bIndole, a bacterial product of tryptophan degradation, has a variety of important applications in the pharmaceutical industry and is a biomarker in biological and clinical specimens. Yet, specific assays to quantitate indole are complex and require expensive equipment and a high level of training. Thus, indole in biological samples is often estimated using the simple and rapid Kovács assay, which nonspecifically detects a variety of commonly occurring indole analogs. We demonstrate here a sensitive, specific, and rapid method for measuring indole in complex biological samples using a specific reaction between unsubstituted indole and hydroxylamine. We compared the hydroxylamine-based indole assay (HIA) to the Kovács assay and confirmed that the two assays are capable of detecting microgram amounts of indole. However, the HIA is specific to indole and does not detect other naturally occurring indole analogs. We further demonstrated the utility of the HIA in measuring indole levels in clinically relevant biological materials, such as fecal samples and bacterial cultures. Mean and median fecal indole concentrations from 53 healthy adults were 2.59 mM and 2.73 mM, respectively, but varied widely (0.30 mM to 6.64 mM) among individuals. We also determined that enterotoxigenic Escherichia coli strain H10407 produces 3.3 ؎ 0.22 mM indole during a 24-h period in the presence of 5 mM tryptophan. The sensitive and specific HIA should be of value in a variety of settings, such as the evaluation of various clinical samples and the study of indole-producing bacterial species in the gut microbiota. Indole is widely distributed in the environment and is a component of diverse important compounds that occur in nature. In the pharmaceutical industry, synthesized indoles and their modified derivatives are popularly known for their medicinal properties. Indole analogs are significant components of a number of products, including vitamin supplements, dye, over-the-counter drugs, flavor enhancers, and perfumery. They are also used in the agricultural and plastics industries. Indole has been shown to play a role in regulating bacterial biofilm formation and virulence and influences diverse physiological processes, including host immune response (1-7).Indole is produced by about 85 bacterial species, including Gram-positive and Gram-negative bacteria, through the enzymatic degradation of tryptophan (8). Once produced, indole can be chemically modified within the same bacterial cell or taken up and modified by non-indole-producing bacteria. The most common naturally occurring indole analog is 3-methylindole (skatole), although other analogs, such as indoxyl sulfate and indole-3-propionic acid, can be found (9-11).Indole production by bacteria is an important phenotypic characteristic that has long been used to differentiate, identify, and diagnose enteric bacterial infections (12). Currently, the Kovács assay (13-17) is the most widely used method for detecting indole-producing bacteria. However, the key component, para-dimethylaminobe...

The trillions of microbes that make up the gut microbiome are an important contributor to health and disease. With respect to xenobiotics, particularly orally administered compounds, the gut microbiome interacts directly with drugs to break them down into metabolic products. In addition, microbial products such as bile acids interact with nuclear receptors on host drug-metabolizing enzyme machinery, thus indirectly influencing drug disposition and pharmacokinetics. Gut microbes also influence drugs that undergo enterohepatic recycling by reversing host enzyme metabolic processes and increasing exposure to toxic metabolites as exemplified by the chemotherapy agent irinotecan and non-steroidal anti-inflammatory drugs. Recent data with immune checkpoint inhibitors demonstrate the impact of the gut microbiome on drug pharmacodynamics. We summarize the clinical importance of gut microbe interaction with digoxin, irinotecan, immune checkpoint inhibitors, levodopa, and non-steroidal anti-inflammatory drugs. Understanding the complex interactions of the gut microbiome with xenobiotics is challenging; and highly sensitive methods such as untargeted metabolomics with molecular networking along with other in silico methods and animal and human in vivo studies will uncover mechanisms and pathways. Incorporating the contribution of the gut microbiome to drug disposition, pharmacokinetics, and pharmacodynamics is vital in this era of precision medicine.