BACKGROUND: A solid-liquid two-phase partitioning bioreactor (TPPB) was used in the biotransformation of indene to cis-(1S,2R)-indandiol by Pseudomonas putida 421-5 (ATCC 55687). Metered substrate feeding in single-phase operation, or delivery from an immiscible liquid, have previously been employed to regulate the exposure of the biocatalyst to inhibitory concentrations of the substrate. In contrast, the solid-liquid platform provided in situ substrate addition (ISSA) as well as simultaneous in situ product removal (ISPR) as a means of overcoming substrate and product toxicity. Three different modes of operation were compared for their effects on the performance of this biotransformation: single-phase, fed-batch operation was carried out as a benchmark in 2.75 L aqueous medium, and subsequently with the inclusion of either 700 g liquid silicone oil or 700 g solid polymer beads. RESULTS: Biphasic modes achieved a 3-fold productivity improvement with respect to single-phase (30 to 90mg L(-1) h(-1)), and solid-liquid productivity was similar to liquid-liquid operation while achieving more extensive removal of inhibitory compounds resulting in a slightly higher product titer (1.29 vs 1.16 g L(-1)). The operability of the reactor was improved by the phase stability of the solid polymer beads relative to immiscible organic solvents, preventing emulsion formation and facilitating analytics. CONCLUSION: Solid polymer beads replaced the immiscible liquid auxiliary phase for substrate delivery while performing simultaneous inhibitory molecule sequestration. (C) 2011 Society of Chemical Industry