The metabolic stability test of drugs is a key step in drug discovery and achieving low clearance is frequently the goal in the design of drug. Increased drug metabolism stability can reduce drug dosage, enhance drug exposure and prolong drug half-life. Accurately assessing the metabolic stability parameters of low clearance drugs and predicting human pharmacokinetics has become a challenge. Traditional tools in vitro including microsomes and suspended primary hepatocytes are limited by incubation time, which is not long enough to make sufficient metabolic conversion. Determination of intrinsic clearance or metabolic pathways and mechanisms of drug are implicated. Novel models tend to further mimic the in vivo environment in order to prolong lifetime of hepatocytes and achieve sufficient metabolic turnover of drugs for monitoring. In vitro-in vivo correlation of intrinsic clearance of methodologies has evaluated to support the reliability in predicting human pharmacokinetics. Application of these methodologies greatly decreases the forthputting of experimental animals and the release of expensive clinical trials during the acquisition of pharmacokinetic parameters. In this review, we summarized the principles, advantages and disadvantages of the novel in vitro methodologies for metabolic stability dealing with low-turnover drugs, including hepatocyte relay method, plated human hepatocytes, coculture system and microfluidic devices. Future prospect is proposed for in vitro metabolic models and it provides reference and optimization in metabolic stability for early lead compounds.