Development of an environmental friendly and cost-effective process for microbial degradation of caffeine to non-toxic compounds are promising to solve the problems of physiochemical extraction of caffeine in the treatment the caffeine containing agro-industrial effluents. Thirteen bacterial strains, isolated from tea plantation soils in the north region of Iran, were screened to show their abilities in using caffeine as the sole source of carbon and nitrogen. The intrinsic tolerance of the isolated strains to the caffeine substrate was measured in a defined and complex medium by using the agar dilution method. Based on the tolerance efficiency, isolate TPS8 which showed maximum tolerance to caffeine was selected and identified as Pseudomonas pseudoalcaligenes strain TPS8 (GenBank accession number KF414528) according to the cultural and physiochemical characteristics and also 16S rDNA gene sequencing. Growing cells of P. pseudoalcaligenes TPS8 were used for the biodecaffeination experiments. The maximum removal of caffeine (80.2%) was reached after a 72 h incubation using 2.5 g/l of caffeine substrate without further optimization. Our results show that growing cells of P. pseudoalcaligenease TPS8 can thus be efficiently used as a simple and cheap process for preparative decaffeination from agro-industrial effluents. The present survey is the first report on biodecaffeination using Pseudomonas pseudoalcaligenes.