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This study investigated the enzymatic production of bioactive peptides from hemp (Cannabis sativa L.) protein with potent pancreatic lipase (PL) inhibitory activity. Through a comparative evaluation of proteases, papain yielded a hydrolysate with the highest PL inhibitory activity and was consequently selected for process optimization using Box-Behnken response surface methodology. The optimal conditions were determined as follows: enzyme-to-substrate ratio, 4.3%; hydrolysis temperature, 62 ℃, and pH6.1, resulting in 57.06%±1.64% PL inhibition with an IC50 value of 5.12±0.31 mg/mL. The hydrolysate exhibited an irregular amorphous microstructure, accompanied by a reduced average molecular weight and an increased proportion of 1~3 kDa peptides. After enzymatic hydrolysis, the β-sheet proportion of the protein decreased significantly, while the α-helix and β-turn proportions increased remarkably. Additionally, the levels of amino acids such as glycine, alanine, proline, serine, and arginine underwent significant changes. A total of 1195 peptides were identified by LC-MS/MS, from which 15 candidate peptides (with sequences FEW, LLFSP, LAF, LLF, FYL, LSF, VLF, LLSF, LFL, LFY, LVF, LFR, LFF, LLLF, and FLF) were screened based on two criteria: relative content >1% and PeptideRanker score ≥0.6. The cumulative relative content of these 15 candidate peptides reached 28.79%. Molecular docking results indicated that the binding energies between these peptides and PL ranged from −7.4 to −10.0 kcal/mol, suggesting their potential as PL inhibitors. In conclusion, papain can efficiently generate PL-inhibitory peptides from hemp protein. Additionally, the enzymatic hydrolysis process significantly alters the structural characteristics of hemp protein. The optimized hydrolysis conditions established here, along with the identification of 15 potential bioactive peptides, provide a solid foundation for the high-value development of hemp protein and its industrial application in functional foods for the intervention of obesity.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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