We have developed a convenient method for the detection of Lophophora williamsii using loop-mediated isothermal amplification (LAMP). We designed six species-specific primers for L. williamsii, including two loop primers. This L. williamsii-specific primer set was used for LAMP of total DNA extracted from L. williamsii and from L. diffusa. Real-time monitoring of LAMP was achieved by measuring turbidity due to the formation of magnesium pyrophosphate. Amplification occurred in samples mixed with total DNA from L. williamsii, but not in those mixed with total DNA from L. diffusa. We could also visually observe L. williamsii by adding fluorescent detection reagent to the reaction tube and exposing it to UV light. This new method amplified L. williamsii selectively and is expected to be applicable to the detection of peyote.