Expanding the use of green solvents for the isolation of melittin from honeybee venom

The intrinsic complexity of natural products, which are made of different chemical constituents, poses significant challenges in their direct therapeutic use and their application as a source of new drug candidates, primarily due to the difficulty of isolating high-value bioactive compounds. One notable example of such complex matrix is bee venom, which contains a large variety of bioactive molecules, including melittin, alongside many allergenic compounds. The latter must be depleted to enable its safe clinical application. To this aim, purification methods need to be developed based on advanced preparative chromatographic approaches. However, one major drawback is the employment of toxic solvents, such as acetonitrile (ACN) and the generation of large amount of waste. In this context, this work aims to advance the green transition of melittin isolation through the development of more sustainable chromatographic methods able to recover as much product as possible, while simultaneously reducing the solvents consumption and enhancing the safety for operators. In this study, ACN is replaced with greener alternatives, including two alcohols, such as ethanol (EtOH) and isopropanol (IPA), as well as a mixture of dimethyl carbonate (DMC) and IPA in order to enhance process sustainability. The four methods were compared and evaluated based on four performance parameters: purity, recovery, productivity, and solvent consumption. To assess the greenness of each method, different green metrics were also employed. Most of them demonstrate that the calculated scores are much more favorable for DMC and alcohols, confirming their suitability as green alternative to ACN.