A micro flow device and a method for generating a fluid with pH gradient are provided. The micro flow device includes a first and second substrates, an ion exchange membrane, and at least an electrode unit. The second substrate having a second flow path is disposed corresponding to the first substrate that has a first flow path. The ion exchange membrane is disposed between the first substrate and the second substrate to separate an electrolyte solution inside the first and second flow paths. The electrode unit includes at least two electrodes disposed in the first and second flow paths respectively. When the pair of electrodes is driven to electrolyze the electrolyte solution, the ion exchange membrane retards the mixing of an anode product and a cathode product produced by electrolyzing the electrolyte solution, such that a liquid having pH gradient is generated inside the first and second flow paths.