An integral part of the intestinal microbiota is undoubtedly formed by lactic acid bacteria (LAB). Their presence in the digestive tract is essential for its proper functioning. During inflammatory bowel disease, such as ulcerative colitis (UC), LAB occurrence is reduced while sulfate-reducing bacteria (SRB) occur widely in the intestine resulting in an increase of their metabolite, hydrogen sulfide. Inhibitory concentration and mechanism of action of hydrogen sulfide on LAB are not fully known yet. The aim of this paper is to find the proper testing methods for evaluation of the interaction between hydrogen sulfide and lactic acid bacteria, including minimal inhibitory concentration (MIC) determination. Spectrophotometric, colorimetric, agar plate methods and combination of these methods were tested. Thousands of people from all over the world are affected by inflammatory bowel disease every year. The gained results could help to understand and improve the stability of intestinal microbiome, improve the treatment of bowel inflammation diseases, or prevent bowel disease altogether.