Diabetes seriously endanger human health,and noninvasive glucose sensing is the expectation of both doctors and patients. Physiological background is complicated,volatile and mixed with a variety of tissue information,resulting in direct measurement of the body’s near infrared spectra difficult to truly reflect the concentration change in glucose. As a matter of fact,blood volume is always changing,but human tissue background and the concentration of blood components are constant in a short period. Taking advantage of this,subtracted blood volume spectrometry is propounded,which could eliminate the interference of human tissue background and obtain effective spectrum information of blood. To verify the effectiveness of the method,a experimental system was developed. The system noise is better than 20 μAU,and the signal to noise ratio of the effective spectrum signal at 1 250 nm is 20 000:1. Finally,the feasibility and advantages of subtracted blood volume spectrometry are clarified in clinical application of near infrared non-invasive glucose sensing.