Cell / Cell culture products

Quantification of mucin in feces by fluorescence measurement

Fecal mucin assay kit

Background

Mucin is a type of glycoprotein and a major component of mucus such as saliva, tears, gastric juice, and intestinal juice. The basic structure is a glycoprotein containing a large amount of sugars with a molecular weight of 1 to 10 million. The core is a repeating peptide structure of 10 to 80 residues. N-acetylgalactosamine (GalNAc) at the reducing end is frequently linked via O-glycosidic bonds (see Figure 1).

It is a molecule in which sugar chains are attached to a peptide backbone in the form of branches, and the heterogeneity of the structure of the branched sugar chains creates diversity, which realizes various physiological functions. Among these, sugar chains that have a specific molecular recognition function and recognize proteins on the surface of viruses and bacteria are also known. Due to these properties, in the intestinal tract, it works to prevent viruses, pathogenic bacteria, and toxins derived from bacteria from crossing the intestinal wall and moving into the blood. ).

 

Figure 2 Intestinal barrier function by mucin

Intended use

• It is possible to measure the mucin content in feces by measuring the fluorescence intensity obtained by degrading O-glycans by β-elimination under alkaline conditions and simultaneously labeling the reducing ends of sugar chains with fluorescence.
• It can be used for the development of functional foods, research on intestinal flora, food and agriculture.

Kit Components

• Buffer A: 100 mL x 3
• Buffer B: 25 mL x 1 bottle
• Buffer C: 25 mL x 1 bottle
• Reagent A: 1.0 mL x 1 bottle
• Reagent B: 1.5 mL x 2 bottles
• Standard solution ( N -acetylgalactosamine: 250 μg/mL): 1.0 mL x 1 bottle
• Enzyme solution: 1.5 mL x 1 bottle

Required but not included in the kit

• Purified water
• 99.5% ethanol
• Fluorescent microplate reader and black plate
• Micro test tube (2 mL, 1.5 mL）
• Micro test tube (2 mL, 1.5 mL）

(When you measure with a light spectrophotometer, please prepare a microcell)

 

Ordering Information

Catalog Number	Product Name	Size	Datasheet
CSR-FFA-MU-K01	Fecal Mucin Assay Kit	1 Kit	Download

 Principle and method

Heat denatures glucosidase in feces to prevent mucin degradation.

Crude mucin is extracted.

Enzyme solutions are used to degrade dietary starch.

Purify the mucin by ethanol precipitation.

Reagent A is added to the purified mucin and heat-treated under alkaline conditions.
(Reagent B reacts with the sugar chain reducing end of mucin generated by alkali treatment and emits fluorescence.) *

After adding buffer C, fluorescence is measured (excitation: 336 nm, fluorescence: 383 nm).

*At this time, the mucin content (as N -acetylgalactosamine equivalent) can be quantified by simultaneously reacting with N -acetylgalactosamine as a standard.

Experimental example

Effect of polyphenol administration on the intestinal environment of rats fed a high-fat diet

Figure 1 Calibration curve for mucin determination	Fig. 2 Mucin content in feces

 

 Citations:

Kawakami, Shinpei, et al. "Intake of a mixture of sake cake and rice malt increases mucin levels and changes in intestinal microbiota in mice." Nutrients 12.2 (2020): 449. https://www.mdpi.com/2072-6643/12/2/449/pdf

Higashimura, Yasuki, et al. "Dietary intake of yacon roots (Smallanthus sonchifolius) affects gut microbiota and fecal mucin and prevents intestinal inflammation in mice." Journal of Clinical Biochemistry and Nutrition (2021): 20-203. https://www.jstage.jst.go.jp/article/jcbn/advpub/0/advpub_20-203/_pdf

Forgie, Andrew J., et al. "Phytochemical-induced mucin accumulation in the gastrointestinal lumen is independent of the microbiota." bioRxiv (2022): 2022-03. https://www.biorxiv.org/content/10.1101/2022.03.11.483917.full.pdf

Tian, Yuan, et al. "Early Life Short-Term Exposure to Polychlorinated Biphenyl 126 in Mice Leads to Metabolic Dysfunction and Microbiota Changes in Adulthood." International journal of molecular sciences 23.15 (2022): 8220. https://www.mdpi.com/1422-0067/23/15/8220/pdf

Horinouchi, Ayumu, et al. "Intestinal immunomodulatory activity of indigestible glucan in mice and its utilization by intestinal bacteria in vitro." Journal of Functional Foods 87 (2021): 104759. https://www.sciencedirect.com/science/article/pii/S1756464621004084

Takahashi, Masanobu, et al. "Improvement of psoriasis by alteration of the gut environment by oral administration of fucoidan from Cladosiphon okamuranus." Marine Drugs 18.3 (2020): 154. https://www.mdpi.com/1660-3397/18/3/154/pdf

Ishibashi, Riko, et al. "Isoliquiritigenin attenuates adipose tissue inflammation and metabolic syndrome by modifying gut bacteria composition in mice." Molecular Nutrition & Food Research 66.10 (2022): 2101119. https://onlinelibrary.wiley.com/doi/pdf/10.1002/mnfr.202101119

Nakashima, Takako, et al. "Novel gut microbiota modulator, which markedly increases Akkermansia muciniphila occupancy, ameliorates experimental colitis in rats." Digestive Diseases and Sciences (2021): 1-13.

Fujisaka, Shiho, et al. "Bofutsushosan improves gut barrier function with a bloom of Akkermansia muciniphila and improves glucose metabolism in mice with diet-induced obesity." Scientific Reports 10.1 (2020): 5544. https://www.nature.com/articles/s41598-020-62506-w

Matsumoto, Kenji, et al. "Resistant Starch-Supplemented Udon Noodles Prevent Impaired Glucose Tolerance and Induce Intestinal Immunoglobulin-A Secretion in Mice." Starch-Stärke 71.9-10 (2019): 1900042.

Tanaka, Miku, et al. "Comparison of the effects of roasted and boiled red kidney beans (Phaseolus vulgaris L.) on glucose/lipid metabolism and intestinal immunity in a high-fat diet-induced murine obesity model." Journal of food science 84.5 (2019): 1180-1187.

Matsumoto, Kenji, et al. "Comparison of the Effects of 3 Forms of Soluble Dietary Fiber on the Production of IgA in BALB/cAJcl and BALB/cAJcl-nu/nu Mice." The Journal of Nutrition 153.5 (2023): 1618-1626.

Niibo, M., et al. "Probiotic Lactobacillus gasseri SBT2055 improves insulin secretion in a diabetic rat model." Journal of dairy science 102.2 (2019): 997-1006.https://www.sciencedirect.com/science/article/pii/S002203021831083X

Kawashima, Rei, et al. "Histamine H2-receptor antagonists improve non-steroidal anti-inflammatory drug-induced intestinal dysbiosis." International journal of molecular sciences 21.21 (2020): 8166. https://www.mdpi.com/1422-0067/21/21/8166/pdf 

Shikano, Riho, et al. "Effects of proportions of carbohydrates and fats in diets on mucin concentration and bile composition in gallbladder of dogs." Journal of Veterinary Medical Science 84.11 (2022): 1465-1468. https://www.jstage.jst.go.jp/article/jvms/84/11/84_22-0126/_pdf

Tian, Yuan, et al. "Early-life exposure to a potent Aryl hydrocarbon receptor ligand results in persistent changes to the microbiota and host glucose homeostasis." (2023). https://www.researchsquare.com/article/rs-2781053/latest.pdf

Forgie, Andrew J., et al. "Pea polyphenolics and hydrolysis processing alter microbial community structure and early pathogen colonization in mice." The Journal of Nutritional Biochemistry 67 (2019): 101-110. https://drive.google.com/file/d/1q8uys9tGTS3GSNupntQOE0AsRxqi8SuC/view

Do, Moon Ho, et al. "Consumption of salt leads to ameliorate symptoms of metabolic disorder and change of gut microbiota." European Journal of Nutrition 59 (2020): 3779-3790.

Mezhibovsky, Esther, et al. "Impact of grape polyphenols on Akkermansia muciniphila and the gut barrier." AIMS microbiology 8.4 (2022): 544. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9834079/

Ansia, I., and J. K. Drackley. "Evaluation of 3 methods to determine mucin protein concentration in ileal digesta of young preweaning calves." Journal of dairy science 103.7 (2020): 6250-6257.https://www.sciencedirect.com/science/article/pii/S0022030220303246

Nakano, Hironobu, et al. "Effects of Amazake Produced with Different Aspergillus on Gut Barrier and Microbiota." Foods 12.13 (2023): 2568. https://www.mdpi.com/2304-8158/12/13/2568/pdf 

Nakamura, Akihisa, et al. "The potential of soluble CD14 in discriminating nonalcoholic steatohepatitis from nonalcoholic fatty liver disease." Hepatology Research 52.6 (2022): 508-521.

Watanabe, Yoshiyuki, et al. "Isoxanthohumol improves obesity and glucose metabolism via inhibiting intestinal lipid absorption with a bloom of Akkermansia muciniphila in mice." bioRxiv (2023): 2023-06. https://scholar.google.com/scholar?output=instlink&q=info:EDslOanu4ysJ:scholar.google.com/&hl=en&as_sdt=0,5&as_ylo=2019&as_yhi=2023&scillfp=11793773440774658088&oi=lle 

Kameyama, Akihiko, Risa Nishijima, and Kimi Yamakoshi. "Bmi-1 regulates mucin levels and mucin O-glycosylation in the submandibular gland of mice." Plos one 16.1 (2021): e0245607. https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0245607

Neto, Dionísio Pedro Amorim, et al. "Akkermansia muciniphila induces mitochondrial calcium overload and a-synuclein aggregation in an enteroendocrine cell line." Iscience 25.3 (2022). https://scholar.google.com/scholar?output=instlink&q=info:wFnEnB7sPC4J:scholar.google.com/&hl=en&as_sdt=0,5&as_ylo=2019&as_yhi=2023&scillfp=6773396684589809885&oi=lle 

Zhou, Xiaoliang, et al. "Aryl hydrocarbon receptor activation coordinates mouse small intestinal epithelial cell programming." Laboratory Investigation 103.2 (2023): 100012. https://www.laboratoryinvestigation.org/article/S0023-6837(22)00012-5/fulltext

Pedro, Dionisio, et al. "Akkermansia muciniphila induces mitochondrial calcium overload and a-synuclein aggregation in an enteroendocrine cell line." https://www.academia.edu/download/83193857/Paper_Akkermansia.pdf  

Forgie, Andrew. "Understanding the role of dietary phytochemicals and vitamin B12 in host-microbe interactions to support host gut integrity and health." (2022). https://era.library.ualberta.ca/items/6c1763f1-4746-4b1a-9d9e-2c85c79b7c34/download/1841a2c1-0960-40f8-99d8-e145d22a037c