Integrated deep eutectic solvent (DES) extraction and microwave-assisted process for algal protein extraction: Process improvement, characterizations, functional properties, and antioxidant and anti-colorectal cancer potentials

• DOI: 10.1016/j.jafr.2025.101673
• Publication Date: 2025-01-22T01:15:12Z
• Authors: May Thu Zin; Thida Kaewkod; Jeeraporn Pekkoh; Wasu Pathom-aree; Supakit Chaipoot; Gochakorn Kanthakat; Phisit Seesuriyachan; Yan-Yu Chen; Kuan Shiong Khoo; Benjamas Cheirsilp; Sirasit Srinuanpan
• Keywords: Deep eutectic solvent; Algae; Nutrition. Foods and food supply; Protein; Agriculture (General); TX341-641; Biomass; Microwave; S1-972

Microalgae and cyanobacteria are recognized as an alternative of traditional protein for their high protein content and nutritional benefits, but existing extraction methods often suffer from low efficiency, high energy consumption, and potential degradation of bioactive compounds This study aimed to develop an innovative method for extracting proteins from Chlorella and Spirulina biomass by combining deep eutectic solvent (DES) extraction with microwave-assisted processing, optimizing key extraction parameters for enhanced efficiency. Results found that the optimized extraction process, utilizing ChCl/Urea DES at 160 W microwave power, a biomass-to-solvent ratio of 1:40 for Spirulina and 1:30 for Chlorella, and a 10-min duration, achieved notable yields: 30.48 % for Spirulina and 15.53 % for Chlorella. The SDS-PAGE analysis demonstrated a diverse molecular weight range, with four distinct bands (10 kDa, 20–25 kDa, 50–75 kDa and >250 kDa) identified for Chlorella proteins and three bands (10 kDa, 50–75 kDa and >250 kDa) for Spirulina. The extracted proteins demonstrated balanced amino acid profiles, containing 39–40 % essential amino acids, comparable to high-quality plant proteins, and demonstrated strong functional properties, including foaming capacities of 96–100 % and emulsifying capacities of 98–100 %. Despite lower solubility due to isoelectric points, the proteins exhibited significant bioactivity, including DPPH scavenging activity ranging from 0.98 to 1.06 mg GAE/g extract, ABTS scavenging activity from 18.20 to 161.39 mg TEAC/g extract, and PFRAP activity from 2.90 to 3.45 mg GAE/g extract. More importantly, anticancer assays indicated substantial effects against colorectal cancer cells, with IC50 values of 8.60–9.28 mg/mL for Caco-2 cells and 14.16–15.99 mg/mL for HT-29 cells, while showing minimal toxicity toward normal cells. Flow cytometry further confirmed the pro-apoptotic effects of the extracted proteins. These findings highlight the potential of Spirulina and Chlorella proteins as sustainable, algal-based ingredients for functional foods and nutraceutical applications while addressing challenges in optimization, scalability, and biomass variability.