Bioactive Peptides from Walnut Residue Protein - PMC

5.1. Antioxidant Peptide

Since Harman put forward the theory of free radicals, it has been recognized that the free radicals produced by oxidation in human body are related to human aging and many diseases [45]. The research on reactive oxygen species and free radicals has also become afrontier and hot spot of modern life science. Therefore, the preparation and extraction of antioxidants have developed rapidly, and the use of antioxidants has become more and more extensive. They are not only used as antioxidants in food processing, but also as functional factors in the development of health food and cosmetics. There are many kinds of antioxidants. However, the chemical synthesis of antioxidants such as BHA, BHT, etc. has its own toxic and side effects that are not conductive to human health. Governments of various countries have issued mandatory policies to regulate their acceptable daily intakes (ADI) value, and excessive addition is strictly prohibited. Based on the above reasons, screening and evaluation of natural resources with strong antioxidant activity has become a new trend in biological, medical and food science research. Antioxidant peptide is a kind of bioactive peptide with antioxidant function, which can effectively remove the excess active oxygen free radicals in the body, protect the normal structure and function of cells and mitochondria, prevent the occurrence of lipid peroxidation, and then play the role of delaying aging and preventing diseases [45,46]. In addition, it also has many advantages, such as high water solubility, low foaming, high safety, wide application range, etc. Therefore, it has a good market prospect to develop and utilize high-efficiency antioxidant peptides.

You can find more information on our web, so please take a look.

In recent years, walnut residue, as a potential plant protein resource, has attracted more and more attention. Many researchers have successfully extracted walnut peptide products with strong antioxidant activity from walnut protein, characterized the structure of these products, identified and isolated a number of small peptides with antioxidant activity ( ), and explored and discussed the antioxidant mechanism of walnut peptide.

Table 2

HydrolaseSITACS *RMM orM/CRefPepsinRP-HPLC-ESI-MSADAF423.23 Da[19]TrypsinUPLC-ESI-MS / MSWSREEQEREE Da[43]TrypsinUPLC-ESI-MS / MSADIYTEEAGR Da[43]Neutral proteaseRP-HPLC,nanoLC-ESI&#;MS/MS,de novo sequencingLAGNPDDEFRPQ. Da[26]Neutral ProteaseRP-HPLC,nanoLC-ESI&#;MS/MS, de novo sequencingVEDELVAVV971. Da[26]PapainRP-HPLCCTLEW651. Da[47]Tripsin, Viscozyme LUPLC-ESI-Q-TOF-MS/MSGGW319. Da[48]Tripsin, Viscozyme LUPLC-ESI-Q-TOF-MS/MSVYY444. Da[48]Tripsin, Viscozyme LUPLC-ESI-Q-TOF-MS/MSLLPF489.Da[48]PancreatinESI-MS/MSYS269.11 m/z[12]PancreatinESI-MS/MSYSVH505.23 m/z[12]PancreatinESI-MS/MSYK310.14 m/z[12]PancreatinESI-MS/MSYT283.13 m/z[12]PancreatinESI-MS/MSLPC331.16 m/z[12]PancreatinESI-MS/MSEM279.13 m/z[12]PancreatinESI-MS/MSCA192.14 m/z[12]PancreatinESI-MS/MSSQK362. m/z[12]PancreatinESI-MS/MSCR288. m/z[12]PancreatinESI-MS/MSCHC362. m/z[12]PancreatinESI-MS/MSGHC316. m/z[12]PancreatinESI-MS/MSYA253. m/z[12]PancreatinESI-MS/MSYG239. m/z[12]PancreatinESI-MS/MSNW318. m/z[12]Alcalase and protamexHPLC-FTMS, de novo sequencing, scoring methodVEGNLQVLRPR [44]Alcalase and protamexHPLC-FTMS, de novo sequencing, scoring methodLAGNPHQQQQ [44]Alcalase and protamexHPLC-FTMS, de novo sequencing, scoring methodHNLDTQTESDV [44]Alcalase and protamexHPLC-FTMS, de novo sequencing, scoring methodAGNDGFEYVTLK [44]Alcalase
protamexHPLC-FTMS, de novo sequencing, scoring methodWSVWEQELEDR [44]Alcalase and protamexHPLC-FTMS, de novo sequencing, scoring methodQQRQQQGL [44]Alcalase and protamexHPLC-FTMS, de novo sequencing, scoring methodAELQVVDHLGQTV [44]Alcalase and protamexHPLC-FTMS, de novo sequencing, scoring methodEQEEEESTGRMK [44]Alcalase and protamexHPLC-FTMS, de novo sequencing, scoring methodVEDELVAVV [44]AlcalaseLC-ESI-QTOFAGGA [42]Simulating gastrointestinal digestionRP-HPLC-UPLC-QTOF-MSTY [21]Simulating gastrointestinal digestionRP-HPLC-UPLC-QTOF-MSSGGY [21]Open in a separate window

The mechanism of antioxidant action of bioactive peptides, the relationship between antioxidant function and the molecular structure of bioactive peptides has been explored more and more. It is reported that the antioxidant properties of active peptides are closely related to many factors, such as their ability to remove reactive oxygen, the ability to donate hydrogen atoms or transfer electrons to free radical compounds, and the ability to chelate transition metals such as copper or iron [45,49]. Antioxidant peptides can scavenge free radicals by inhibiting hydrogen atom transfer and electron transfer, or prevent the production of free radicals and active oxygen based on instantaneous metal ion chelation [50,51]. The scavenging ability of active peptides to 2,2&#;-azino-bis (3-ethylbenzthiazoline-6-sulfonicacid) cation radicals (ABTS+), hydroxyl radicals (.OH), 1,1-diphenyl-2-picryl-hydrazyl free radicals (DPPH), and other radicals, the scavenging ability of active peptides to superoxide and other active oxygen and the chelating ability of active peptides to metal ionsare the scientific evaluation criteria for the antioxidant ability of active peptides. In addition, the amino acid composition, sequence, molecular size and other information of the active peptide are very important to explore the molecular mechanism of the antioxidant activity of the active peptide. Gu et al. isolated fourteen new antioxidant peptides from walnut protein trypsin hydrolysate, identified and analyzed the structure of these peptides, found that most of these fourteen antioxidant peptides contain tyrosine or cysteine, and speculated that tyrosine and cysteine may contribute a lot to the antioxidant activity of the active peptide. The possible mechanism is that cysteine residue has reductive SH group and potential antioxidant activity, while tyrosine has the ability of donating hydrogen atom, which is a good material to remove ABTS+ and other free radicals [12,52]. Feng et al. isolated six walnut peptides from the simulated digestion products of walnut meal protein, and characterized their antioxidant activity and molecular structure, respectively. According to the structure-activity relationship of the six peptides, they proposed that the amino acid sequence and composition of the peptides play an important role in the antioxidant activity [21]. The report from Chen et al. [43] is slightly different from that of Feng et al. [21]. Chen et al. identified seventy-seven peptides from the trypsin hydrolysate of defatted walnut residue, and isolated WSREEQEREE and ADIYTEEAGR, two walnut peptides with strong protective effect on PC12 cells damaged by H2O 2. They found that the amino acid mixture with the same composition as these two peptides had no protective effect on H2O2 damaged cells, so they proposed that the main factor affecting the antioxidant effect is the amino acid sequence, not the amino acid composition [43]. Sheng et al. identified more than peptides from walnut hydrolysates by de novo sequencing. On this basis, they established a scoring method based on structure and activity, and screened eight new antioxidant peptides of walnut. It was found that the content of Glu, Arg, ASP, Gly and other specific amino acids was closely related to the scavenging ability of hydroxyl radicals of peptide [44]. As far as we know, the antioxidant mechanism of walnut peptide has not been completely cleared up at present, and further research is needed.

In addition, many literatures reported that walnut peptides with high antioxidant activity also have excellent anti-tumor activity [22,24], improve memory activity [43,48,53,54], enhance immunity and anti-fatigue activity [54], etc. Wang et al. found that the mixed solution of low molecular weight walnut peptide improved the memory deficit behavior induced by sleep deprivation in rats (666 mg/kg peptide daily recommended). They identified three highly active neuroprotective peptides (GGW, VYY and LLPF) from the solution. Their experimental results showed that these three peptides could effectively protect PC12 cells from apoptosis induced by glutamine, and the relative survival rate of cells were 78.29 ± 3.09%, 80.65 ± 1.74% and 83 97 ± 3.06%. In addition, all of these peptides can reduce the consumption of antioxidant enzymes (SOD and GSH PX), inhibit the production of ROS, regulate the expression of apoptosis related proteins (Bax and Bcl-2) [48]. Therefore, walnut peptide may be considered as a potential health care product against neurodegenerative diseases related to memory impairment. Ma et al. found that the walnut peptide prepared by papain enzymatic hydrolysis of walnut protein under the optimum conditions (substrate concentration 5%, enzyme substrate ratio 10%, temperature 60 °C, time 3 h) not only had superior antioxidant activity, but also showed excellent anti-cancer activity. The amino acid sequence of walnut peptide with high anticancer activity purified by them is CTLEW, with a molecular weight of 651. Da. It is a new type of biological peptide with amphiphilic structure, which can induce apoptosis and autophagy of cancer cells, significantly inhibit the growth of cancer cells, but has no cytotoxic effect on normal cells. In addition, CTLEW can promote the proliferation of mouse spleen lymphocytes and the secretion of IL-2, enhance the phagocytic ability of mouse macrophages, and improve the immunity of the body [47]. Fang et al. reported that low molecular weight (is less than 3 kDa) WBPS (800 mg/kg daily recommended) can significantly extend the swimming time of mice, increase the liver glycogen content in mice, reduce the lactic acid content in mice, increase the immune organ index of mice, promote the proliferation of mouse intestinal T lymphocytes, and has significant anti-fatigue and immunomodulatory effects [46]. These results show that walnut protein extracted from walnut residue can be hydrolyzed moderately to prepare highly active anticancer peptide, and walnut protein hydrolysates and purified anticancer peptides can be fully considered for the development of functional foods and anticancer drugs.

What is Collagen? Your Ultimate Guide to Collagen Supplements

Collagen supplements are one of the most hyped ingredients in the health and beauty industry, but are they really worth the hype? Consider this a registered dietitian&#;s complete guide to collagen supplements and its benefits.

One of the most common questions I get is &#;what are your thoughts on collagen supplements&#;, and rightfully so! Collagen is one of those trendy ingredients showing up everywhere. You can find collagen creamers, collagen protein bars, and collagen waters without too much trouble. Take a walk down the supplement aisle and you&#;ll find collagen supplements in the form of powders, capsules and gummies. 

But what exactly is collagen and what are the benefits? Consider this your ultimate guide to collagen supplements. It&#;s a simple but science-based look into collagen and its benefits.

I also wrote a book all about collagen with Sterling Publishing, it&#;s benefits and 30 different recipes called Collagen: Self-Care Secrets to Eat, Drink and Glow. 

WHAT IS COLLAGEN?

Collagen is the most abundant protein in the body, making up about a third of all protein. Think of it as the &#;glue&#; that holds your body together. It&#;s one of the main structural components of your skin and connective tissues, like tendons and cartilage. It also plays an important role in our joints, bones, hair, nails, and even the gut (1). Needless to say, it&#;s a pretty important part of the body!

Since collagen is a protein, it is made up of amino acids. (For a refresher on protein, check out the Protein 101: A Simple Guide to Understanding Protein post). What makes collagen unique is that the majority of the total amino acid content comes from glycine, proline, and hydroxyproline.

TYPES OF COLLAGEN

There are 28 different types of collagen found in different parts of the body. The different amino acid makeup of each type of collagen depends on the type of structure it&#;s a part of (ex. joint cartilage vs. gut lining). However, there are five main types of collagen, with 80-90% of all collagen in the body consisting of Type I, Type II and Type III. 

Type I &#; this is the most abundant type of collagen and found in almost every tissue of your body. This includes tendons, skin, bones, cartilage and connective tissues.

Type II &#; found mostly in cartilage. This includes joints between bones, ends of ribs, ears & nose, etc.

Type III &#; often found alongside Type I collagen, as well as in muscles, organs, arteries and some connective tissues in the liver, spleen and more.

COLLAGEN SYNTHESIS

Our bodies naturally produce (aka synthesize) collagen. Just like how our bodies pull from the pool of amino acids to make new muscles, it does the same to make collagen. The body uses the necessary amino acids along with the help of certain vitamins and minerals like vitamin C, zinc, and copper to make collagen protein when and where it&#;s needed.

That being said, as we age, collagen in the body begins to deteriorate and production slows. This process starts in the late 20s early 30s and is most noticeable in our skin as fine lines and wrinkles. As we produce less collagen, other noticeable issues include brittle hair and nails, digestion issues, more pronounced cellulite and joint pain.

WHAT CAUSES LOSS OF COLLAGEN?

Age is one of the biggest players in collagen breakdown. Unfortunately, we can&#;t turn back time or reverse the process but we can make diet and lifestyle changes that lessen the impact. These include: 

• • minimizing a diet high in refined sugars

  • reducing stress

  • preventing nutrient deficiencies such as protein and other vitamins and minerals

  • not smoking

  • be smart sun exposure

  • reducing exposure to air pollution

DIFFERENT TYPES OF COLLAGEN SUPPLEMENTS

Collagen supplements come from connective tissue of cows, fish and sometimes chicken. Most supplements you&#;ll find are made up of bovine collagen which are sourced from cows. However, marine collagen, which is sourced from fish, is growing in popularity due to it being absorbed up to 1.5 times more efficiently. 

Hydrolyzed Collagen  (aka collagen peptides)

To make this form of collagen, collagen proteins are broken down by an enzymatic process called hydrolysis, into a smaller, more easily absorbable form called peptides. This is the same thing as collagen peptides. 

This is a tasteless, odorless powdered form of collagen that can be added into either hot or cold liquids. It essentially just dissolves into whatever it is mixed with, making it extremely easy to take.

Capsules

These are convenient in the sense that you don&#;t have to add it to a liquid or food in order to take it. You just have to remember to take it! If you already have a supplement regimen in place, adding a collagen supplement in capsule form may be an easy addition.

Gummies

I typically do not recommend gummy supplements. Most of the time they are sugar-based and have added ingredients and fillers.

Keep in mind quality matters when it comes to supplements, especially collagen supplements. I recommend purchasing high-quality collagen made from grass-fed, pasture-raised cows or wild caught fish. Also, make sure the products have been third-party tested for purity.

FOOD SOURCES OF COLLAGEN

Whole food sources of collagen come from meat, fish, and eggs. When you consume these foods you are getting collagen. However, since the best sources of collagen are found in skin, tendons and cartilage, it&#;s likely you aren&#;t getting a lot from lean animal protein. Another whole food source of collagen is bone broth, which is made by simmering the collagen-rich parts of the animal and bones (hence the name &#;bone&#; broth). Bone broth can be used in cooking or sipping as a warm, savory beverage.

Other nutrients play an important role in collagen formation and protect the body&#;s collagen from damage and deterioration. Vitamin A, Vitamin C, iron, zinc and copper are all necessary for proper collagen production. Thankfully a whole food, plant-based diet can easily provide these nutrients. Good sources of these nutrients include carrots, sweet potatoes, kale (vitamin A), strawberries, bell peppers, broccoli, kiwi (vitamin C), pumpkin seeds, cocoa powder, cashews (zinc), cashew, sunflower seeds, chickpeas (copper), spinach, lentils and black beans (iron.

Omega-3 fatty acids protect the body&#;s collagen stores from damage and work to help reduce free radicals and inflammation. The best sources of omega-3s include hemp seeds, chia seeds, walnuts, flaxseeds and fatty fish like salmon.

WHAT ARE THE BENEFITS OF COLLAGEN?

There are numerous claims supporting the benefits of collagen. These range from decreasing wrinkles and improving joint health, to aiding in weight loss and healing the gut. But what does science say?

The research on collagen is limited since it&#;s a fairly new topic of study. Most of the research has looked at collagen in regards to skin, joint and bone health. Other studies look at the main amino acids (glycine and proline) in collagen and their benefits or a collagen as a protein source. But the research looks promising!

Skin 

Various forms and dosages of collagen have been studied showing improvements in skin elasticity, keeping skin looking younger and preventing signs of aging by keeping skin firm and hydrated (2, 3, 4). Research also shows that this can increase the smoothness and help reduce the appearance of cellulite (5).

Link to SEMNL

Hair and Nails

As collagen production slows, hair and nails can become brittle and weak. Collagen can help hydrate your hair and nails, helping them grow faster and thicker (6).

Bone and Joint Health

Collagen has been shown to help reduce inflammation (7). This can be extremely beneficial for those with Rheumatoid Arthritis, Osteoporosis or other inflammatory disorders (8, 9, 10). Collagen also helps in preventing the breakdown of joints, especially in individuals with osteoarthritis. Another study found that athletes given the collagen supplement had less joint pain than those that did not receive the supplement (11).

Gut Lining

Numerous studies have shown that those with inflammatory bowel diseases have less collagen in the lining of their intestines (12, 13, 14). Collagen is 30% glycine. Glycine is shown to help repair leaky-gut, a condition where the lining of the small intestine becomes damaged, causing undigested food particles, waste products and bacteria to &#;leak&#; through the intestines into the bloodstream (15). Collagen can help restore the lining of the stomach and intestines and improves nutrient absorption (16).

Muscle gain and satiety

Protein is an important part of the diet and out of all the macronutrients provides the most satiety &#; meaning it keeps you full longer. Adding collagen can also help with muscle gain when combined with strength training by providing necessary amino acids for muscle growth and recovery following exercise (14).

COMMONLY ASKED QUESTIONS:

IS COLLAGEN VEGAN?

There are no sources of collagen that are vegan. Collagen is only made in humans and animals. You may have seen &#;vegan collagen&#; supplements but these only contain &#;collagen-boosting&#; nutrients like vitamin C, zinc, and amino acids derived from plant sources. They do not contain actual  collagen derived from animal connective tissues.

ARE THERE PLANT-BASED SOURCES OF COLLAGEN?

No. But there are foods that boost our natural collagen production. Even though your body can produce the necessary amino acids for collagen production, it&#;s important to make sure you are getting enough proline and glycine to ensure your body has enough to support this process. Soy, beans, spinach, cabbage, cauliflower, avocados, asparagus, and sunflower seeds.

This is a fantastic resource on plant-based foods that support collagen production.

CAN COLLAGEN REPLACE MY PROTEIN POWDER?

Collagen can be a good replacement for your traditional protein powder if you are eating a well-balanced diet and getting other sources of protein. Collagen is an incomplete protein, meaning it doesn&#;t contain all of the 9 essential amino acids in adequate amounts. If you&#;re adding protein powder as a main protein source or to increase muscle mass, you may be better off with a different source of protein. 

HOW MUCH COLLAGEN SHOULD I TAKE?

The recommended serving size is about 10-20g of collagen peptides (about 1-2 scoops). This ensures a good amount of protein and collagen. Remember more isn&#;t always better. You want a variety of protein in your diet to get the full amount of essential amino acids your body needs. 

WHAT IS THE DIFFERENCE BETWEEN COLLAGEN AND GELATIN?

Collagen and gelatin are not the same. Gelatin is made from collagen that has gone through a heating process that alters the state of the amino acids. This means that collagen can be dissolved in both cold and warm liquid, whereas gelatin &#;gels&#; in cold liquid but dissolves in hot liquid. Think of Jell-o, puddings or gummies which are made from gelatin.

Nutritionally, collagen and gelatin have same amino acid profile and the same benefits. 

CAN I TAKE COLLAGEN WHILE PREGNANT OR NURSING?

Yes, collagen is just another form of protein and is safe to take while pregnant or nursing. In fact, glycine needs greatly increase during pregnancy making collagen an excellent choice for pregnant women. 

Additionally, most women aren&#;t meeting their protein needs during pregnancy and postpartum. Pregnant women need protein for blood sugar management, minimizing nausea, and growing baby&#;s cells and the placenta. Collagen is in an optimal form for all of this. It also supports joints, pelvic floor tissue, skin elasticity, and postpartum hair and skin.

Make sure you read labels and make sure your supplement doesn&#;t contain any other ingredients or fillers. As always, it is best to talk with your doctor about supplements before taking any at this special time in your life.

HOW TO CHOOSE A COLLAGEN SUPPLEMENT?

Not all collagen is created equal. Since collagen is an animal product, choosing collagen from a reputable source is important. Look for collagen that comes from grass-fed, pasture-raised animals or wild-caught fish. The collagen should not have any additional fillers or additives. 

Overall, when choosing a collagen supplement opt for a brand that is transparent about their processes and sourcing. Also, look for collagen that is certified by a third-party quality-testing company, like NSF International or United States Pharmacopeia (USP).

I like these brands:

• • Needed Collagen Peptides (code NOURISHEDBYNUTRITION)
  • Vital Proteins
  • Ancient Nutrition

DO YOU USE COLLAGEN?

Yes, I try to incorporate collagen as a daily ritual. It&#;s easy to use and I like that it dissolves into basically anything. I also wrote a book all about collagen with Sterling Publishing, it&#;s benefits and 30 different recipes called Collagen: Self-Care Secrets to Eat, Drink and Glow. 

Keep in mind. collagen isn&#;t my only protein source and I also use a plant-based protein powder. I love the results I&#;ve seen with my hair and skin and it&#;s easier on my digestion than some of the other protein powders I&#;ve tried. 

It&#;s also nice to take individual packets of collagen me on the go, especially on trips. I keep one in my purse just in case I&#;m in a pinch or feel like I need a source of protein to round out a snack or meal when the options are limited.

HOW TO INCORPORATE COLLAGEN INTO MY DIET?

Adding collagen into your diet can be pretty easy since it dissolves into pretty much anything. Try adding it to a liquid such as a smoothie, juice, water, tea or coffee, or boosting the protein content of your favorite foods: oatmeal, baked goods, raw treats, snack balls, pancakes, salad dressings and sauces.

For more easy recipe to get you started to try these Cashew Coconut Collagen Bites or Date Sweetened Collagen Hot Chocolate. I have 30 more recipes is my book Collagen: Self-Care Secrets to Eat, Drink and Glow. 

THE BOTTOM LINE: SHOULD YOU TAKE A COLLAGEN SUPPLEMENT?

The current research on the effects of collagen is very promising, especially for skin, joint and gut health. 

I recommend considering your diet as a whole. Whole foods have a variety of nutrients that support collagen production and protect it from breaking down due to environmental damage. Your body has the ability to produce the amino acids to make most types of collagen. A well-balanced diet with adequate protein can provide you with enough of the necessary essential amino acids for your body to support collagen.

Remember, consuming collagen doesn&#;t mean the body uses it directly to make more collagen. The collagen is broken down into amino acids during the digestion process just like other proteins. The body is smart and prioritizes which proteins to make from the pool of amino acids based on where proteins are needed in your body.

That being said, if you are looking for an easy way to boost your protein intake, an excellent source of glycine or not eating animal products or much protein, taking a collagen supplement is a great option!

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