According to the European Monitoring Center for Drugs and Drug Addiction (EMCDDA), there were 179 different synthetic cannabinoids reported as of 2017. In the USA, 5F-MDMB-PINACA, or 5F-ADB, accounted for 28% of cannabinoid seizures 2016-2018. The synthetic cannabinoid, 5F-MDMB-PICA, is structurally similar to 5F-MDMB-PINACA with an indole group replacing the indazole. Limited data exist from in vivo or in vitro metabolic studies of these synthetic cannabinoids, so potential metabolites to identify use may be missed. The goals of this study were to (a) investigate 5F-MDMB-PICA and 5F-MDMB-PINACA in vitro metabolism utilizing human hepatocytes; (b) to verify in vitro metabolites by analyzing authentic case specimens; and (c) to identify the potency and efficacy of 5F-MDMB-PICA and 5F-MDMB-PINACA by examining activity at the CB1 receptor. Biotransformations found in this study included phase I transformations and phase II transformations. A total of 22 5F-MDMB-PICA metabolites (A1 to A22) were identified. From hepatocyte incubations and urine samples, 21 metabolites (B1 to B21) were identified with 3 compounds unique to urine specimens for 5F-MDMB-PINACA. Phase II glucuronides were identified in 5F-MDMB-PICA (n = 3) and 5F-MDMB-PINACA (n = 5). For both compounds, ester hydrolysis and ester hydrolysis in combination with oxidative defluorination were the most prevalent metabolites produced in vitro. Additionally, the conversion of ester hydrolysis with oxidative defluorination to pentanoic acid for the first time was identified for 5F-MDMB-PICA. Therefore, these metabolites would be potentially good biomarkers for screening urine of suspected intoxication of 5F-MDMB-PICA or 5F-MDMB-PINACA. Both 5F-MDMB-PICA and 5F-MDMB-PINACA were acting as full agonists at the CB1 receptor with higher efficacy and similar potency as JWH-018.