Figure 2 - uploaded by Jose Angel Gómez Ruiz
Content may be subject to copyright.
Source publication
Adapted, follow-up, probiotic follow-up, toddler, and probiotic toddler infant formulas were subjected to an in vitro enzymatic procedure simulating physiological digestion. The formation and identification of casein phosphopeptides (CPPs) in the milk-based infant formulas were studied using reversed phase high-performance liquid chromatography cou...
Contexts in source publication
Context 1
... Formula. The chromatographic profile of peptides released after simulated digestion of the adapted infant formula is reported in Figure 2A. A total of seven CPPs, two of them with the cluster sequence SpSpSpEE and with 2-5 phosphate groups, are identified ( Table 1). ...
Citations
... This activity is due to the presence of the amino acid phosphorylated serine, which can make salts with minerals, such as calcium. Enzymatic digestion of milk produces a diverse group of these peptides [207]. The type of amino acid composition present in the phosphorylated region plays an essential role in the amount of calcium-binding activity in this group of peptides [208]. ...
Bioactive peptides are a group of biological molecules that are normally buried in the structure of parent proteins and become active after the cleavage of the proteins. Another group of peptides is actively produced and found in many microorganisms and the body of organisms. Today , many groups of bioactive peptides have been marketed chemically or recombinantly. This article reviews the various production methods and sources of these important/ubiquitous and useful biomolecules. Their applications, such as antimicrobial, antihypertensive, antioxidant activities, blood-lipid-lowering effect, opioid role, antiobesity, ability to bind minerals, antidiabetic, and anti-aging effects, will be explored. The types of pathways proposed for bioactive applications will be in the next part of the article, and at the end, the future perspectives of bioactive peptides will be reviewed. Reading this article is recommended for researchers interested in various fields of physiology , microbiology, biochemistry, and nanotechnology and food industry professionals.
... In Ca-binding peptides, both the oxygen atoms of the carboxyl group and the nitrogen atom of the amino group seem to be involved in the formation of chelate with the mineral. Ca-binding capacity has been largely studied in casein phosphopeptides (CPPs), which are phosphorylated CN-derived peptides that can be released by in vitro or in vivo enzymatic hydrolysis of as1, as2, and b-CN (Miquel, Gomez, et al. 2005). Most CPPs contain the cluster SpSpSpEE, a highly polar acidic sequence of 3 phosphoseryl groups followed by 2 Glu residues, which are a binding site for minerals such as Ca, Fe, and Zn, and play an important role in mineral bioavailability Miquel et al. 2006). ...
Bioactive peptides derived from food protein sources have been widely studied in the last years, and scientific researchers have been proving their role in human health, beyond their nutritional value. Several bioactivities have been attributed to these peptides, such as immunomodulatory, antimicrobial, antioxidant, antihypertensive, and opioid. Among them, metal-binding capacity has gained prominence. Mineral chelating peptides have shown potential to be applied in food products so as to decrease mineral deficiencies since peptide-metal complexes could enhance their bioavailability. Furthermore, many studies have been investigating their potential to decrease the Fe pro-oxidant effect by forming a stable structure with the metal and avoiding its interaction with other food constituents. These complexes can be formed during gastrointestinal digestion or can be synthesized prior to intake, with the aim to protect the mineral through the gastrointestinal tract. This review addresses: (i) the amino acid residues for metal-binding peptides and their main protein sources, (ii) peptide-metal complexation prior to or during gastrointestinal digestion, (iii) the function of metal (especially Fe, Ca, and Zn)-binding peptides on the metal bioavailability and (iv) their reactivity and possible pro-oxidant and side effects.
... Nevertheless, the highly disordered N-terminal region of β-casein is characterized by a cluster of five phosphorylated serine residues (Fig. 8) and proteins carrying phosphorylated serine residues are known to resist proteolysis in the course of in vivo and in vitro gastrointestinal digestion. 50,51 This resistance to proteases can be explained by the supramolecular organisation of β-casein involving its phosphorylated and negatively charged N-terminal region and bridges formed in the presence of calcium and sodium ions. These interactions promote the association of β-casein molecules in small aggregates or micelles, [52][53][54] which results in the low accessibility of the N-terminal end to proteases. ...
Rabbit gastric extracts (RGE) is a source of gastric enzymes for in vitro digestion studies. While its gastric lipase activity has been characterized and compared to other lipases, its pepsin activity has not been studied. We measured pepsin activity in RGE using both hemoglobin and azocoll a substrates, and identified the protein separated by SDS-PAGE as a type II-4 mature pepsin of 328 amino acid residues using Edman sequencing, LC-MS/MS analysis and intact mass measurement. As a proof of concept that RGE was suitable for in vitro digestion of both proteins and lipids, it was used for studying the proteolysis of β-casein under conditions mimicking the early stages of intragastric digestion. β-casein was presented either in solution or at the surface of a β-casein-stabilized rapeseed oil emulsion to investigate the impact of lipids and lipolysis on proteolysis. Proteolysis of β-casein was quantified based on the kinetics of β-casein disappearance, the identification of various peptides generated upon digestion and their variation with time. The results obtained with RGE were highly similar to those obtained with equivalent amounts of porcine pepsin used a reference standard. Digestion of β-casein was slower when it was presented at the oil-water interface and some degradation peptides were transiently observed at higher levels and for a longer time than with β-casein in solution, or accumulated upon digestion. N-terminal sequencing of the main isolated peptides revealed a sequential action of pepsin starting from the hydrophobic C-terminal end of β-casein and which was impaired by β-casein interaction with lipids.
... In contrast, some ACE-inhibitory peptides exhibit high activity in vitro but have no effects in vivo. For example, the peptide FFWAP derived from αs1-CN [49][50][51] is a potent ACE inhibitor in vitro but has no hypotensive effect in vivo [15]. Generally, the difficulty of establishing a direct relationship between in vitro and in vivo activity may depend on different reasons but it is clear that bioavailability after oral administration plays a key role. ...
Milk provides a wide range of biologically active compounds that protect humans against diseases and pathogens. The purpose of this work is to describe the main aspects and research lines concerning bioactive peptides: from their chemistry, bioavailability, and biochemical properties to their applications in the healthcare sector. In this context, the uses of bioactive peptides in nutraceutical and functional foods have been highlighted, also taking into account the perspective of innovative applications in the field of circular bioeconomy.
... Phosphopeptide formation and resistance has been followed during simulated gastrointestinal digestion. The use of sequential hydrolysis with pepsin and pancreatin has been shown to release phosphorylated sequences previously reported in vivo, also in infant formula employing suitable conditions (Miquel et al., 2005). Moreover, isolation of the CPP fraction by selective precipitation or TiO2 chromatography allowed to identify several sequences containing the phosphorylated cluster (Miquel et al., 2006;Picariello et al., 2010). ...
During the last decade, there has been a growing interest in understanding food's digestive fate in order to
strengthen the possible effects of food on human health. Ideally, food digestion should be studied in vivo
on humans but this is not always ethically and financially possible. Therefore, simple in vitro digestion
models mimicking the gastrointestinal tract have been proposed as alternatives to in vivo experiments.
Thus, it is no surprise that these models are increasingly used by the scientific community, although their
various limitations to fully mirror the complexity of the digestive tract. Therefore, the objective of this
article was to call upon the collective experiences of scientists involved in Infogest (an international
network on food digestion) to review and reflect on the applications of in vitro digestion models, the
parameters assessed in such studies and the physiological relevance of the data generated when compared
to in vivo data. The authors provide a comprehensive review in vitro and in vivo digestion studies
investigating the digestion of macronutrients (i.e. proteins, lipids and carbohydrates) as well as studies of the bioaccessibility and bioavailability of micronutrients and phytochemicals. The main conclusion is that
evidences show that despite the simplicity of in vitro models they are often very useful in predicting
outcomes of the digestion in vivo. However, this has relies on the complexity of in vitro models and their
tuning towards answering specific questions related to human digestion physiology, which leaves a vast
room for future studies and improvements.
... 7,8 Casein phosphopeptides (CPPs) are phosphorylated casein-derived peptides that can be released via in vitro or in vivo enzymatic digestion of s1 -, s2 -and -casein. 9,10 They are reported to have multiple bioactive functionalities, including improved mineral absorption (mainly calcium), immunomodulatory, anti-oxidative, antithrombotic, angiotensin-converting enzyme inhibitory activity and enhanced intestinal 1940 www.soci.org N Lebetwa et al. ...
Background:
Current research on the gastro-intestinal digestion of milk-casein strongly suggests the existence of novel bioactive peptides having antiviral activities that are attributable to their immunostimulatory effects. Here, we investigated the antiviral activity of casein peptides rich in phosphate groups, such as casein phosphopeptide (CPP-III).
Results:
We prepared two types of CPP with different phosphorylation levels to clarify the role of the phosphate group. Further phosphorylation of CPP-III was conducted by dry heating with sodium pyrophosphate, while dephosphorylation was performed enzymatically using alkaline phosphatase and by alkaline treatment. Feline calicivirus (FCV) strain F9, a typical norovirus surrogate, and Crandell-Rees feline kidney cells were used as the target virus and host cells, respectively. Antiviral activity was determined based on MTT assay and qPCR quantification of antiviral cytokine mRNA expression. Higher cell viability was observed in the host cells treated with phosphorylated CPP-III, and a significant up-regulation of type 1 interferon expression was induced, compared with that treated with native CPP-III. However, dephosphorylation of CPP-III resulted in a decrease in the anti-FCV effect.
Conclusion:
CPP effect was enhanced by introduction of additional phosphates and conversely weakened by their elimination. Therefore, CPP-III phosphorylation represents an emerging approach to the production of food-grade antiviral agents.
... Another area of interest in studies of milk digestion is release of casein phosphopeptides (CPP), which are phosphorylated CN-derived peptides that can be released through enzymatic hydrolysis of α S1 -, α S2 -, and β-CN during digestion. These peptides form soluble complexes with calcium and other minerals, allowing them to be absorbed through the intestinal wall at pH levels that would otherwise cause them to precipitate (Miquel et al., 2005). The presence of CPP during intestinal digestion of milk and variations due to type of processing may have implications on the bioavailability of minerals. ...
... Following the procedure of Miquel et al. (2005), 0.3 g of lyophilized defatted milk sample was weighed into a 50-mL centrifuge tube and dissolved in 10 mL of 1 M Tris, pH 8. While the mixture was being stirred, 20 mL of 10% (wt/vol) CaCl 2 was added, followed by slow addition of 20 mL of 100% ethanol. ...
... The CPP function primarily to bind and solubilize minerals and are important in enhancing the bioavailability of calcium, zinc, and iron (Meisel and Fitzgerald, 2003;Miquel et al., 2005). Casein phosphopeptides with known mineral binding capabilities include α S1 -CN f(46-58), α S1 -CN f(59-79), α S2 -CN f(1-32), α S2 -CN f(55-64), β-CN f(1-25), and β-CN f(33-48) (Meisel and Fitzgerald, 2003). ...
Central to commercial fluid milk processing is the use of high temperature, short time (HTST) pasteurization to ensure the safety and quality of milk, and homogenization to prevent creaming of fat-containing milk. Ultra-high-temperature sterilization is also applied to milk and is typically used to extend the shelf life of refrigerated, specialty milk products or to provide shelf-stable milk. The structures of the milk proteins and lipids are affected by processing but little information is available on the effects of the individual processes or sequences of processes on digestibility. In this study, raw whole milk was subjected to homogenization, HTST pasteurization, and homogenization followed by HTST or UHT processing. Raw skim milk was subjected to the same heating regimens. In vitro gastrointestinal digestion using a fasting model was then used to detect the processing-induced changes in the proteins and lipids. Using sodium dodecyl sulfate-PAGE, gastric pepsin digestion of the milk samples showed rapid elimination of the casein and α-lactalbumin bands, persistence of the β-lactoglobulin bands, and appearance of casein and whey peptide bands. The bands for β-lactoglobulin were eliminated within the first 15 min of intestinal pancreatin digestion. The remaining proteins and peptides of raw, HTST, and UHT skim samples were digested rapidly within the first 15 min of intestinal digestion, but intestinal digestion of raw and HTST pasteurized whole milk showed some persistence of the peptides throughout digestion. The availability of more lipid droplets upon homogenization, with greater surface area available for interaction with the peptides, led to persistence of the smaller peptide bands and thus slower intestinal digestion when followed by HTST pasteurization but not by UHT processing, in which the denatured proteins may be more accessible to the digestive enzymes. Homogenization and heat processing also affected the ζ-potential and free fatty acid release during intestinal digestion. Stearic and oleic acids were broken down faster than other fatty acids due to their positions on the outside of the triglyceride molecule. Five different casein phosphopeptide sequences were observed after gastric digestion, and 31 sequences were found after intestinal digestion, with activities yet to be explored. Processing affects milk structure and thus digestion and is an important factor to consider in design of foods that affect health and nutrition.
... Moreover, bioactive peptides released by microbial enzymes could be attacked by other enzymes present in GIT and produce some other peptides (Shimizu, 2007;Moller et al., 2008). Although bioactive peptides released by the action of site specific protease like trypsin or chymotrypsin on milk protein are well (Miquel et al., 2005). ...
Dairy products are associated with numerous health benefits. They are good source of nutrients like carbohydrates, protein (bioactive peptides), lipids, minerals and vitamins which are essential for growth, development and maintenance of the human body. Accordingly, dairy bioactive peptides are one of targeted compounds present in different dairy products. Dairy bioactive compounds can be classified as anti-hypertensive, anti-oxidative, immmunomodulant, anti-mutagenic, anti-microbial, opoid, anti-thrombotic, anti-obesity and mineral-binding agents depending upon biological functions. These bioactive peptides can easily be produced by enzymatic hydrolysis, during fermentation and gastrointestinal digestion. For the reason, fermented dairy products like yogurt, cheese and sour milk are gaining popularity worldwide and considered excellent sources of dairy peptides. Furthermore, fermented and non-fermented dairy products are associated with lower risks of hypertension, coagulopathy, stroke and cancer insurgences. The current review article is an attempt to disseminate general information about dairy peptides and their health claims to scientists, allied stakeholders and certainly readers.
... Similarly, after SGID of an adapted and a follow-up milk-based infant formula, CPPs were detected with 50 % of the fragments containing the acidic motif. In addition, the CPP sequences initially present in the infant formulas were still found after SGID, indicating that they were not degraded during digestion and would have the potential to play a physiological role in the organism (Miquel et al. , 2005 ). β-CN (f 1-25) binding to different minerals (Ca, Mg, Zn and Cu) has been studied in conditions mimicking the ileum (pH 8 and 37 °C) using isothermal titration calorimetry. ...
Milk proteins and milk protein-derived peptides have been widely studied for their health enhancing properties. This chapter presents the updated scientific knowledge on the bioactive properties of milk protein-derived peptides. The different bioactive properties which have been attributed to milk protein-derived peptides are discussed. These include mineral binding, cardioprotective, antidiabetic, satiating, opioid, antimicrobial, immunomodulatory, anticancer and antioxidant activities. The structure-function relationship is presented for the aforementioned bioactive properties based on current scientific knowledge. For each bioactive property, the data obtained in vitro is discussed, followed by an analysis of the information obtained from animal and human intervention studies. To date, most studies have been conducted in vitro. However, an increasing number of in vivo studies testing the efficacy of milk protein-derived peptides are being conducted. In certain instances, the in vivo studies have confirmed the bioactivity of specific milk protein-derived peptides or milk protein hydrolysates. However, conflicting data still exist in the scientific literature, which demonstrates that the bioactive properties observed in vitro do not always translate in vivo. Detailed knowledge of the peptide sequences responsible for the bioactive properties, together with a better understanding of the bioavailability and stability of these peptides in vivo may help to enhance the development of milk protein hydrolysates with health promoting capabilities in humans. Ultimately, this may lead to the approval of health claims by the relevant regulatory agencies.
... The size of the hydrolyzed fragments could be critical in terms of the antihypertensive activity as di-and tripeptides are easily absorbed in the intestine and may reach the blood and other target sites (Hara et al. 1984). Furthermore, SGID had a Bovine (Miquel et al. 2005) Antimicrobial κ-casein f (106-169) -Bovine (Brück et al. 2003) positive impact on the unhydrolyzed samples and did not affect the hydrolyzed wheybased formulas with respect to ACE inhibition. This suggests that whey hydrolysates survive physiological digestion. ...
... According to the authors, calcium could be bound preferentially to CPPs with the cluster sequence SpSpSpEE and iron and zinc to CPPs with the phosphorylated cluster and phosphoserine residues. The release of CPPs with metal-chelating properties following SGID of infant formulas was also reported in previous studies (Miquel et al. 2005). Many of these CPPs were only detected in probiotic formulas indicating that they are formed by the action of bifidobacteria. ...
