Effects of ethanol and sex on propionate metabolism evaluated via a faster 13C-propionate breath test in rats

Abstract

BACKGROUND

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.

AIM

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

METHODS

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 ¹³CO₂ (Δ¹³CO₂‰), 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.

RESULTS

Males weighed significantly more than females (P < 0.001); CRs weighed significantly more than ERs (P < 0.008). Δ¹³CO₂ reached a peak (C_max) 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 C_max 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.

CONCLUSION

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.