Basic Study
Copyright ©The Author(s) 2023. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Jun 7, 2023; 29(21): 3269-3279
Published online Jun 7, 2023. doi: 10.3748/wjg.v29.i21.3269
Effects of ethanol and sex on propionate metabolism evaluated via a faster 13C-propionate breath test in rats
Yosuke Sasaki, Naoyuki Kawagoe, Tsunehiko Imai, Yoshihisa Urita
Yosuke Sasaki, Naoyuki Kawagoe, Tsunehiko Imai, Yoshihisa Urita, Department of General Medicine and Emergency Care, Toho University School of Medicine, Tokyo 143-8541, Japan
Author contributions: Sasaki Y and Urita Y coordinated the study; Sasaki Y, Kawagoe N, Imai T, and Urita Y performed the experiments; Sasaki Y and Urita Y acquired and analyzed the data; Sasaki Y and Urita Y interpreted the data; and Sasaki Y and Urita Y wrote the manuscript; All authors approved the final version of the article.
Supported by the Japan Society for the Promotion of Science KAKENHI Grant, No. JP21K18089.
Institutional review board statement: The study was reviewed and approved by the Toho University School of Medicine Institutional Review Board, No. 21-51-496.
Conflict-of-interest statement: All the authors report no relevant conflicts of interest for this article.
Data sharing statement: Technical appendix, statistical code, and dataset available from the corresponding author at
ARRIVE guidelines statement: The authors have read the ARRIVE guidelines, and the manuscript was prepared and revised according to the ARRIVE guidelines.
Open-Access: This article is an open-access article that was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution NonCommercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See:
Corresponding author: Yosuke Sasaki, MD, PhD, Assistant Professor, Lecturer, Department of General Medicine and Emergency Care, Toho University School of Medicine, 6-11-1 Omori-Nishi, Ota-Ku, Tokyo 143-8541, Japan.
Received: February 2, 2023
Peer-review started: February 2, 2023
First decision: March 8, 2023
Revised: March 13, 2023
Accepted: May 6, 2023
Article in press: May 6, 2023
Published online: June 7, 2023

Alcoholism is regarded as a risk factor for vitamin B12 (VB12) deficiency. Because VB12 serves as a coenzyme of methylmalonyl-CoA mutase, a key enzyme in propionate metabolism, the 13C-propionate breath test (PBT) has been studied as a non-invasive diagnostic modality for VB12 deficiency. However, the conventional PBT requires 2 h, which is inconvenient in clinical practice. We hypothesized that a faster PBT can be used to evaluate propionate metabolism and is more easily adaptable for clinical practice.


To evaluate a faster PBT for assessing the effects of long-term ethanol consumption on propionate metabolism in ethanol-fed rats (ERs).


ERs were obtained by replacing standard drinking water (for control rats, CRs) with 16% ethanol solution in descendants of F344/DuCrj rats. Faster PBT was performed by administering 13C-propionate aqueous solution to male and female ERs and CRs by inserting a metal tubule from the mouth to the stomach; exhaled gas was collected in a bag to measure its 13CO2/12CO2 isotope ratio via infrared isotope spectrometry. Serum VB12 and alanine transaminase (ALT) levels were measured via chemiluminescence immunoassay and the lactate dehydrogenase-ultraviolet method, respectively. We evaluated statistical differences in mean body weight, change in 13CO213CO2‰), peak Δ13CO2‰, and serum VB12 and ALT, between males and females and between ERs and CRs using the t-test and Mann-Whitney U test for normally and non-normally distributed variables, respectively.


Males weighed significantly more than females (P < 0.001); CRs weighed significantly more than ERs (P < 0.008). Δ13CO2 reached a peak (Cmax) at 20 min and 30 min in females and males, respectively, decreasing after 20-30 min without rebound in all groups. Males had significantly higher Cmax and Δ13CO2 at 15-45 min than females (P < 0.05; for all pairs). Propionate metabolism was enhanced in male ERs relative to male CRs, whereas metabolism did not differ markedly between ERs and CRs for females. Males had higher serum VB12 levels than females, without prominent differences between the ER and CR groups. Male CRs had notably higher ALT levels than male ERs. Thus, chronic ethanol consumption may trigger fatty acid production via intestinal bacteria and changes in gut microbiome composition.


Faster PBT shows that 16% ethanol consumption promotes propionate metabolism without inducing liver injury. This PBT may be used clinically to evaluate gut flora status.

Keywords: Alcoholism, Breath test, Carbon isotope, Gut flora, Propionate, Vitamin B12

Core Tip: Alcoholism is a risk factor for vitamin B12 (VB12) deficiency. The 13C-propionate breath test (PBT) is a diagnostic modality for VB12 deficiency, but requires 2 h for completion. We applied a faster PBT to evaluate propionate metabolism using an ethanol-fed rat model. After 13C-propionate administration, the 13CO2/12CO2 isotope ratio of gas collected every 5 min for 60 min was measured using infrared isotope spectrometry. The Δ13CO2 peak occurred within 30 min. Ethanol-fed males showed marked propionate metabolism without associated liver injury. This study demonstrates the potential of the faster PBT to evaluate propionate metabolism under various clinical conditions.