Gastrointestinal Digestion System Research Articles

Optimization of a green tea seed oil-loaded double-layer emulsion and its preventive effects on dextran sulfate sodium-induced colitis in mice.

Green tea seed oil (GTSO; Camellia sinensis) is rich in bioactive compounds and has great potential for preventing intestinal inflammation. Conversely, high-fat diets have been shown to promote or aggravate gastrointestinal inflammation, and the bioactive ingredients of GTSO face difficulty passing through the gastrointestinal tract while remaining intact. This study employed whey protein isolate (WPI) and sodium carboxymethyl cellulose (CMC-Na) to prepare a GTSO-loaded double-layer emulsion. The optimal parameters were as follows: WPI (2% w/w) and 35% oil phase for the primary emulsion preparation, CMC-Na (0.8% w/w) for the final emulsion, and 450W of ultrasonic power for homogenization. No significant changes in particle size or coalescence in the emulsions were observed after 30days of storage at 4°C. In addition, in a simulated gastrointestinal digestion system, more than 60% of the encapsulated GTSO was able to remain intact while passing through the gastric and small intestinal environment. In mice with dextran sulfate sodium-induced colitis, pretreatment with the GTSO emulsion significantly prevented the further development of colitis, whereas an empty-carrier-plus-free-GTSO treatment had no such protective effects and even tended to aggravate the disease. The results of the present study suggest that encapsulated GTSO is a reliable alternative approach for colitis prevention. PRACTICAL APPLICATION: The green tea seed oil-loaded double-layer emulsion demonstrates good storage stability when kept at 4°C, exhibits excellent slow-release performance, and achieves superior outcomes in the prevention of colitis. Our current study provides a reliable alternative approach for the prevention of colitis.

Intestinal-Targeted Digestion of Heme Chloride by Forming Inclusion Complexes In Vitro.

Hemin, a heme-like compound with significant biological activity, shows promise as an iron supplement for humans. Nonetheless, its poor solubility in water greatly impedes its absorption and utilization. To surmount this obstacle, researchers have chosen various cyclodextrins with distinct cavity sizes and derivative groups to act as hosts, forming inclusion complexes with hemin chloride. Among these, γ-cyclodextrin has been identified as the optimal carrier, based on a thorough evaluation of its encapsulation efficiency, solubility, and molecular docking. Multiple characterization techniques further confirmed the formation of these inclusion complexes. Results from IEC-6 cell experiments indicated that the cytotoxicity of the inclusion complexes was lower than that of FeSO4. Static and dynamic gastrointestinal simulation digestion systems were established, and the results showed that the bioavailability of the inclusion complex was significantly higher than that of raw hemin. Additionally, only about 0.29% of hemin chloride is digested by gastric enzymes, whereas 9.52% is digested by pancreatic enzymes in the static gastrointestinal simulation digestion system, with similar outcomes observed in the dynamic system. These findings suggest that targeted digestion in the intestine significantly enhances the bioavailability of hemin chloride by forming inclusion complexes in vitro.

Development of novel functional thickened drinks enriched with vitamin D3 for the older adult population – behaviour under dynamic in vitro digestion

The older adult population is greatly affected by malnutrition. This condition can arise from inappropriate micro and micronutrient intake, as well as due to experienced age-related changes. For instance, a significant number of older adults suffer from dysphagia. Thus, the development of foods with adequate consistency/viscosity are vital for overcome this condition. Within this framework, novel functional thickened drinks were developed and tailored for the older adult population, by using chia seed gum as the thickening agent enriched with vitamin D3-loaded solid lipid nanoparticles (SLN). Rheological properties of different thickened drinks formulated with water, semi-skimmed milk or orange juice were optimized using a central composite rotational design. Results showed that SLN delivered high vitamin D3 encapsulation efficiency with high concentration (i.e., 32 IU/mg). The different drink matrices attained distinct rheological properties, being observed that the viscosity and consistency attained with the same thickening agent concentration was higher in more complex matrices. An optimized formulation containing the tolerable upper intake level (ca. 4000 IU) was selected to study the vitamin D3 release and bioaccessibility on older adults’ population using a dynamic in vitro gastrointestinal digestion system. Once more, strong matrix effects were observed and significantly affected vitamin D3 release profile, bioaccessibility and recovery during the in vitro digestion. Additionally, the drinks enrichment with SLN-VitD3 did not display a negative effect on cells viability. The development of novel functional thickened drinks tailored for older adults could provide a safer swallowing and improved vitamin D3 intake.

Human Oral Phase Coupled with In Vitro Dynamic Gastrointestinal Digestion for Assessment of Plant Sterol Bioaccessibility from Wholemeal Rye Bread.

A dynamic gastrointestinal digestion system (simgi) after a human oral phase was used, for the first time, to assess the bioaccessibility of plant sterols (PS) from wholemeal rye bread (74.8 ± 2.2 mg of PS/100 g d.m.) and PS-enriched wholemeal rye bread (PS-WRB) (1.6 ± 0.04 g of PS/100 g of fresh bread). The use of these solid food matrices requires a novel adaptation of the gastric phase of the system. The PS identified in the breads are campesterol, campestanol, stigmasterol, β-sitosterol, sitostanol, Δ5-avenasterol, Δ5,24-stigmastadienol, Δ7-stigmastenol, and Δ7-avenasterol. The bioaccessibility of the total PS, only quantifiable in PS-WRB, is 19.9%, with Δ7-avenasterol being the most bioaccessible and Δ5-avenasterol being the least (p < 0.05). As shown in this study, PS-WRB can be considered to be a good choice to include in the daily diet. Furthermore, although the use of dynamic digestion methods for evaluating bioaccessibility implies high costs and technical complexity, their application means a closer approximation to in vivo scenarios.

Investigation of formation of AGEs precursors, hydroxymethylfurfural and malondialdehyde in oleogel added cakes using an in vitro simulated gastrointestinal digestive system

The baking process has the potential to generate health-risk compounds, including products from lipid oxidation and Maillard reaction. Pre- and post-digestion levels of hydroxymethylfurfural (HMF), malondialdehyde (MDA), glyoxal (GO), and methylglyoxal (MGO) were studied in cakes formulated with hazelnut and sunflower oil, along with their oleogels as margarine substitutes. The concentration of HMF in oil and oleogel-formulated cakes increased after digestion compared to cakes formulated with margarine. The MDA values were between 82 and 120 μg/100 g in oil and oleogel formulated cakes before digestion and a decrease was observed after digestion. The substitution of margarine with oil and oleogels resulted in the production of high amounts of GO and MGO in cakes. However, the highest bioaccessibility as 318.2% was found in cakes formulated by margarine for GO. Oleogels may not pose a potential health benefit compared to margarines due to the formation of HMF, MDA, GO, and MGO.

In vitro assessment of addition of soy protein isolate on milk protein digestion and conformational behaviour

Soy protein (SP) and milk protein are nutritionally complementary proteins, while they have different postprandial kinetic manifestations. In this study, different sources of proteins (casein, whey protein and SP isolate) were compounded in different concentrations and ratios. An in vitro dynamic gastrointestinal digestive system for children was used to study the digestive behaviour. For all the concentrations assessed, a consistently high level of small peptide release was demonstrated at 2.67 g/100 mL protein concentration, while the addition of 10% SP isolate would increase the release of small peptides. Overall, the study offers novel perspectives on children's products with diverse protein sources.

Dynamic digestion of a high protein beverage based on amaranth: Structural changes and antihypertensive activity

An amaranth beverage (AB) was subjected to a simulated process of dynamic gastrointestinal digestion DIDGI®, a simple two-compartment in vitro dynamic gastrointestinal digestion system. The structural changes caused to the proteins during digestion and the digesta inhibitory capacity of the angiotensin converting enzyme (ACE) were investigated. In gastric compartment the degree of hydrolysis (DH) was 14.7 ± 1.5 % and in the intestinal compartment, proteins were digests in a greater extent (DH = 60.6 ± 8.4 %). Protein aggregation was detected during the gastric phase. The final digesta obtained both at the gastric and intestinal level, showed ACE inhibitory capacity (IC50 80 ± 10 and 140 ± 20 μg/mL, respectively). Purified fractions from these digesta showed even greater inhibitory capacity, being eluted 2 (E2) the most active fraction (IC50 60 ± 10 μg/mL). Twenty-six peptide sequences were identified. Six of them, with potential antihypertensive capacity, belong to A. hypochondriacus, 3 agglutinins and 3 encrypted sequences in the 11S globulin. Results obtained provide new and useful information on peptides released from the digestion of an amaranth based beverage and its ACE bioactivity.

Advances in the delivery of bioactive compounds using starch-based nanogels: Formation mechanism, contemporary challenges, and prospects

Despite the widespread interest in starch-based nanogels due to their excellent swelling capacity and biocompatibility, there is a lack of in-depth analysis in the existing literature on the effects of their preparation methods, internal structure, and physicochemical properties on the release, retention, and protection of bioactive when used as delivery systems. In addition, there is a lack of systematic integration and discussion on the theoretical basis of how to design starch-based nanogels with specific structures and functions and the specific functions of these nanogels in oral-gastrointestinal digestive delivery systems. In this review, the preparation methods, characterization, and functions of starch-based nanogels are discussed in association with the delivery process to further understand the influence that the preparation method, as well as the structural and physicochemical characteristics have on the construction of the nanogel. This review aims to offer a theoretical foundation for designing starch-based nanogels with specific structures and functions. Summarizing functions of starch-based nanogels in an oral–gastrointestinal digestive delivery system can lay the foundation for the further development of starch-based nanogels as delivery systems. Furthermore, it can also deliver valuable insights and direct the development of healthy food items in an industrial setting.

Assessment of formation of the Glyoxal and Methylglyoxal Levels in Various Sugar-Free or Sweetened Products under in Vitro Gastrointestinal Digestive System

This study aimed to determine the formation of glyoxal (GO) and methylglyoxal (MGO) in various sweetener-containing products under in vitro digestion. Products obtained from different markets in Istanbul, Turkey. The amount of GO and MGO was determined by High-Performance Liquid Chromatography using pre-column derivatization. Initial GO and MGO values were ranged between 21.1-234.4 µg/100 g and 109.7-3164.5 µg/100 g for GO and MGO respectively. After digestion formation of GO and MGO were increased up to 2546% and 607.9%, respectively. Biscuit sample had the highest GO and MGO values, whereas, after digestion, the wafer exhibited the highest formation rate; both result may be related to high fat content, low moisture and, high heat treatment. Our results revealed that sweetener-containing products such as diabetic jams, chocolate, candy, had high GO and MGO content and this content may increase with digestion. Further studies examining the effects of different sugar types on GO and MGO formation are needed.

Calcium absorption by Alaska pollock surimi protein hydrolysate promotes osteoblast differentiation.

The calcium-binding capacity and osteoblast proliferation and differentiation were studied in Alaska pollock surimi hydrolysate (APSH) using a system that mimics the gastrointestinal digestive system. Evaluation of the calcium absorption-promoting ability of APSH revealed that the best calcium-binding ability was achieved after hydrolysis with a combination of pepsin, α-chymotrypsin, and trypsin, and separation into <3kDa (APSH-I), 3-5kDa (APSH-II), 5-10kDa (APSH-III), and <10kDa (APSH-IV) fractions. Scanning electron microscopy with energy-dispersive X-ray spectroscopy analysis confirmed that the hydrolysate and calcium ions formed a complex. Comparison of the calcium absorption capacity using Caco-2 cells showed that calcium absorption was promoted by these hydrolysates. Measurement of the osteoblast activation revealed higher alkaline phosphatase activity, collagen synthesis, and mineralization effect for the low-molecular-weight hydrolysate (LMH) than for the other hydrolysates. In addition, LMH promoted the expression of osteocalcin, osteopontin, and bone morphogenetic protein-2 and -4, which are hormones related to bone formation. Expression of the Runx2 transcription factor, which regulates the expression of these hormones, also increased. These results suggest that Alaska pollock surimi protein hydrolysates prepared using a system that mimics gastrointestinal hydrolysis may result in better osteoblast proliferation and bone health than those prepared using other proteases.

Chinese Yam and Its Active Components Regulate the Structure of Gut Microbiota and Indole-like Metabolites in Anaerobic Fermentation In Vitro.

As a medicinal and edible plant, Chinese yam (CY) can promote the enrichment of intestinal probiotics. Mucilage polysaccharides, diosgenin and taxifolin are the dominant components of CY. The purpose of this study was to investigate whether the impact of Chinese yam on gut microbiome structure and metabolism is attributable to its components. In the in vitro gastrointestinal digestion and colon fermentation system, the changes in gut microbiota composition and function were determined by 16S rRNA sequencing, and the levels of bacterial metabolites including short-chain fatty acids (SCFAs) and indole-like metabolites were detected by gas chromatography and an enzyme-linked immunoassay. The results show that CY, mucilage polysaccharides, diosgenin and taxifolin could increase the microbial diversity index. Furthermore, probiotics including Lactobacillus and Bacteroides were significantly increased, while harmful bacteria such as Escherichia and Proteus declined. CY could increase the production of SCFAs including acetic acid and butyric acid. Of note, CY and diosgenin displayed similar impacts on enhancing the abundance of Clostridium and promoting the production of indole-3-lactic acid and lactic acid. These findings provide evidence supporting Chinese yam as a natural food to regulate intestinal health. Diosgenin as a component of CY contributes mostly to the impact on regulating intestinal flora.

Developing garlic polysaccharide-Fe (III) complexes using garlic pomace to provide enhanced iron-supplementing activity in vivo

This study investigated the potential of garlic polysaccharides (GPs) from garlic pomace as iron carriers. The obtained GP-Fe (III) complexes had a higher molecular weight (5646 Da) and more fructose (90.46 %) than the GPs did and contained 9.7 % Fe (III). GPs were mainly composed of → 2)-β-d-Fruf (1 → and → 2)-β-d-Fruf (6 → residues, and their interactions with Fe (III) reduced the crystallinity, increased the thermal stability, and altered the morphological features through targeting the OH stretching vibrations of the hydroxyl groups and affecting the COC and OCO structures. The GP-Fe (III) complexes had high stability under simulated gastrointestinal digestion system and showed better therapeutic effects on iron deficiency anemia in mice than FeSO4 did, evidenced by improved hematological parameters, restored iron levels, and attenuated oxidative damage. Thus, GP-Fe (III) complexes are promising as novel Fe (III) supplements for Fe-deficient individuals, and promote the high-value utilization of garlic pomace.

Effects of six different microbial strains on polyphenol profiles, antioxidant activity, and bioaccessibility of blueberry pomace with solid-state fermentation.

To explore the effect of different microbial strains on blueberry pomace with solid-state fermentation (SSF), three fungi strains and three lactic acid bacteria (LAB) strains were utilized to investigate with respect to polyphenol profiles, antioxidant capacities, and bioaccessibility. Different strains exhibited different capacities for metabolizing polyphenolic compounds in blueberry pomace. The contents of 10 phenolic acids and 6 flavonoids (except (+)-catechin) were increased in blueberry pomace fermented by Lactobacillus acidophilus (LA). A similar tendency was observed in blueberry pomace fermented by Aspergillus niger (AN) and Lactobacillus plantarum (LP), where the concentration of 8 phenolic acids and 5 flavonoids was enhanced, with the following exceptions: (+)-catechin, ferulic acid, vanillic acid, and quercitrin. Chlorogenic acid and quercetin were the maximum phenolic acids and flavonoids in blueberry pomace with SSF, upgraded at 22.96 and 20.16%, respectively. Contrary to the growth of phenolic acids and flavonoid compounds, all individual anthocyanins showed a decreased trend. Only in the blueberry pomace fermented by AN, all anthocyanidins exhibit a rising trend. After SSF, 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), 2,2-diphenylpicrylhydrazyl (DPPH), and ferric reducing antioxidant power (FRAP) radical scavenging abilities were increased by up to 33.56, 59.89, and 87.82%, respectively. Moreover, the simulated gastrointestinal digestion system revealed that SSF improved the bioaccessibility of polyphenolic compounds. Compared with other strains, LA, LP, and AN showed better excellent capacities for metabolizing polyphenolic compounds, which led to a greater increase in antioxidant activity and bioaccessibility in fermented blueberry pomace.

DETERMINATION OF CONTENT, COOKING LOSS, AND BIOACCESSIBILITY OF THIAMINE IN RED MEATS USING IN VITRO SIMULATED HUMAN GASTROINTESTINAL DIGESTION SYSTEM

Meats are considered a valuable part of the human diet and a best source of B-group vitamins. Thus, investigations that measure the losses by cooking and bioaccessibilities of micronutrients are crucial to knowing the vitamin amounts in meats. The aim of this research was to investigate the content and loss of thiamine in meats by four different cooking practices, boiling, roasting, grilling, and frying, and to determine the bioaccessibility of thiamine in cooked meats by a simulated in vitro human gastrointestinal digestion system. High-performance liquid chromatography was used to detect the contents of thiamine in each raw and cooked meat. In raw meats, the measured thiamine amounts varied from 58.67 to 159 µg/100 g. After cooking, the amounts of thiamine ranged between 21 – 93 µg/100 g. While the lowest thiamine loss was determined in the boiled mutton thigh (8%), the highest loss was found in roasted veal thigh and mutton sleeve with 64%. The bioaccessibilities of thiamine in meats after digestion ranged from 12 to 83 µg/100 g. The highest thiamine bioaccessibility was in the grilled veal chop with 91%, while the lowest bioaccessibility was found in the roasted mutton sleeve with 12%. The bioaccessibility of vitamins may be remarkably influenced by factors such as stability, pH of the gastrointestinal tract, and temperature. Determining the bioaccessibility of thiamine in meats after cooking and digestion is important for the correct evaluation of the daily intake values ??of these vitamins.

Comparative analysis of fat digestion characteristics in sow milk and piglet formulas based on dynamic in vitro gastrointestinal digestion

Piglet formula is an important substitute for piglets when sow milk is unavailable. Triacylglycerols (TAGs) are the major components of sow milk and formulas. Lipolysis of TAGs in sow milk and five piglet formulas, mainly PF1 containing high similarity of sow milk fat substitute, PF2 containing coconut oil, and PF3-PF5 containing different amounts of 19.7%∼38.5% of medium chain fatty acids, was studied using in vitro dynamic gastrointestinal digestion system of piglets, and to elucidate the mechanism on digestive behaviors affected by different types of fats. The results showed that sow milk exhibited the largest particle size (4.22 μm) and the lowest absolute zeta-potential (12.8 mV) compared with formulas, contributing to its highest lipolysis degree (60.6%). It was followed by PF1 (56.2%), which had a similar fat composition to sow milk, especially the high content of sn-2 C16:0. Similarly, sow milk released the highest level of free fatty acids (56.9%, mainly long-chain species) after digestion, followed by PF1 (54.2%), and the other three formulas (33.8%–50.7%). The releases of diacylglycerides (e.g., L/L, P/L, P/O, O/O, and O/L) and monoacylglyceride-C16:0 were positively correlated with sn-2 C16:0 concentration, and the later will be re-esterified and reutilized for providing energy and essential fatty acids to piglets. It was indicated that the difference of fat digestive behaviors between sow milk and piglet formulas was associated to fat compositions and interfacial properties of their fat droplets. In this regard, digestion of fats for piglet formulas contributes to improving piglet health.

Acori Tatarinowii Rhizoma: A comprehensive review of its chemical composition, pharmacology, pharmacokinetics and toxicity.

Acori Tatarinowii Rhizoma (ATR, Shi Chang Pu in Chinese), a natural product with multiple targets in various diseases. This review provides the comprehensive summary of the chemical composition, pharmacological effects, pharmacokinetics parameters and toxicity of ATR. The results indicated that ATR possesses a wide spectrum of chemical composition, including volatile oil, terpenoids, organic acids, flavonoids, amino acids, lignin, carbohydrates and so on. Accumulating evidence from various studies has shown that ATR exerts a wide range of pharmacological properties, including protecting nerve cells, alleviating learning and memory impairment, anti-ischemic, anti-myocardial ischemia, anti-arrhythmic, anti-tumor, anti-bacterial, and anti-oxidant activities. Currently, ATR is widely used in the central nervous system, cardiovascular system, gastrointestinal digestive system, respiratory system in China, and for the treatment of epilepsy, depression, amnesia, consciousness, anxiety, insomnia, aphasia, tinnitus, cancers, dementia, stroke, skin diseases, and other complex diseases. Pharmacokinetic studies indicated that β-asarone, α-asarone, cis-methylisoeugenol, and asarylaldehyde, the active components of ATR, were absorbed slowly after oral administration of ATR. Moreover, toxicity studies have suggested that ATR has no carcinogenic, teratogenic and mutagenic toxicity. Nevertheless, long term or high-dose toxicity testing in animals to explore the acute and chronic toxicity of acori Tatarinowii Rhizoma is still lacking. In view of good pharmacological activities, ATR is expected to be a potential drug candidate for the treatment of Alzheimer's disease, depression, or ulcerative colitis. However, further studies are needed to elucidate its chemical composition, pharmacological effects, molecular mechanisms and targets, improve its oral bioavailability, and clarify its potential toxicity.

Bee pollen and bee bread nutritional potential: Chemical composition and macronutrient digestibility under in vitro gastrointestinal system

Bee pollen (BP) and bee bread (BB) have been often investigated as potential functional foods. Both bee products are generally characterized by their high nutritional content, with BB being referred as more digestible than BP, however, there is a lack of scientific studies proving this claim. Here, we present a comparative evaluation of the macronutrient digestibility of BP and BB after applying a simulated in vitro gastrointestinal digestive system, together with the evaluation of its nutritional value and chemical composition. The digestibility scores for protein content were calculated on average as 69% and 76% for BP and BB, respectively, whereas digestibility scores for soluble sugars varied depending on bee product and sugar type. The results demonstrated that the nutritional values of both bee products changed depending on their botanical origin but BB is more accessible in the intestinal lumen, especially regarding protein.

Development of a human milk protein concentrate from donor milk: Impact of the pasteurization method on static in vitro digestion in a preterm newborn model

The impact of high temperature short time (HTST, 72 °C, 15 s), Holder pasteurization- (63 °C, 30 min) and high hydrostatic pressure (HHP, 600 MPa-10 min) was evaluated on the digestibility of human milk protein concentrate (HMPC) by using a static in vitro gastrointestinal digestion system. The results showed that the processing steps used to produce the HMPC induced a decrease in readily available nitrogen (non-protein nitrogen and peptides). Overall, digestibility was similar between pasteurized and raw HMPC (degree of hydrolysis ranged from 26 to 34 %). Lactoferrin was more susceptible to gastric and intestinal digestion after thermal pasteurization. Additionally, the resistance of β-casein to digestion increased after HHP and Holder pasteurization due to aggregation and changes in protein structure. During intestinal digestion, Holder pasteurization induced a higher release of arginine, phenylalanine and tyrosine from HMPC compared to raw and HHP-treated HMPC. Overall, protein structural changes induced by human milk (HM) processing (freeze–thawing and filtration) and pasteurization treatments affected HMPC proteolysis during in vitro digestion. However, protein digestion behaviors were quite similar for raw and HHP-treated HMPC compared to the thermal-treated HMPC, with no effect on lactoferrin digestion. Consequently, pasteurization of HMPC by HHP represents an interesting non-thermal process that preserves the HM bioactive proteins during digestion.

The effect of in vitro simulated gastrointestinal digestive system on the biodegradation of B group vitamins in bread

Today, there is a growing interest in the consumption of whole grain products and the development of bread enriched with vitamins that have functional properties. Considerable losses arise in naturally found vitamins with food processing. Therefore, it is recommended to add vitamins to bread to obtain a satisfactory level. The aim of the current research was to investigate and assess the bioaccessibilities of the vitamins B1, B2, B3, and B6 in enriched commercial whole wheat breads by an in vitro digestion model. The average bioaccessibility of vitamin B1, B2, B3, and B6 in enriched breads after digestion was 80%, 64%, 79%, and 64%, respectively. After digestion, the bioaccessibilities of vitamins were affected. Mainly, vitamins B2 and B6 had the lowest bioaccessibility than vitamins B1 and B3. In vitro bioaccessibility was 70.9–90.2%, 54.2–89.7%, 42.1–94.9%, and 44.1–92.5% for vitamins B1, B2, B3, and B6, respectively in enriched commercial whole wheat bread. Vitamin B3 was seen with predominantly higher levels among the breads. Knowing the content of these vitamins in breads after digestion is necessary for the healthy nutrition of the population and for determining daily intake.

Identification of Gastrointestinal Digestion Stable Antihypertensive Fish Peptides from Atlantic Mackerel (Scomber scombrus).

High blood pressure has been recognized as one of the main risk factors of cardiometabolic syndrome by health organizations. Hypertension is medicated by various classes of synthetic drugs; however, adverse effects have repeatedly been reported. Moreover, natural alternatives such as fish peptides have been effective in the treatment and prevention of hypertension. The aim of our study was to fractionate and identify antihypertensive peptides. Fractions were produced using different techniques of solid-phase extraction (SPE), pressure-driven ultrafiltration (UF), and electro04dialysis with UF membrane. According to our results, the hydrophobic fraction of SPE (IC50 5 μg) was the most potent anti-angiotensin converting enzyme (ACE) product. Findings of the study suggest that the separation technique plays an important role in the isolation efficiency of antihypertensive biopeptides. Importantly, the hydrophobic fraction's activity was retained through a static model of an in vitro gastrointestinal digestion system. In conclusion, polarity regardless of charge and size was the most important factor for anti-ACE activity of an Atlantic mackerel biopeptide. In addition, the presence of leucine at either of the extremities (C- or N-terminal) and/or leucine-rich motifs could well explain the hypotensive effect of our active fraction.