Mitochondrial DNA (mtDNA) mutations played crucial roles on affecting the susceptibility to cancer. In his study, to investigate whether mitochondrial DNA mutations contributed to the genetic susceptibilty of Chinese tongue cancer patients, mtDNA control regions of 105 Chinese tongue cancer patients were amplified and sequenced, the mutations were recorded by comparing with the revised Cambridge Reference Sequence (rCRS), which were attributed to certain mtDNA haplogroups based on he specific variations motif of each patients. The Miao Chinese group (a Chinese ethnic minority) from surrounding region has no essential difference with tongue cancer group, which was taken as the matched control group with principal component analysis by taking the haplogroups frequency of 105 tongue cancer individuals and 354 healthy individuals of eight groups from the similar geographic regions as input factors. This was supported by the smallest genetic distance between tongue cancer and Miao_2 groups. Further, the statistical analysis based on mtDNA variations of hypervariable sequence I (HVSI) indicated that 13 variations including 16,124, 16,148, 16,182C, 16,183C, 16,227, 16,266A, 16,249, 16,272, 16,291, 16,327, 16,335, 16,497, and 16,519 have significant differences between tongue cancer group and matched control group. Comparison of mtDNA haplogroups between tongue cancer and control groups indicated that mtDNA haplogroups C, F2*, and M10 have significant differences. It's worth noting that 16,327 and 16,291 was the defining variation of haplogroups C and F2*, respectively. Our results suggested that mitochondria' DNA may play a crucial role for the maternal genetic susceptibility of tongue cancer patients from Hunan, central of China.