Widespread antibiotic use has resulted in increased frequency of clinically important bacteria acquiring single or multiple antibiotic resistance.^{1, 2} Even antibiotic therapies for relatively trivial afflictions, such as acne,³ have promoted development of microbial antibiotic resistance. The availability of non-prescription antibiotics in some areas has also resulted in improper and/or irrational self-medication, further exacerbating this problem.^{4, 5} New antibacterial agents with broad-spectrum impact against both Gram-positive⁶ and Gram-negative⁷ species, as well as against drug-resistant strains such as methicillin-resistant Staphylococcus aureus⁸ are needed for wound care and to treat topical and dermatological infections.⁹ A chance observation in our laboratory revealed that creatinine (CRN; creatinine hydrochloride, CRN-HCl) halted the growth of bacteria on nutrient agar plates. CRN is the naturally occurring breakdown product of creatine phosphate, a high-energy molecule used to store and then donate, a high-energy phosphate to ADP for the synthesis of ATP in metabolism. Occurring normally in human blood at concentrations ranging approximately between 50–100 μM and in urine at slightly higher levels, CRN is accepted to be a naturally produced inert waste product with no active function,¹⁰ although a recently published study has challenged this dogma.¹¹ We characterized the ability of CRN-HCl to inhibit the growth of a wide array of bacterial species, including methicillin-resistant Staphylococcus aureus.

Drug-resistant bacterial strains and other bacterial species were assayed for sensitivity to inhibition of replication by CRN-HCl using a disc diffusion assay. Assays were performed as described¹² with the following modifications. Twenty-five microliters of 2 M CRN-HCl in water was added to 30 mg of a powdered carrier (Eridex; Cargill Inc., Cedar Rapids, IA, USA) and stirred into a thickened slurry in order to apply the maximum amount of CRN-HCl on the disc. Fifty microliters of the slurry containing 5 mg CRN-HCl were applied to 6 mm diameter sterile dry paper discs (Whatman no. 3 filter paper; Whatman, Piscataway, NJ, USA) that were then inverted onto LBG agar plates containing test bacteria. The LBG agar plates were prepared 15 min before use by spreading the test bacteria (diluted in phosphate-buffered saline to 1–5 × 10⁵ c.f.u. ml⁻¹ from overnight cultures) to the plate with a sterile cotton swab. The plates with discs were incubated for 15 h at the temperature appropriate for the particular bacterial species. Clear zones around the discs, indicative of growth inhibition, were measured. As a point of comparison, gentamicin-impregnated discs (GM-10; Becton Dickinson) were tested. For all bacteria but drug-resistant and anaerobic bacteria, for which assays were repeated twice, discs were tested in triplicate on two different days and zones of inhibition were recorded as the average of these six measurements. Variation was ⩽2 mm. Results for the diverse bacterial species assayed using this approach are shown in Table 1. All bacteria tested in this manner, including drug-resistant strains, were inhibited by CRN-HCl with a range of zone sizes between 16–40 mm.