BACKGROUND: Major royal jelly protein 1 (MRJP1) is a 55-57 kDa glycloprotein of royal jelly. Due to its several potential medical applications to human health, its production and purification is required for further studies. In this work, aqueous two-phase systems (ATPS) is proposed as an initial step to establish a practical strategy for the recovery of recombinant MRJP1 from Pichia pastoris fermentation culture. RESULTS: MRJP1 showed high affinity for top phase in PEG/phosphate systems when low MW PEG was used (PEG600 and PEG1000). It was recovered in the top phase of a PEG600/phosphate system with 45.2% w/w TLL from a cell-free supernatant with 95.8% recovery and 81% purity. In the same way, MRJP1 was concentrated in the top phase (83.6% recovery and80% purity) of a PEG1000/phosphate system with 47.2% w/w TLL when the whole fermentation broth, including cells, was processed. CONCLUSION: This study proved the potential of using ATPS for the primary recovery of recombinant MRJP1 from the fermentation culture of P. pastoris with cells. The results reported here represent the first step in a route to establish an ATPS-based process with integration of the initial separation steps. (C) 2014 Society of Chemical Industry