Pumpkin seed cake, a byproduct of cold-pressed oil production, represents a food waste material with a great potential for valorization. The objective of the present study is to optimize the papain enzymatic hydrolysis process of pumpkin seed cold-pressed oil cake (CPC) to obtain protein hydrolysates with the highest antioxidant activity. Box–Behnken Response Surface Methodology (RSM) was used to optimize the simultaneous effects of an enzyme concentration of papain, a temperature, and a reaction time on the process of enzymatic hydrolysis on pumpkin seed cold-pressed oil cake (CPC). For these three input factors, different values are used—1, 2, and 3% for papain concentration, 20, 30, and 40 °C for temperature, and 60, 120, and 180 min for hydrolysis time. Thus, the design generated a total of 21 experimental runs. The aim is to obtain protein hydrolysates with the highest antioxidant activity. The responses DPPH and ABTS were calculated and the determined regression models were statistically analyzed and validated. The results revealed that optimal conditions included a papain concentration of 1.0%, a temperature of 40 °C, and a hydrolysis time of 60 min to retrieve the highest level of bioactive compounds.

Hemp seed oil cake, a by-product of hemp seed oil extraction, is characterized by its high protein content and bioactive components, making it a valuable resource for the development of functional products through enzymatic hydrolysis. Hemp seed oil itself is renowned for its rich content of essential fatty acids, vitamins, and antioxidants, contributing to its widespread use in health and wellness products. Consequently, the residual cake presents significant potential for the food, pharmaceutical, and cosmetic industries as a source of high-quality protein ingredients. The optimization of enzymatic hydrolysis conditions is crucial for maximizing the efficiency and quality of the resulting protein hydrolysates. This study aims to optimize the hydrolysis process of hemp seed oil cake with bromelain, focusing on three key factors: enzyme concentration (E/S ratio), temperature, and time, to achieve hydrolysates with superior antioxidant activity. Response Surface Methodology (RSM) was applied using a Box–Behnken design to model and optimize the hydrolysis conditions. The experimental design involved three levels for each factor: 1%, 2%, and 3% for bromelain concentration; 20 °C, 30 °C, and 40 °C for temperature; and 60, 120, and 180 min for hydrolysis duration, resulting in 21 experimental runs. The antioxidant activity was assessed via DPPH and ABTS radical scavenging assays (%RSA), and the derived regression models were statistically analyzed and validated. The findings indicate that the optimal conditions for obtaining protein hydrolysates with the highest antioxidant activity are a bromelain concentration of 3.0%, a temperature of 40 °C, and a hydrolysis time of 60 min.

The use of non-traditional oilseeds for food purposes has increased significantly in recent years. Due to the high content of unsaturated fatty acids, the production of cold-pressed oils from pumpkin and hemp seeds is growing at a steady pace. This results in the accumulation of a by-product (oilseed press cake), from which high-quality proteins can be derived. Protein hydrolysates based on these raw materials offer further potential for valorization. This review summarizes classical and modern methods for obtaining protein isolates from pumpkin and industrial hemp oilseed cakes, as well as possible approaches for their modification to enhance their nutritional and functional properties. Furthermore, it provides an overview of the methods for producing protein hydrolysates rich in bioactive peptides. The presence of peptides with antioxidant, antihypertensive, or hypoglycemic properties makes hydrolysates a valuable source of nutraceutical components with potential applications in the development of functional foods or dietary supplements. However, most studies to date have been conducted using in vitro systems. Further research involving animal models or clinical observations is required to determine dosage regimens and applications of these hydrolysates. Regarding protein isolates, the challenges for their broader use in the food industry are related to increasing solubility, removing anti-nutritional factors, and improving techno-functional properties.

The potential of plant-based bioactive peptides to be used as an alternative or complementary therapy for the safe management of metabolic diseases such as diabetes is garnering increasing attention. In this in vivo study, the hypoglycemic effect of bromelain hydrolysates of hemp seed cake protein is analyzed in streptozotocin-induced diabetic mice. The results demonstrate that treatment of mice with the hydrolysate at a dose of 200 mg/kg/day for 4 weeks leads to a significant decrease in blood glucose levels (by 39%), serum triglyceride levels (16.44%), total cholesterol (13.50%), and low-density lipoprotein LDL (23.60%) compared to the untreated streptozotocin (STZ) group. The obtained values are comparable to those of the group treated with acarbose, which is used as a reference drug in the study. The applied hydrolysate significantly reduces serum alanine aminotransferase levels (by 39.38%) compared to the values of the untreated STZ group, while the change in superoxide dismutase levels was comparable to that of the group treated with acarbose. This study demonstrates the capacity of the applied hydrolysate to improve hyperglycemic conditions and reveals opportunities for future research into its potential as a functional ingredient.