Objective: To investigate potential associations of preoperative total testosterone (TT) with tumor volume (TV) and grade of prostate cancer (PCa). Methods: Patients who were under medications impacting on the hypothalamic-pituitary-adrenal-testis-prostate axis were excluded. TT was measured preoperatively at least 1 month after biopsies and TV was calculated on the removed prostate specimen. Other continuous variables included total prostate specific antigen (PSA), percentage of positive cores (P+) and weight (W) of the removed prostate. Patients were categorized according to the pathologic Gleason score (pGS) in 3 groups (pGS 6, 7 and > 7). Invasion of the seminal vesicles was coded as seminal vesicle invasion (SVI). Results: The median levels of TT were significantly and increasingly higher from pGS 6 (14.7 nmol/L) to pGS 7 (15.0 nmol/L) and pGS > 7 (18.8 nmol/L). The median values of TV were also detected significantly and increasingly higher from pGS 6 (5.6 mL) to pGS 7 (8.1 mL) and pGS > 7 (14.8 mL). The median preoperative levels of PSA were also increasing from pGS 6 (5.9 μg/L) to pGS 7 (6.2 μg/L) and pGS > 7 (7.7 μg/L). There was a significant and positive correlation of TV to PSA, TT and P+. Multiple linear regression analysis showed that TV was significantly and independently predicted by TT, PSA and P+. High grade PCa (pGS > 7) independently associated with TV, TT, P+ and SVI. The median density values of TT relative to TV (quotient TT/TV) significantly decreased from pGS 6 (2.6 nmol/L/mL) to pGS 7 (1.9 nmol/L/mL) and pGS > 7 (1.4 nmol/L/mL). The median density values of PSA relative to TV (quotient PSA/TV) also significantly decreased from pGS (1.1 μg/L/mL) to pGS 7 (0.7 μg/L/mL) and pGS > 7 (0.6 μg/L/mL). Conclusion: The investigation shows that TT relates to volume and grade of PCa; moreover, the density of TT relative to TV inversely associates with rate of increase of cancer that depends on the grade of the tumour.