Aims To examine the antioxidant activity of Bacterioruberin (Bctr)-rich extracts isolated from a hyperpigmented, genetically modified Haloferax volcanii strain (HVLON3) and to investigate the effect on cold-sensitive ram sperm cells. Methods and Results The strain HVLON3 produces higher Bctr amounts than most haloarchaea (220 +/- 13 mg g(-1) DW). HVLON3-Bctr extract has higher antioxidant activity than beta-carotene (threefold) as evaluated using 2,2 diphenyl-1-picrylhydrazyl combined with Electron Paramagnetic Resonance analysis (EC50 4.5 x 10(-5) mol l(-1) vs 13.9 x 10(-5) mol l(-1) respectively). Different concentrations of HVLON3-Bctr extracts were assayed on ram sperm after freezing/thawing and physiologically relevant parameters were examined. Extracts containing 7 and 20 mu mol l(-1) Bctr significantly improved cell viability (P < 0.0001), total and progressive motility (P < 0.0001) and sperm velocities (P = 0.0172 for curvilinear velocity VCL, P = 0.0268 for average path velocity VAP and P = 0.0181 for straight line velocity VSL) and did not affect other parameters evaluated. Conclusions HVLON3 is an excellent source of natural microbial C-50 carotenoids with applicability in Biotechnology, Biomedical and Veterinary fields. HVLON3 Bctr extract improves the quality of cryopreserved ram sperm cells and could be applied to increase insemination yields. Significance and Impact of the Study: This study provides an insight on the bioactive properties of a bioproduct derived from haloarchaea (carotenoids) which are so far underexploited.