The growth thermogenic curves of Escherichia coli (E. coli) affected by berberine, coptisine and palmatine were determined quantitatively by microcalorimetry. The power-time curves of E. coli with and without the three berberines alkaloids (BA) were acquired, meanwhile the extent and duration of inhibitory effects on the metabolism were evaluated by growth rate constant (k), half-inhibitory ratio (IC50), peak time of maximum heat-output power (tp), total heat-production (Qt) and so on. The inhibitory effects of BA on E. coli revealed that the sequence of their antimicrobial activity was berberine > coptisine > palmatine. The functional groups methylenedioxy at C2 and C3 on phenyl ring improve antimicrobial activity more remarkably than methoxyl at C2 and C3 on phenyl ring. However, the antimicrobial activity does not vary significantly with methylenedioxy or methoxyl at C9 and C10 on phenyl ring.