To overcome the drawbacks of prism and concave gratings commonly used as dispersive devices in the traditional spectrophotometer, including low splitting efficiency, low resolution and little luminous flux, a high performance micro-spectrophotometer for biochemical analyzer (BCA) based on charge coupled devices (CCD) and virtual instrument technique is successfully developed, in which a plane grating is employed as the dispersive device, a linear CCD and a high speed peripheral component interconnect (PCI) bus data acquisition card are combined as the spectral detection system. Meanwhile, a custom-built Czerny-Turner (CT) typed monochrometer with a symmetric crossed structure is designed as the light splitting apparatus in this spectrophotometer. The experimental results demonstrate that this spectrophotometer realizes the capacity of detecting absorption intensity larger than 1000 (a.u.) in the ultra-violet and visible wavebands, the wavelength repeatability is less than 0.5 nm, the stray-light is less than 0.5% at the wavelength of 360 nm, its spectral accuracy is less than ±0.5 nm and its spectral measurement range and resolution reaches 300~800 nm and 1 nm, respectively. In addition, this system has the characteristics of post light-splitting and multi-channel parallel analysis, etc. This spectrophotometer for BCA has the potential value by virtue of optimized imaging system, high accuracy and low cost, etc.