Abstract The dependence of formal potential ( E 0′) on temperature for cytochrome c in phosphate buffer solution at various pH was investigated by the microband gold electrode modified with a new promoter 4,6-dimethyl-2-mercaptopyrimidine (DMMP). The temperature dependence of the E 0′ showed a biphasic character in slightly alkaline solution with a turning point at ca. 45 °C. However, the E 0′ values decreased monotonically with the increasing temperature in acidic and neutral media. The thermodynamic parameters of the electron transfer reaction for cytochrome c at the chemically modified microband electrode were estimated. The E 0′ and the changes of standard Gibb's free energy ( ΔG 0), entropy ( ΔS 0) and enthalpy ( ΔH 0) for the reduction of ferricytochrome c at 25 °C and pH 7.0 were 0.272 V (vs. NHE), −26.2 kJ mol −1, −123.7 J mol −1 K −1 and −63.1 kJ mol −1, respectively. The effect of buffer component on the E 0′ was also studied in binding (phosphate) and nonbinding (Tris/cacodylic acid) neutral buffer media. The results showed that the E 0′ in Tris/cacodylic acid buffer solution shifted positively by about 10 mV in comparison with that in phosphate buffer solution.