (watery). Fecal scores were calculated using the following formula: (0 × number of score 0 stools + 1 × number of score 1 stools + 2 × number of score 2 stools + 3 × number of score 3 stools) / total number of stools. After the dosing period, animals were anaesthetized with isoflurane and euthanized after portal blood collection. Blood chemistry and plasma organic acid analyses were conducted using a biochemical autoanalyzer (JCA-BM6050; Nihon Denshi Co., Ltd., Japan) and gas chromatography-mass spectrometry (GCMS-QP2010Ultra; Shimadzu Corporation, Japan), respectively. The whole cecum and partial cecal contents/feces were weighed and then dried at 50 °C for 48 hours. The dried feces were homogenized, and the average pH was calculated using three values recorded using a pH meter (Eutech, Singapore). Ammonium concentration was measured by ion chromatography (Dionex Integrion HPLC; Thermo Fisher Scientific, Japan) using frozen samples stored at -80 °C. The concentrations of the fecal putrefaction products were measured using gas chromatography-mass spectrometry (5977A; Agilent Technologies, USA). Fecal water content was calculated using the weight of each sample. Ceca were collected from the animals, immersed, fixed in 10% neutral buffered formalin, embedded in paraffin blocks, and sectioned using standard procedures. All specimens were stained with hematoxylin and eosin (HE) and labelled with Ki-67 primary (rabbit monoclonal SP6, ab16667; Abcam, UK) and secondary (Histofine Simple Stain RAT MAX-PO (MULTI); Nichirei, Japan) antibodies. All specimens were examined histopathologically using a light microscope blindly by in house pathologist. Statistical analysis Data are presented as mean ± standard deviation, except for those of the histopathological examination. Fecal scores were analyzed using repeated measures analysis of variance (ANOVA). Other measurements were analyzed using one-way ANOVA, and then statistically analyzed using Tukey's multiple comparison test. Statistical significance was set at P < 0.05. All statistical analyses were performed using Stat Preclinica Ver 4.1 (Takumi Information Technology Inc., Japan). Results Representative images of the fecal samples are shown in Figure 1. In the FF and GG groups, fecal appearance deteriorated chronologically to muddy and watery stools. Antibiotic treatment in each group led to a shift in the stool appearance to being more unshaped and waterier. In contrast, normal stools were consistently observed in the LM group throughout the administration period following antibiotic treatment. The fecal scores are shown in Figure 1. Fecal scores in the FF and GG groups increased over time, which was consistent with the macroscopic images (Figure 2). In contrast, fecal scores remained consistently low in the LM group and significantly lower than those in the FF and GG groups from day 2 onwards. A comparison of fecal scores between each liquid diet group with or without antibiotic treatment showed low scores in all dysbiosis groups during the early phases of administration, likely due to impaction or decrease in the number of stools caused by antibiotic treatment (data not shown). The FF-D and GG-D groups exhibited higher fecal scores over time; a more synchronized transition was observed between the LM and LM-D groups, with no trend or minor differences observed on day 7. Plasma ammonia and urea nitrogen levels were lower in the LM group than in the FF group and were diminished by antibiotic treatment (Figure 3). Lower plasma levels of propionic acid and isovaleric acid were observed in the LM group; no significant differences were observed for the other organic acids, including butyric acid. All values tended to decrease after antibiotic treatment in each group (Figure 3). No statistically significant differences in water content corresponding to fecal scores were observed between the groups. A higher fecal pH was observed in the LM group, and antibiotic treatment led to a higher pH in all groups (Figure 4). Fecal ammonia, indole, and p-cresol levels were lower in the LM group than in the FF group. Dysbiosis treatment resulted in lower values in all groups for each putrefaction product (Figure 4). Histopathologically, villous atrophy was specifically observed in all groups treated with antibiotics and was frequently observed to be high in the FF-D and GG-D groups; Ki-67-labelled cells were scarce in crypts in these groups, corresponding to mucosal atrophy and suggesting low proliferative activity in the mucosal epithelium (Figure 5). Discussion It has been reported that unfermented and excreted pectin can hold excess water within the formed gel to generate normal feces in the lower intestine ( 14) .Additionally,antibiotic-associated diarrhea might be secondary to impaired colonic fermentation in otherwise predisposed subjects, resulting in the accumulation of luminal carbohydrates and/or decreased short-chain fatty acid (SCFA)-stimulated sodium and water absorption (15 ) . Taking these mechanisms and pathophysiology into account, LM pectin may solely affect water retention in affecting fecal appearance, although its effects may also be due to other mechanisms. The fecal scores deterioration in the FF-D group compared with that in the FF group (completely withdrawn from dietary fiber plus antibiotic treatment) suggests that an additional mechanism is in play. Mucosal atrophy, observed histopathologically in most groups (including GG-D), is considered another exacerbating factor for lower fecal scores, as it leads to decreased water absorption from the intestinal lumen and secondary deterioration of fecal characteristics. In the LM-D group, mucosal atrophy was suppressed with a proper background of fecal scores, indicating that stool formation was an accelerator of peristaltic movement and a counterfactor for disuse atrophy of the intestinal mucosa. Regarding the impacts of LM pectin on peristalsis, rats fed the liquid diet containing LM pectin showed similar contractile motility of the descending colon, compared with rats fed a solid chow diet (1 4) . The mucosa may come in physical contact with the opposing mucosa during contraction, generating shear stress, compression, and other forces. Although a large food bolus can aid in mucosal deformation as it passes through the intestine ( 16) , this does not explain the negative impact of mucosal atrophy on water absorption. Depletion of SCFAs is another key player and bacterial metabolite involved in mucosal homeostasis. The FF-D and GG-D groups showed mucosal atrophy along with a significant decrease in butyrate, serving as fuel in the distal bowel (1 7) .Overall, the mode of action of LM pectin in maintaining fecal characteristics in the disbiotic rat model was attributed to the water absorption by dietary fiber and its inhibitory effect on mucosal atrophy. In contrast, the deterioration of fecal characteristics in the FF-D and GG-D groups, which underwent dysbiosis treatment, was presumed to be the result of complex effects, including depletion of SCFAs and impaired peristaltic movement, ultimately leading to mucosal atrophy. Even without dysbiosis treatment, the advantages of LM pectin went beyond fecal characteristics; lower plasma ammonia and urea nitrogen levels and lower fecal ammonia, indole, and p-cresol levels were also observed compared with those in the FF group. Microbial interactions with dietary polysaccharides produce SCFAs, which are important energy and signaling molecules. It is becoming increasingly accepted that butyrate-producing bacteria and butyrate may be beneficial for human health (18 ) .However, when easily accessible carbohydrates become scarce, microbial activity shifts towards fermentation of dietary or mucosal proteins instead, resulting in the formation of potentially deleterious compounds, such as phenols, indoles, ammonia, and hydrogen sulphide (H 2 S) (19 ) .The series of changes observed in the FF group is likely attributable to the cessation of dietary fiber intake, leading to the autodigestion and fermentation of mucosal proteins. The benefits of dietary fiber supplementation are undisputed based on various studies. Providing fiber to patients in the ICU is a controversial but important issue. Previously, the risk of infusing fiber-containing formulas was overestimated, as evidence now supports that providing fiber may be a better nutrition therapeutic strategy ( 20) .The reduction of putrefaction products observed in this study presents evidence that clinicians indifferent to fiber supplementation in ICU patients should choose EN-containing dietary fiber/LM pectin. Water-soluble dietary fiber (PHGG) has a wide range of applications in clinical nutrition. Due to its low viscosity, it is widely used in EN products and has clinical benefits beyond its generic properties in intestinal immunity. In a previous study, PHGG reduced the incidence of diarrhea in septic patients receiving total EN and symptoms of irritable bowel syndrome ( 21) .PHGG can be considered the main player in dietary fiber production; however, the selection of LM pectin as a source of dietary fiber may be a wildcard. New insights into the effects of LM pectin on fecal characteristics would pave the way for developing new EN-containing LM pectin. Additionally, there was no difference between LM pectin and PHGG in terms of butyrate production, a key feature of PHGG ( 22) , when evaluated using batch fermentation systems used to simulate the unstudiable gut environment of free-living humans ( 21) . Conclusion Low fecal scores were observed in antibiotic-treated dysbiotic rats fed liquid diets containing LM pectin, demonstrating a reconciliation between diarrhea control and antibiotic management. We hope to address these unmet medical needs in patients receiving EN by using LM pectin in clinical settings. Abbreviations ANOVA analysis of variance CD Clostridium difficile EN enteral nutrition FF fiber-free GG PHGG-based based liquid diet HE hematoxylin and eosin IAD incontinence-associated dermatitis ICU intensive care unit LM low-methoxyl PHGG partially hydrolyzed guar gum PPIs proton pump inhibitors SCFA short-chain fatty acid Declarations Ethics approval and consent to participate All experimental protocols and animal experiments were reviewed and approved by the Animal Ethics Committee of Otsuka Pharmaceutical Factory, Inc. (OPFCAE-2023239). All animals were treated in accordance with committee guidelines. Consent for publications Not applicable Availability of data and materials All data analysed during this study are included in this published article. Competing interests The authors declare that they have no competing interests. Funding No funding. Author contributions MT: conceptualization, carrying out of the animal experiment, formal analysis, writing – original draft. RA: preparing test materials, writing – review & editing. IY: conceptualization, supervision. All authors contributed to the manuscript and approved the final version for submission. Acknowledgements We thank Oriental Yeast Co., Ltd; Nutrition and Pathology Laboratory Co., Ltd; TechnoSuruga Laboratory Co., Ltd; and Biopathology Institute Co., Ltd for their technical support. We also would like to thank Erika Mori for fine technical assistance and Editage (www.editage.jp) for English language editing. References Cataldi-Betcher EL, Seltzer MH, Slocum BA, Jones KW. Complications occurring during enteral nutrition support: a prospective study. JPEN J Parenter Enter Nutr. 1983;7:546–52. Scheppach W, Burghardt W, Bartram P, Kasper HE. Addition of dietary fiber to liquid formula diets: the pros and cons. JPEN J Parenter Enter Nutr. 1990;14:204–9. Gray M, Bliss DZ, Doughty DB, Ermer-Seltun J, Kennedy-Evans KL, Palmer MH. Incontinence-associated dermatitis: a consensus. 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Advancing human gut microbiota research by considering gut transit time. Gut. 2023;72:180–91. McClave SA, Omer E, Eisa M, Klosterbauer A, Lowen CC, Martindale RG. The importance of providing dietary fiber in medical and surgical critical care. Nutr Clin Pract. 2024;39:546–56. Slavin JL, Greenberg NA. Partially hydrolyzed guar gum: clinical nutrition uses. Nutrition. 2003;19:549–52. Velázquez M, Davies C, Marett R, Slavin JL, Feirtag JM. Effect of oligosaccharides and fiber substitutes on short-chain fatty acid production by human fecal microflora. Anaerobe. 2000;6:87–92. Tables Table 1 is available in the Supplementary Files section. Additional Declarations No competing interests reported. Supplementary Files GraphicalAbstract.pptx floatimage1.jpeg Table 1 Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-5412851","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":382443313,"identity":"bebf3b78-1c2a-4b52-8789-90e45b65a504","order_by":0,"name":"Makoto Tanaka","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABCklEQVRIiWNgGAWjYBAC+wMMDIwNDAyJDTwgbgWIYANiHtxaQNIgLcVALUD6DIME0VrqwVoY22Ba8AA29rMHP86oYcjt7zl8/MHHeTZ18jPSEj8wyNzBrYUnL1lywzGG3Bln2xIbZ25LkzC4kXZYgoHnGR6H5RhIPmBjyG04z2PYzLvtsISBRHoDUMth3Fr43xj/fPCPIXE+WMucwxLyM9Kbf+DVIpFjJrmxjSFxw9keoJYGoJtupB3Db4vEGzPLmX0SiRvPHEucOeNYmuSGM8/SLBLw+YU/x/hmzzebxHlnkg98+FBjwy/fnmZ842MP7hCDAgk0fmLPAUJaMMAP0rWMglEwCkbBsAUAAvpazPsjWNYAAAAASUVORK5CYII=","orcid":"","institution":"Otsuka (Japan)","correspondingAuthor":true,"prefix":"","firstName":"Makoto","middleName":"","lastName":"Tanaka","suffix":""},{"id":382443314,"identity":"5d8f3814-0e3f-48d2-b7b6-c70acc446bc0","order_by":1,"name":"Ryosuke Akiyama","email":"","orcid":"","institution":"Otsuka (Japan)","correspondingAuthor":false,"prefix":"","firstName":"Ryosuke","middleName":"","lastName":"Akiyama","suffix":""},{"id":382443315,"identity":"fe990797-2480-4f72-ae6e-c6f946f1966e","order_by":2,"name":"Ippei Yamaoka","email":"","orcid":"","institution":"Otsuka (Japan)","correspondingAuthor":false,"prefix":"","firstName":"Ippei","middleName":"","lastName":"Yamaoka","suffix":""}],"badges":[],"createdAt":"2024-11-08 01:53:19","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-5412851/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-5412851/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":71055460,"identity":"b1a8cebc-5aa0-4543-ab8a-ea413b7f3f5b","added_by":"auto","created_at":"2024-12-10 16:09:04","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":450411,"visible":true,"origin":"","legend":"\u003cp\u003eFecal scoring\u003c/p\u003e\n\u003cp\u003eAbbreviations: D1-D7, day 1-day 7 after dosing; FF, liquid diet without dietary fiber; GG, liquid diet with partially hydrolyzed guar gum; LM, liquid diet with low-methoxyl pectin; FF-D, GG-D, LL-D, dysbiosis treatment prior to each liquid diet administration.\u003c/p\u003e","description":"","filename":"fig1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-5412851/v1/7bd53c19bc23506c8da8ca4d.jpg"},{"id":71055470,"identity":"657bfaf2-dcf4-4198-88ab-fdca6ba2b38d","added_by":"auto","created_at":"2024-12-10 16:09:05","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":34788526,"visible":true,"origin":"","legend":"\u003cp\u003eRepresentative macroscopic images of the feces.\u003c/p\u003e\n\u003cp\u003eRepresentative fecal images on the final day of liquid diet administration. Deterioration of fecal characteristics from muddy to watery stools in the FF and GG groups as well as worsening of fecal characteristics during dysbiosis treatment. Normal fecal appearance in LM and LM-D groups. FF, liquid diet without dietary fiber; GG, liquid diet with partially hydrolyzed guar gum; LM, liquid diet with low-methoxyl pectin; FF-D, GG-D, LL-D, dysbiosis treatment prior to each liquid diet administration.\u003c/p\u003e","description":"","filename":"fig2.png","url":"https://assets-eu.researchsquare.com/files/rs-5412851/v1/9e9f22f47fe88b6d0fe43849.png"},{"id":71055461,"identity":"69d0f82b-e70a-48bb-bb8f-8f0cc7ead0af","added_by":"auto","created_at":"2024-12-10 16:09:04","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":284474,"visible":true,"origin":"","legend":"\u003cp\u003eBlood chemistry and plasma organic acids\u003c/p\u003e\n\u003cp\u003eAbbreviations: FF, liquid diet without dietary fiber; GG, liquid diet with partially hydrolyzed guar gum; LM, liquid diet with low-methoxyl pectin; FF-D, GG-D, LL-D, dysbiosis treatment prior to each liquid diet administration.\u003c/p\u003e","description":"","filename":"fig3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-5412851/v1/c9456377baf00b5b1e987aaf.jpg"},{"id":71056412,"identity":"b5dd98bc-ba29-46a3-99b4-cd6e4c132e12","added_by":"auto","created_at":"2024-12-10 16:17:04","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":281548,"visible":true,"origin":"","legend":"\u003cp\u003eWeight of cecal contents and water amounts. Fecal pH, ammonia, indol and p-cresol concentrations.\u003c/p\u003e\n\u003cp\u003eAbbreviations: FF, liquid diet without dietary fiber; GG, liquid diet with partially hydrolyzed guar gum; LM, liquid diet with low-methoxyl pectin; FF-D, GG-D, LL-D, dysbiosis treatment prior to each liquid diet administration.\u003c/p\u003e","description":"","filename":"fig4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-5412851/v1/a032f069939bccd391211dea.jpg"},{"id":71055466,"identity":"d46daa35-760a-4a20-bf48-0b00c694fa9d","added_by":"auto","created_at":"2024-12-10 16:09:04","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":54659873,"visible":true,"origin":"","legend":"\u003cp\u003eResult of the histopathological examination\u003c/p\u003e\n\u003cp\u003eResults of cecum histopathological examination. HE staining with low magnification (bar = 500 μm) and high magnification (bar = 100 μm). Ki-67 immunostaining (bar = 100 μm). Villous atrophy was observed frequently and was high in the FF-D and GG-D groups. Few Ki-67-positive cells were seen in crypts in the FF-D and GG-D group (arrowheads). HE, hematoxylin and eosin; FF-D, GG-D, dysbiosis treatment prior to each liquid diet administration.\u003c/p\u003e","description":"","filename":"fig5.png","url":"https://assets-eu.researchsquare.com/files/rs-5412851/v1/6246480beaf789d2627f27eb.png"},{"id":108601096,"identity":"5b4b739b-abb8-4f5f-804a-c8e7047be836","added_by":"auto","created_at":"2026-05-06 11:28:12","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":69621429,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-5412851/v1/b24e8735-8b03-4c56-932f-bf57afb653df.pdf"},{"id":71055464,"identity":"c257ebd5-3cae-42d7-8a32-d1df2aa74d21","added_by":"auto","created_at":"2024-12-10 16:09:04","extension":"pptx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":1096621,"visible":true,"origin":"","legend":"","description":"","filename":"GraphicalAbstract.pptx","url":"https://assets-eu.researchsquare.com/files/rs-5412851/v1/2862016e62d92e845ad735ac.pptx"},{"id":71056414,"identity":"191846b3-fe06-4797-aa85-14403a671784","added_by":"auto","created_at":"2024-12-10 16:17:04","extension":"jpeg","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":353864,"visible":true,"origin":"","legend":"\u003cp\u003eTable 1\u003c/p\u003e","description":"","filename":"floatimage1.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-5412851/v1/a743c89b02d48553e8d191fa.jpeg"}],"financialInterests":"No competing interests reported.","formattedTitle":"Diarrhea control using low-methoxyl pectin-containing enteral nutrition. ","fulltext":[{"header":"Background","content":"\u003cp\u003eGastrointestinal complications are the most frequently occurring (52%) complications in enteral nutrition (EN), with diarrhea being the most common side effect\u003csup\u003e(\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e)\u003c/sup\u003e. Experiencing loose stools is inconvenient for patients as well as nursing staff\u003csup\u003e(\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e)\u003c/sup\u003e and can lead to incontinence-associated dermatitis (IAD)\u003csup\u003e(\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e)\u003c/sup\u003e. It is important to provide a holistic approach to incontinence management, considering both the financial and psychosocial impacts on the patients\u003csup\u003e(\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e)\u003c/sup\u003e. Taking these factors into account, diarrhea management is an unmet medical need in EN. The complexities of EN are widely recognized and typically managed symptomatically; however, intrinsic issues for patients, such as malabsorption associated with diarrhea and energy loss in stools, have not received much attention. In a previous study, approximately half of patients with diarrhea had malabsorption during enteral feeding, indicating that greater awareness regarding managing diarrhea and nourishing patients with appropriate EN is required\u003csup\u003e(\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e)\u003c/sup\u003e. Although diarrhea management for EN is needed, there is a gap in the selection of commercial EN for this purpose. Prioritizing nutritional content sometimes worsens diarrhea, leading to intolerance to EN continuation. Because of this, biphasic features, appropriate nutritional content, and suppressive potential for diarrhea should be implemented in EN and selected for patients. External factors in EN formulations, such as osmolality, insoluble dietary fiber, and infusion rate, are non-specific factors contributing to diarrhea\u003csup\u003e(\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e)\u003c/sup\u003e. Suppliers have improved EN composition, adding external factors to address diarrhea. Choosing dextrin and peptides over monosaccharides and amino acids is a basic countermeasure to reduce EN osmolarity. Additionally, viscosity-variable EN, forming a gel of low-methoxyl (LM) pectin and calcium ions in the stomach at low pH, could be an alternative to control gastrointestinal transition\u003csup\u003e(\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e)\u003c/sup\u003e. With advancements in formulations, external factors have become controllable; despite this, EN-related diarrhea is triggered by internal factors such as altered physiological response, use of antibiotics/medication, and enteropathogenic infections\u003csup\u003e(\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e)\u003c/sup\u003e. Of these intrinsic factors, the use of medication is regarded as a relatively closer factor as most patients often not only need to be nourished by EN but also have multiple diseases requiring medication. For instance, antibiotics and proton pump inhibitors (PPIs) are used for infection control and as mucosal protective agents in 61% and 28% of patients receiving enteral feeding, respectively\u003csup\u003e(\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e)\u003c/sup\u003e; however, their negative side effects on mucosal homeostasis are underrated. \u003cem\u003eClostridium difficile\u003c/em\u003e (CD) is a major cause of antibiotic-associated diarrhea and colitis. The incidence of CD infection is increasing in hospitals worldwide owing to the widespread use of broad-spectrum antibiotics. Unnecessary antibiotic use makes the treatment of CD-associated diseases more difficult and potentially less effective\u003csup\u003e(\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e)\u003c/sup\u003e. Similarly, PPIs prescribed routinely in the intensive care unit (ICU) due to the high incidence of stress-related mucosal damage appear to cause an alteration in the microbiota, which may also create a niche for CD colonization. The usual acidic environment of the stomach is fatal to CD, but once the pH rises to \u0026gt;\u0026thinsp;5, it can survive gastric exposure\u003csup\u003e(\u003cspan additionalcitationids=\"CR11\" citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e)\u003c/sup\u003e. Negative effects are observed in patients using these medicines alongside EN. EN-related diarrhea is significantly correlated with antibiotic or PPI use\u003csup\u003e(\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e)\u003c/sup\u003e. Additionally, antibiotics are recommended to be withdrawn for diarrhea lasting more than 2 days in the algorithm for clinical management of EN-associated diarrhea\u003csup\u003e(\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e)\u003c/sup\u003e. This complexity reflects the dilemma between EN and antibiotic use, indicating the difficulty in maintaining both effective nutritional management and infection control. Patients with diarrhea and their relatives may feel that EN product suppliers are indifferent. Commercial EN is a homogenous product that is always formulated with an optimal nutritional composition; in contrast, individual EN products that cover patient medications are rare, meaning that profitability and interpreting unmet medical needs are always conflicting themes for suppliers. In this study, we report an approach to address these unmet medical needs, reconciling diarrhea control and antibiotic management during EN use in an antibiotic-induced dysbiosis rat model using an LM pectin-containing liquid diet.\u003c/p\u003e"},{"header":"Methods","content":"\u003cp\u003e\u003cem\u003eEthical approval\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eAll experimental protocols and animal experiments were reviewed and approved by the\u0026nbsp;Animal Ethics Committee of Otsuka Pharmaceutical Factory, Inc. (OPFCAE-2023239). All animals were treated in accordance with committee guidelines.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAnimals and experimental protocols\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;Forty-two male Sprague-Dawley rats (7\u0026ndash;8 weeks old) were purchased from Jackson Laboratory, Japan, and housed in polycarbonate cages (3 rats/cage) with ALPHA-dry bedding and free access to a standard AIN-93G diet (Oriental Yeast Co., Ltd., Japan) and tap water under a 12-hour light-dark cycle. After five days acclimation, the rats were stratified in accordance with their body weight and randomly allocated to six groups (n = 7) using the same software as used for the statistical analysis. The rats in three groups underwent dysbiotic treatment before administering the liquid diet. Following the protocol for a germ-free mouse model\u0026nbsp;\u003csup\u003e(\u003c/sup\u003e\u003csup\u003e13)\u003c/sup\u003e, the antibiotic cocktail was formulated with metronidazole (1\u0026nbsp;g/L), neomycin sulphate (1 g/L), vancomycin hydrochloride (0.5 g/L), and ampicillin sodium salt (1 g/L) (all purchased from Fuji Film Wako Pure Chemical Corporation, Japan) in distilled water. The cocktail was dosed for 4 weeks in freely accessed drinking water to induce dysbiosis. After 3 weeks of\u0026nbsp;antibiotic treatment for dysbiosis groups, all animals were individually housed in metabolic cages, and liquid diets were administered for 1 week with three liquid diets for each of the groups based on the commercial liquid diet HINEX\u003csup\u003e\u0026reg;\u003c/sup\u003e - EGel (Otsuka Pharmaceutical Factory, Inc., Japan): Fiber-free (FF) with LM pectin extracted, LM pectin-containing liquid diet (LM), and a partially hydrolysed guar gum (PHGG)-based liquid diet (GG) replacing LM pectin with PHGG. The dysbiosis groups were registered as FF-D, LM-D, and GG-D, respectively. Diet compositions are summarized in Table 1. All liquid diets were dosed using gastric tubes and syringe pumps at an infusion rate and caloric intake of 5.0 mL/h and 80 kcal/day (2.5 mL/h and 40 kcal/day for the first day of administration), respectively. In total, two animals were excluded due to the catheter-related troubles from FF and GG-D groups throughout the experimental period.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAnalytical methods\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eThe fecal score was calculated daily throughout the dosing period. Each fecal sample was registered as either 0 (normal), 1 (loose), 2 (muddy), or 3 (watery). Fecal scores were calculated using the\u0026nbsp;following formula: (0 \u0026times; number of score 0 stools + 1 \u0026times; number of score 1 stools + 2 \u0026times; number of score 2 stools + 3 \u0026times; number of score 3 stools) /\u0026nbsp;total number of stools. After the dosing period, animals were anaesthetized with isoflurane and euthanized after portal blood\u0026nbsp;collection. Blood chemistry and plasma organic acid analyses were conducted using\u0026nbsp;a biochemical autoanalyzer (JCA-BM6050; Nihon Denshi Co., Ltd., Japan) and gas chromatography-mass spectrometry (GCMS-QP2010Ultra; Shimadzu Corporation, Japan), respectively. The whole cecum and partial cecal contents/feces were weighed and then dried at 50 \u0026deg;C\u0026nbsp;for 48 hours. The dried feces were homogenized,\u0026nbsp;and the average pH was calculated using three values recorded\u0026nbsp;using a pH meter (Eutech, Singapore). Ammonium concentration was measured by ion chromatography (Dionex Integrion HPLC; Thermo Fisher Scientific, Japan) using frozen samples stored at -80\u0026nbsp;\u0026deg;C. The concentrations of the fecal putrefaction products were measured using gas chromatography-mass spectrometry (5977A; Agilent Technologies, USA). Fecal water content was calculated using\u0026nbsp;the weight of each sample. Ceca were collected from\u0026nbsp;the animals, immersed, fixed in 10% neutral buffered formalin, embedded in paraffin blocks,\u0026nbsp;and sectioned\u0026nbsp;using standard procedures. All specimens were stained with hematoxylin and eosin (HE) and labelled with Ki-67 primary (rabbit monoclonal SP6, ab16667; Abcam, UK) and secondary (Histofine Simple Stain RAT MAX-PO (MULTI); Nichirei, Japan) antibodies. All specimens were examined histopathologically using a light microscope blindly by in house pathologist.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eStatistical analysis\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eData are presented as mean \u0026plusmn; standard deviation, except for those of the histopathological examination. Fecal scores were analyzed using repeated measures analysis of variance (ANOVA). Other measurements were analyzed using one-way ANOVA, and then statistically analyzed using Tukey\u0026apos;s multiple comparison test. Statistical significance was set at P \u0026lt; 0.05. All statistical analyses were performed using Stat Preclinica Ver 4.1 (Takumi Information Technology Inc.,\u0026nbsp;Japan).\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003eRepresentative images of the fecal samples are shown in Figure 1. In the FF and GG groups, fecal appearance deteriorated chronologically to muddy and watery stools. Antibiotic treatment in each group led to a shift in the stool appearance to being more unshaped and waterier. In contrast, normal stools were consistently observed in the LM group throughout the administration period following antibiotic treatment. The fecal scores are shown in Figure 1. Fecal scores in the FF and GG groups increased over time, which was consistent with the macroscopic images (Figure 2). In contrast, fecal scores remained consistently low in the LM group and significantly lower than those in the FF and GG groups from day 2 onwards. A comparison of fecal scores between each liquid diet group with or without antibiotic treatment showed low scores in all dysbiosis groups during the early phases of administration, likely due to impaction or decrease in the number of stools caused by antibiotic treatment (data not shown). The FF-D and GG-D groups exhibited higher fecal scores over time; a more synchronized transition was observed between the LM and LM-D groups, with no trend or minor differences observed on day 7. Plasma ammonia and urea nitrogen levels were lower in the LM group than in the FF group and were diminished by antibiotic treatment (Figure 3). Lower plasma levels of propionic acid and isovaleric acid were observed in the LM group; no significant differences were observed for the other organic acids, including butyric acid. All values tended to decrease after antibiotic treatment in each group (Figure 3). No statistically significant differences in water content corresponding to fecal scores were observed between the groups. A higher fecal pH was observed in the LM group, and antibiotic treatment led to a higher pH in all groups (Figure 4). Fecal ammonia, indole, and p-cresol levels were lower in the LM group than in the FF group. Dysbiosis treatment resulted in lower values in all groups for each putrefaction product (Figure 4). Histopathologically, villous atrophy was specifically observed in all groups treated with antibiotics and was frequently observed to be high in the FF-D and GG-D groups; Ki-67-labelled cells were scarce in crypts in these groups, corresponding to mucosal atrophy and suggesting low proliferative activity in the mucosal epithelium (Figure 5).\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eIt has been reported that unfermented and excreted pectin can hold excess water within the formed gel to generate normal feces in the lower intestine\u003csup\u003e(\u003c/sup\u003e\u003csup\u003e14)\u003c/sup\u003e.Additionally,antibiotic-associated diarrhea might be secondary to impaired colonic fermentation in otherwise predisposed subjects, resulting in the accumulation of luminal carbohydrates and/or decreased short-chain fatty acid (SCFA)-stimulated sodium and water absorption\u003csup\u003e(15\u003c/sup\u003e\u003csup\u003e)\u003c/sup\u003e. Taking these mechanisms and pathophysiology into account, LM pectin may solely affect water retention in affecting fecal appearance, although its effects may also be due to other mechanisms. The fecal scores deterioration in the FF-D group compared with that in the FF group (completely withdrawn from dietary fiber plus antibiotic treatment) suggests that an additional mechanism is in play. Mucosal atrophy, observed histopathologically in most groups (including GG-D), is considered another exacerbating factor for lower fecal scores, as it leads to decreased water absorption from the intestinal lumen and secondary deterioration of fecal characteristics. In the LM-D group, mucosal atrophy was suppressed with a proper background of fecal scores, indicating that stool formation was an accelerator of peristaltic movement and a counterfactor for disuse atrophy of the intestinal mucosa. Regarding the impacts of LM pectin on peristalsis, rats fed the liquid diet containing LM pectin showed similar contractile motility of the descending colon, compared with rats fed a solid chow diet\u003csup\u003e(1\u003c/sup\u003e\u003csup\u003e4)\u003c/sup\u003e. The mucosa may come in physical contact with the opposing mucosa during contraction, generating shear stress, compression, and other forces. Although a large food bolus can aid in mucosal deformation as it passes through the intestine\u003csup\u003e(\u003c/sup\u003e\u003csup\u003e16)\u003c/sup\u003e, this does not explain the negative impact of mucosal atrophy on water absorption. Depletion of SCFAs is another key player and bacterial metabolite involved in mucosal homeostasis. The FF-D and GG-D groups showed mucosal atrophy along with a significant decrease in butyrate, serving as fuel in the distal bowel\u003csup\u003e(1\u003c/sup\u003e\u003csup\u003e7)\u003c/sup\u003e.Overall, the mode of action of LM pectin in maintaining fecal characteristics in the disbiotic rat model was attributed to the water absorption by dietary fiber and its inhibitory effect on mucosal atrophy. In contrast, the deterioration of fecal characteristics in the FF-D and GG-D groups, which underwent dysbiosis treatment, was presumed to be the result of complex effects, including depletion of SCFAs and impaired peristaltic movement, ultimately leading to mucosal atrophy. Even without dysbiosis treatment, the advantages of LM pectin went beyond fecal characteristics; lower plasma ammonia and urea nitrogen levels and lower fecal ammonia, indole, and p-cresol levels were also observed compared with those in the FF group. Microbial interactions with dietary polysaccharides produce SCFAs, which are important energy and signaling molecules. It is becoming increasingly accepted that butyrate-producing bacteria and butyrate may be beneficial for human health\u003csup\u003e(18\u003c/sup\u003e\u003csup\u003e)\u003c/sup\u003e.However, when easily accessible carbohydrates become scarce, microbial activity shifts towards fermentation of dietary or mucosal proteins instead, resulting in the formation of potentially deleterious compounds, such as phenols, indoles, ammonia, and hydrogen sulphide (H\u003csub\u003e2\u003c/sub\u003eS)\u003csup\u003e(19\u003c/sup\u003e\u003csup\u003e)\u003c/sup\u003e.The series of changes observed in the FF group is likely attributable to the cessation of dietary fiber intake, leading to the autodigestion and fermentation of mucosal proteins. The benefits of dietary fiber supplementation are undisputed based on various studies. Providing fiber to patients in the ICU is a controversial but important issue. Previously, the risk of infusing fiber-containing formulas was overestimated, as evidence now supports that providing fiber may be a better nutrition therapeutic strategy\u003csup\u003e(\u003c/sup\u003e\u003csup\u003e20)\u003c/sup\u003e.The reduction of putrefaction products observed in this study presents evidence that clinicians indifferent to fiber supplementation in ICU patients should choose EN-containing dietary fiber/LM pectin. Water-soluble dietary fiber (PHGG) has a wide range of applications in clinical nutrition. Due to its low viscosity, it is widely used in EN products and has clinical benefits beyond its generic properties in intestinal immunity. In a previous study, PHGG reduced the incidence of diarrhea in septic patients receiving total EN and symptoms of irritable bowel syndrome\u003csup\u003e(\u003c/sup\u003e\u003csup\u003e21)\u003c/sup\u003e.PHGG can be considered the main player in dietary fiber\u0026nbsp;production;\u0026nbsp;however, the selection of LM pectin as a source of dietary fiber may be a wildcard. New insights into the effects\u0026nbsp;of LM pectin on fecal characteristics\u0026nbsp;would pave\u0026nbsp;the way for developing new EN-containing LM pectin. Additionally,\u0026nbsp;there was no difference between LM pectin and PHGG in terms of butyrate production, a key feature of PHGG\u003csup\u003e(\u003c/sup\u003e\u003csup\u003e22)\u003c/sup\u003e, when evaluated using batch fermentation systems used to simulate the unstudiable gut environment of free-living humans\u003csup\u003e(\u003c/sup\u003e\u003csup\u003e21)\u003c/sup\u003e.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eLow fecal scores were observed in antibiotic-treated dysbiotic rats fed liquid diets containing LM pectin, demonstrating a reconciliation between diarrhea control and antibiotic management. We hope to address these unmet medical needs in patients receiving EN by using LM pectin in clinical settings.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003cdiv class=\"DefinitionList\"\u003e \u003cdiv class=\"DefinitionListEntry\"\u003e \u003cdiv class=\"Term\"\u003eANOVA\u003c/div\u003e \u003cdiv class=\"Description\"\u003e \u003cp\u003eanalysis of variance\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv class=\"DefinitionListEntry\"\u003e \u003cdiv class=\"Term\"\u003eCD\u003c/div\u003e \u003cdiv class=\"Description\"\u003e \u003cp\u003eClostridium difficile\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv class=\"DefinitionListEntry\"\u003e \u003cdiv class=\"Term\"\u003eEN\u003c/div\u003e \u003cdiv class=\"Description\"\u003e \u003cp\u003eenteral nutrition\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv class=\"DefinitionListEntry\"\u003e \u003cdiv class=\"Term\"\u003eFF\u003c/div\u003e \u003cdiv class=\"Description\"\u003e \u003cp\u003efiber-free\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv class=\"DefinitionListEntry\"\u003e \u003cdiv class=\"Term\"\u003eGG\u003c/div\u003e \u003cdiv class=\"Description\"\u003e \u003cp\u003ePHGG-based based liquid diet\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv class=\"DefinitionListEntry\"\u003e \u003cdiv class=\"Term\"\u003eHE\u003c/div\u003e \u003cdiv class=\"Description\"\u003e \u003cp\u003ehematoxylin and eosin\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv class=\"DefinitionListEntry\"\u003e \u003cdiv class=\"Term\"\u003eIAD\u003c/div\u003e \u003cdiv class=\"Description\"\u003e \u003cp\u003eincontinence-associated dermatitis\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv class=\"DefinitionListEntry\"\u003e \u003cdiv class=\"Term\"\u003eICU\u003c/div\u003e \u003cdiv class=\"Description\"\u003e \u003cp\u003eintensive care unit\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv class=\"DefinitionListEntry\"\u003e \u003cdiv class=\"Term\"\u003eLM\u003c/div\u003e \u003cdiv class=\"Description\"\u003e \u003cp\u003elow-methoxyl\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv class=\"DefinitionListEntry\"\u003e \u003cdiv class=\"Term\"\u003ePHGG\u003c/div\u003e \u003cdiv class=\"Description\"\u003e \u003cp\u003epartially hydrolyzed guar gum\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv class=\"DefinitionListEntry\"\u003e \u003cdiv class=\"Term\"\u003ePPIs\u003c/div\u003e \u003cdiv class=\"Description\"\u003e \u003cp\u003eproton pump inhibitors\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv class=\"DefinitionListEntry\"\u003e \u003cdiv class=\"Term\"\u003eSCFA\u003c/div\u003e \u003cdiv class=\"Description\"\u003e \u003cp\u003eshort-chain fatty acid\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003c/div\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll experimental protocols and animal experiments were reviewed and approved by the Animal Ethics Committee of Otsuka Pharmaceutical Factory, Inc. (OPFCAE-2023239). All animals were treated in accordance with committee guidelines.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publications\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and materials\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAll data analysed during this study are included in this published article.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eThe authors declare that they have no competing interests.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eNo funding.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eMT: conceptualization, carrying out of the animal experiment, formal analysis, writing \u0026ndash; original draft. RA: preparing test materials, writing \u0026ndash; review \u0026amp; editing. IY: conceptualization, supervision. All authors contributed to the manuscript and approved the final version for submission.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe thank Oriental Yeast Co., Ltd; Nutrition and Pathology Laboratory Co., Ltd; TechnoSuruga Laboratory Co., Ltd; and Biopathology Institute Co., Ltd for their technical support. We also would like to thank Erika Mori for fine technical assistance and Editage (www.editage.jp) for English language editing.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eCataldi-Betcher EL, Seltzer MH, Slocum BA, Jones KW. Complications occurring during enteral nutrition support: a prospective study. JPEN J Parenter Enter Nutr. 1983;7:546\u0026ndash;52.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eScheppach W, Burghardt W, Bartram P, Kasper HE. Addition of dietary fiber to liquid formula diets: the pros and cons. JPEN J Parenter Enter Nutr. 1990;14:204\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGray M, Bliss DZ, Doughty DB, Ermer-Seltun J, Kennedy-Evans KL, Palmer MH. Incontinence-associated dermatitis: a consensus. J Wound Ostomy Cont Nurs. 2007;34:45\u0026ndash;54.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBeele H, Smet S, Van Damme N, Beeckman D. Incontinence-associated dermatitis: pathogenesis, contributing factors, prevention and management options. Drugs Aging. 2018;35:1\u0026ndash;10.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003evan Schijndel RS, Wierdsma NJ, Van Heijningen EMB, Weijs PJ, de Groot SD, Girbes AR. Fecal energy losses in enterally fed intensive care patients: an explorative study using bomb calorimetry. Clin Nutr. 2006;25:758\u0026ndash;64.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePitta MR, Campos FM, Monteiro AG, Cunha AG, Porto JD, Gomes RR. Tutorial on diarrhea and enteral nutrition: a comprehensive step-by-step approach. JPEN J Parenter Enter Nutr. 2019;43:1008\u0026ndash;19.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eThakur BR, Singh RK, Handa AK, Rao MA. Chemistry and uses of pectin\u0026mdash;a review. Crit Rev Food Sci Nutr. 1997;37:47\u0026ndash;73.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWhelan K, Schneider SM. Mechanisms, prevention, and management of diarrhea in enteral nutrition. Curr Opin Gastroenterol. 2011;27:152\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHalmos EP, Muir JG, Barrett JS, Deng M, Shepherd SJ, Gibson PR. Diarrhea during enteral nutrition is predicted by the poorly absorbed short-chain carbohydrate (FODMAP) content of the formula. Aliment Pharmacol Ther. 2010;32:925\u0026ndash;33.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMarra AR, Edmond MB, Wenzel RP, Bearman GM. Hospital-acquired Clostridium difficile-associated disease in the intensive care unit setting: epidemiology, clinical course and outcome. BMC Infect Dis. 2007;7:1\u0026ndash;6.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ede Brito-Ashurst I, Preiser JC. Diarrhea in critically ill patients: the role of enteral feeding. JPEN J Parenter Enter Nutr. 2016;40:913\u0026ndash;23.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJump RL, Pultz MJ, Donskey CJ. Vegetative Clostridium difficile survives in room air on moist surfaces and in gastric contents with reduced acidity: a potential mechanism to explain the association between proton pump inhibitors and C. difficile-associated diarrhea? Antimicrob Agents Chemother. 2007;51:2883\u0026ndash;7.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eRakoff-Nahoum S, Paglino J, Eslami-Varzaneh F, Edberg S, Medzhitov R. Recognition of commensal microflora by toll-like receptors is required for intestinal homeostasis. Cell. 2004;118:229\u0026ndash;41.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHino K, Miyatake S, Yamada F, Endo N, Akiyama R, Ebisu G. Undigested low-methoxy pectin prevents diarrhea and induces colonic contraction during liquid-diet feeding in rats. Nutrition. 2020;78:110804.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eClausen MR, Bonn\u0026eacute;n H, Tvede M, Mortensen PB. 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Advancing human gut microbiota research by considering gut transit time. Gut. 2023;72:180\u0026ndash;91.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMcClave SA, Omer E, Eisa M, Klosterbauer A, Lowen CC, Martindale RG. The importance of providing dietary fiber in medical and surgical critical care. Nutr Clin Pract. 2024;39:546\u0026ndash;56.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSlavin JL, Greenberg NA. Partially hydrolyzed guar gum: clinical nutrition uses. Nutrition. 2003;19:549\u0026ndash;52.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eVel\u0026aacute;zquez M, Davies C, Marett R, Slavin JL, Feirtag JM. Effect of oligosaccharides and fiber substitutes on short-chain fatty acid production by human fecal microflora. Anaerobe. 2000;6:87\u0026ndash;92.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"},{"header":"Tables","content":"\u003cp\u003eTable 1 is available in the Supplementary Files section.\u003c/p\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Diarrhea control, low-methoxyl pectin, enteral nutrition, nutritional management","lastPublishedDoi":"10.21203/rs.3.rs-5412851/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-5412851/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eBackground\u003c/h2\u003e \u003cp\u003eDiarrhea is the most common side effect of enteral nutrition and can lead to incontinence-associated dermatitis and malabsorption, indicating an unmet medical need for patients receiving enteral nutrition. Diarrhea related to enteral nutrition often occurs in conjunction with antibiotic use, highlighting the complexity of nutritional management and infection control. Here, we report an approach to address unmet medical needs by using low-methoxyl pectin-containing liquid diets, reconciling diarrhea control and antibiotic management.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e \u003cp\u003eDysbiosis was induced in rats through a 4-week administration of an antibiotic cocktail added to drinking water. Different liquid diets, including fiber-free, low-methoxyl pectin, and partially hydrolyzed guar gum-containing formulas, were administered for 7 days using a gastric fistula. Fecal scoring and cecal histopathological examinations were conducted.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e \u003cp\u003eThe group administered a low-methoxyl pectin-containing liquid diet exhibited normal feces throughout the administration period that did not deteriorate with dysbiosis; additionally, mucosal atrophy was histopathologically suppressed. In contrast, fecal appearance deteriorated chronologically in the other liquid diet groups and worsened further with dysbiosis; frequent and strong villous atrophy was also observed.\u003c/p\u003e\u003ch2\u003eConclusions\u003c/h2\u003e \u003cp\u003eA liquid diet containing low-methoxyl pectin controlled diarrhea in rats with antibiotic-treated dysbiosis. The mechanisms behind the improved fecal characteristics were attributed to the absorption of water and the inhibitory effect of dietary fiber on mucosal atrophy through stool formation. We hope to address these unmet medical needs through the use of low-methoxyl pectin in clinical settings in the future.\u003c/p\u003e","manuscriptTitle":"Diarrhea control using low-methoxyl pectin-containing enteral nutrition. 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