Thiol metabolism is the primary detoxification strategy by which rice plants tolerate arsenic (As) stress. In light of this, it is important to understand the importance of harmonised thiol metabolism with As accumulation and tolerance in rice plant. For this aim, tolerant (T) and sensitive (S) genotypes were screened from 303 rice (Oryza sativa) genotypes on exposure to 10 and 25μM arsenite (As(III)) in hydroponic culture. On further As accumulation estimation, contrasting (13-fold difference) T (IC-340072) and S (IC-115730) genotypes were selected. This difference was further evaluated using biochemical and molecular approaches to understand involvement of thiolic metabolism vis-a-vis As accumulation in these two genotypes. Various phytochelatin (PC) species (PC(2), PC(3) and PC(4)) were detected in both the genotypes with a dominance of PC(3). However, PC concentrations were greater in the S genotype, and it was noticed that the total PC (PC(2)+PC(3)+PC(4))-to-As(III) molar ratio (PC-SH:As(III)) was greater in T (2.35 and 1.36in shoots and roots, respectively) than in the S genotype (0.90 and 0.15in shoots and roots, respectively). Expression analysis of several metal(loid) stress-related genes showed significant upregulation of glutaredoxin, sulphate transporter, and ascorbate peroxidase in the S genotype. Furthermore, enzyme activity of phytochelatin synthase and cysteine synthase was greater on As accumulation in the S compared with the T genotype. It was concluded that the T genotype synthesizes adequate thiols to detoxify metalloid load, whereas the S genotype synthesizes greater but inadequate levels of thiols to tolerate an exceedingly greater load of metalloids, as evidenced by thiol-to-metalloid molar ratios, and therefore shows a phytotoxicity response.