What is Glycation?

Glycation, sometimes called as non-enzymatic glycosylation, is the spontaneous non-enzymatic reaction wherein there is covalent attachment of free reducing sugars such as glucose, fructose, or their derivatives with the free amino groups of proteins, DNA, and lipids. The reaction forms Amadori products which undergo different permanent dehydration and rearrangement reactions that lead to the formation of advanced glycation end products (AGEs). Unlike glycosylation that is enzyme-mediated ATP-dependent attachment of sugars to protein or lipid and a common form of post-translational modification of proteins that is required for the functioning of the mature protein, glycation process leads to a loss of protein function and impaired elasticity of tissues such as blood vessels, skin, and tendons. Moreover, the glycation reaction is significantly increased in situations such as being in the state of hyperglycemia and tissue oxidative stress. Hence, most studies suggest that glycation is responsible for the development of diabetic complications and even aging. The glycation process matches well with the theory that the accumulation of metabolic waste promotes aging because there are no enzymes in the human body capable of removing the advanced glycation end products (AGEs).

As mentioned above, glycation reaction is highly accelerated in the presence of tissue oxidative stress. Oxidative stress has been implicated as a vital factor in the progression of multiple diseases such as chronic disease like diabetes, Alzheimer’s disease, and aging. Oxidative damage to proteins plays an important role in the development of the pathological modifications in the activities of biological mechanisms and cellular processes. Through glucose autoxidation and the non enzymatic covalent attachment of glucose molecules to circulation proteins that lead to the formation of AGEs, there is excessive production of reactive oxygen species that further worsens oxidative stress and damage to cells. 

Advanced glycation end products (AGEs)

Advanced glycation end products (AGEs), as stated above, are end products of glycation. They are proteins or lipids that become glycated as a result of exposure to free reducing sugars such as glucose, fructose, and their derivatives. AGEs were originally identified in the cooking process in a reaction termed as Maillard reaction. This reaction is known as the Maillard reaction. The glycation process is initiated by a chemical reaction between the reactive carbonyl group of a sugar or an aldehyde with a nucleophilic free amino group of a protein, leading to the rapid formation of an unstable Schiff base. This adduct then undergoes rearrangement to form a reversible and more stable Amadori product. These intermediate products undergo further irreversible oxidation, dehydration, polymerization, and cross-linking reactions resulting in the formation of AGEs over the course of several days to weeks. The figure below shows the whole process in the formation of AGEs: 

DEFINITION of GLYCATION and AGEs

(Image lifted from: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5643203/)

AGEs are the specific biomarker implicated in aging and the development, or worsening of many degenerative diseases. Moreover, these end products affect nearly all types of cells and molecules in the human body. They are strongly implicated as the causative role in the vascular complications of diabetes mellitus because they play a role as proinflammatory mediators in diabetes. Moreover, AGEs promote crosslinking of collagen which result in vascular stiffening and entrapment of low-density lipoprotein (LDL) particles in the arterial walls, increasing the risk for the development of cardiovascular diseases. Furthermore, these low-density lipoproteins can also undergo glycation caused by AGEs. With this, the lipoprotein can be oxidized. This oxidized LDL is one of the primary risks for the development of atherosclerosis. AGEs can bind to receptors for advanced glycation end products and can promote further oxidative stress and activation of inflammatory pathways producing a variety of clinical symptoms. 

Structural and functional alterations in plasma and extracellular matrix (ECM) proteins such as those found in the skin also occurs due to the toxic effects of endogenous and exogenous AGEs. This in particular causes crosslinking of proteins. Specifically, in human skin, glucose pane is the most important cross-link product formed with ECM. The glycated-myosin primarily affects myosin velocity and directionality in the human skin. Moreover, AGE accumulation in collagen results to changes in the biochemical and structural property of the components of the basement membrane affecting its elasticity, ionic charge, and thickness, which can produce the signs of aging such as wrinkles, fine lines, sagging skin, and the likes. 

With everything mentioned above, AGEs indeed have multiple pathological effects in the human body such as increasing vascular permeability which can lead to edema; increasing arterial stiffness leading to increased risk for arterial and cardiovascular diseases; impeding with nitric oxide leading to inhibition of vascular dilation; promoting oxidative stress; oxidizing LDL leading to activation of multiple inflammatory pathways; binding cells such as macrophage, endothelial, and mesangial in order to promote the secretion of a variety of cytokines leading to a variety of clinical effects; elevating Hemoglobin-AGE levels in people dealing with diabetes and accumulating with time and increasing from 5-50 fold over periods of 5–20 weeks in the retina, lens and renal cortex of diabetics. Studies show that inhibition of AGE formation significantly decreases the risk of developing nephropathy in diabetics. Therefore, substances that inhibit AGE formation may limit the progression of disease and may offer new tools for therapeutic interventions in the therapy of AGE-mediated disease; and lastly, activating the receptors for advanced glycation end products which have multiple cellular effects. The activation of cellular RAGE on endothelium, mononuclear phagocytes, and lymphocytes induces the production of free radicals and the expression of inflammatory gene mediators. Activation of the transcription factor NF-κB occurs due to the significant increases in oxidative stress. This also promotes the expression of NF-κB regulated genes that have been linked with the development of atherosclerosis.

Diet and Advanced glycation end products (AGEs)

As mentioned, AGEs were originally identified in the cooking process in a reaction termed as Maillard reaction. AGEs also exist in foods in addition to the natural formation within the body. Through the Maillard reaction, formation of new AGEs occur when animal-derived foods are cooked. Moreover, grilling, broiling, roasting, searing, and frying promote and exacerbate formation of new AGE.  The modern diet is a large source of AGEs because a wide range of foods in modern diets are predisposed to cooking or thermal processing for reasons of safety, food preparation, and convenience. Cooking is also greatly emphasized today in order to enhance flavor, color, and appearance of the food. Before, the potential biological role of dietary AGEs was ignored for a long time because of the assumption that they undergo negligible gastrointestinal absorption, thus their role in human health and disease causation was significantly ignored. However, recent studies with the oral administration of a single AGE-rich meal to human beings as well as tagged single protein-AGEs or diets supplemented with specific AGEs to animals coherently indicate that dAGEs are absorbed and contribute highly to the body’s AGE pool. Today, it has been proven that food rich with AGEs significantly undergo intestinal modification and absorption. Dietary ingested AGEs is easily absorbed through the intestine tract where it is rapidly metabolized and then excreted. Some parts of the AGEs are degraded by the intestinal microbiota, and approximately 10% of these reach the blood circulation wherein they are primarily distributed in liver and muscle cells. 

Recent animal studies in mice have shown that administration of AGE-rich diets has been linked with elevated circulating and tissue AGEs and the development of chronic illnesses such as atherosclerosis and kidney disease. Studies also show that dietary restriction of AGE-rich food reduces vascular and kidney dysfunction, development of diabetes type 1 and type 2, improves insulin sensitivity, and promotes wound healing. Longer lifespan was also observed in animal samples who were only given low dietary AGEs compared to those who were administered with High AGEs. 

Studies in healthy human beings have shown that the amount of dietary AGEs is directly proportional with the circulating AGEs in the body, as well as with markers of oxidative stress. Moreover, There is a significant decrease in the markers of oxidative stress and inflammation when subjects with diabetes or kidney disease and healthy subjects are put in dietary AGE restriction. Both findings from animal and human studies strongly recommend reduction and/or avoidance of dietary AGEs for this aid in delaying chronic diseases and aging in animals and possibly in human beings. 

The composition of food, temperature, exposure to the environment such air, humidity, pH, and methods and duration of cooking are all known to be factors that significantly contribute to the formation of AGEs in food. Commonly, foods with relatively high lipid and protein content are found to be the ones with the highest dietary AGE levels. Fat contains 30-fold higher AGE content than carbohydrate meal, while meat contains 12-fold higher AGE content. Moreover, studies have shown that there are significantly higher dietary AGE values in samples grilled at temperatures of 230°C for shorter cooking times when compared to samples boiled in liquid media at 100°C for longer periods. This finding proves that cooking time is less important in the formation of AGEs, while temperature and method of cooking contributes significantly in the AGE formation. Hence, the major sources of dietary AGEs are meat and meat-derived products, processed at high, dry heat such as in broiling, grilling, frying, and roasting. Reduction of up to 50% of the daily dietary AGEs consumption, while keeping the same primary nutrients can be achieved through alternative cooking methods such as boiling and stewing.

Anti-glycation Strategies: Reducing AGEs intake and Formation

Now that the sources and the adverse health effects of AGEs have been present, it is important to discuss how one can avoid and prevent the risk of developing these negative health outcomes through reducing the intrinsic and extrinsic sources of AGEs. There are multiple strategies in order to reduce AGE formation. Here is a list below on the things one can do:

  1. Dietary Intervention and Modifying Cooking Methods

There are multiple strategies to reduce the consumption of dietary AGE. Several natural products like herbs, condiments and spices, have shown some intrinsic antiglycation activity. Such examples are pre-treating meats with lemon, vinegar or with any acidic marinade before cooking has a significantly shown inhibition in the excessive rise of dietary AGE content during cooking. Polyphenols are proven to be potent inhibitors of fructose-mediated protein glycation. These compounds have been observed to be present in many herbs like sage, marjoram, tarragon, and rosemary. Even more potent than herb extracts are the spice extracts such as cloves, ground Jamaican allspice, and cinnamon. These spices were proven to be glycation inhibitors. Food is mainly composed of carbohydrates such as starches, fruits, vegetables, and milk. These types of food contain the lowest AGE concentrations. This may be due to the frequent higher water content or higher level of antioxidants and vitamins in these foods, which may reduce new dietary AGE formation. Moreover, most polysaccharides consist of non-reducing sugars, less likely to give rise to AGEs in this food category. The highest dietary AGE level per gram of food in this category was observed in dry-heat processed foods such as crackers, chips, and cookies. Addition of ingredients such as butter, oil, cheese, eggs, and nuts, which during dry-heat processing substantially accelerate production of dietary AGE, is most likely the cause of this significant increase in AGEs. Even though dietary AGEs in these snack types is relatively low, they still should be consumed moderately for they still present a health hazard especially to those who intake multiple snacks during the day.. Hence, another strategy to reduce dietary AGE consumption is to implement a food diet that consists of these healthy foods, instead of eating full-fat cheeses, meats, and highly processed foods.

Dietary AGE content is also affected by the preparation of common foods under varying conditions of water and heat. For example, Poached or steamed chicken had less than one fourth the dietary AGEs compared to roasted or broiled chicken. Scrambled eggs prepared in an open pan over medium-low heat had about one half the dietary AGEs of eggs prepared in the same way but over high heat. Higher dietary AGEs were observed in all foods that were exposed to higher temperatures and lower moisture levels compared to foods prepared at lower temperatures or with more moisture for equal weight of food. Thus, boiling, poaching, stewing, and steaming produce less dietary AGEs compared to frying, broiling, grilling, and roasting produce mores. It is good to note however that microwaving food does not raise dietary AGE content to the same extent as other dry heat cooking methods. But, it should only be cooked for a relatively short cooking time (6 minutes or less).

Multiple studies have compared different food and their dietary AGEs.  Studies show that meat contains the highest levels of AGEs based on standard serving sizes. Meats will likely contribute more to overall dietary AGE consumption even though fats tend to contain more dietary AGE per gram of weight. This is due to food habits such that meats are usually prepared in larger servings than fats. To elaborate further, when different meats were prepared by identical methods and were compared with each other, findings showed that the highest dietary AGE levels were observed in beef and cheeses followed by poultry, pork, fish, and eggs. Moreover, it is good to note that out of all the available meat in the market, lamb contains significantly low dietary AGEs compared to other meats. When cooked under dry heat, lean red meats and poultry contain high levels of dietary AGEs. This occurs because there are highly reactive amino-lipids and reducing sugars such as glucose and fructose in intracellular components of lean muscle. The presence of such compounds in lean muscles rapidly forms new dietary AGEs when they are exposed to heat such as when cooking.

In addition,  reduced-fat mozzarella, milk cheddar, and cottage cheese all contain lower dietary AGEs compared to higher-fat and aged cheeses including full-fat American and Parmesan. Pasteurization and/or holding times at ambient room temperatures during curing or aging processes can cause animal-derived foods such as cheese to contain larger amounts of dietary AGEs even if not cooked which is one of known mechanisms to drive the generation of new AGEs in foods. Moreover, in spite of the fact that it occurs at a slower rate, glycation-oxidation reactions still persist over time at cool temperatures, resulting in large accumulation of dietary AGEs in food such as cheese. Also, among the highest in dietary AGEs are high-fat spreads such as butter, cream cheese, margarine, and mayonnaise. This list is followed by oils and nuts. Just like the cheeses mentioned earlier, even if uncooked, both butter and different types of oils are still AGE abundant. This may be caused by several extraction and purification procedures which involves heat in combination with air and dry conditions.

Among the lowest of dietary AGEs are grains, legumes, breads, vegetables, fruits, and milk. However, the dietary AGE formation increases when they are prepared with added fats. Moreover, nonfat milk had significantly lower dietary AGEs than whole milk. Milk-related products with a high moisture index such as yogurt, pudding, and ice cream were also found to have low levels of AGEs.

Hence, in order to reduce AGE consumption, one must take note of these foods. Eating foods that have low dietary AGE levels is the key in order to decrease adverse health effects such as insulin resistance and type 2 diabetes. It is highly recommended that dietary restrictions of these AGE-rich foods must be followed. Moreover, eating habits with low dietary AGE levels may aid in preserving the body’s natural defenses against a wide array of chronic disease through reducing the risk of developing oxidative stress and inflammation. Moreover, restricting dietary AGEs helps protect against premature aging and results in a lower risk of heart and kidney disease, increased insulin sensitivity, and lower levels of AGEs in blood and tissues by approximately 53%. However, it is good to note that even though a reduction in dietary AGEs has been proven to offer a wide range of health benefits, as of the present there are no guidelines regarding safe and optimal intake. But just to get a rough idea, a high-AGE diet is frequently referred to as anything significantly above 15,000 kilounits daily, and anything below this value is considered low. So if you are consuming too many AGEs, might as well consider your diet.

  1. AGEs Inhibitors

There are a lot of different AGE inhibitors that are being studied today that can be used as a potential controller of AGE formation, reducing the risk of developing its adverse health effects. These AGEs inhibitors include Guanidine compounds, such as aminoguanidine and metformin that can trap α‐dicarbonyl compounds hence preventing their further reactions with free amino groups of proteins, and Vitamin B complexes such as pyridoxamine and thiamine pyrophosphate. 

  • Aminoguanidine

Aminoguanidine is the prototype therapeutic agent for the inhibition of AGEs formation. It was originally introduced as a nucleophilic hydrazine compound. This compound is utilized in order to trap reactive carbonyls formed during the Maillard reaction. It specifically targets Amadori intermediates so that it would halt the formation of AGEs. Other than reacting with Amadori carbonyl groups of glycated proteins, it has also been shown to affect the dicarbonyl compounds such as methylglyoxal, glyoxal, and 3-deoxyglucosone. Animal studies have shown that aminoguanidine can prevent the arterial wall protein cross-linking. Thus,  having the capacity to improve cardiovascular risk in diabetic patients. Moreover, aminoguanidine prevented the formation of AGEs‐modified apoA‐1, thereby improving the capacity of apoA‐1 to promote cholesterol efflux from human monocytes. This strongly suggests that the compound may exert atheroprotective properties through promoting reverse cholesterol transport in diabetic patients. Furthermore, aminoguanidine prevents oxidative modification of low‐density lipoprotein. It also binds reactive aldehydes produced during lipid peroxidation, preventing their subsequent conjugation and lipid oxidation.  Hence with these evidences, aminoguanidine is a promising therapeutic agent for the management of diabetic dyslipidemia.

In addition, arterial elasticity is impaired due structural modification of collagen by AGEs. Aminoguanidine has been shown to increase vascular elasticity, decrease vascular permeability, and improve left‐ventricular‐arterial coupling. It also prevents decreases in myocardial compliance and risk for cardiac hypertrophy. Aminoguanidine also reduces plaque formation in the aorta.  These findings suggest aminoguanidine may play a vital role in improving symptoms with people who have diabetes‐induced vascular impairment through inhibiting the AGEs formation on collagen in the arterial wall and the heart and as a potential treatment for accelerated atherosclerosis in diabetes.

  • Metformin

This drug is used as an antihyperglycemic agent for patients with type 2 diabetes. This compound belongs to the guanidine family like aminoguanidine, suggesting that it may also have a potential role in the inhibition of glycation reactions. Several studies show inhibitory effects of metformin on protein glycation. Research has shown that metformin can decrease the expression of receptor for AGEs and can consequently block the downstream signaling of AGEs in cultured endothelial cells via a redox‐sensitive nuclear factors such as NF‐κB. Metformin also has been proven to have the ability to inhibit glycation of Low density Lipoproteins and to prevent the in vitro formation of foam cells. Moreover, functional and structural alterations of the diabetic myocardium can be associated with glycation, however chronic metformin treatment can be used in order to prevent such diabetic complications. Hence, blockade of glycation reaction is probably one of the possible mechanisms that can prove the beneficial effects of metformin on diabetic vascular complications.

  • Vitamin B complex

Recent studies have shown that vitamin B complexes such as pyridoxamine and thiamine pyrophosphate may have a potential role in inhibiting the formation of AGEs. Pyridoxamine is a post-amadori inhibitor of AGEs formation. During lipid peroxidation reactions, it also prevents the formation of advanced lipoxidation end products on protein. It is effective at inhibiting AGEs formation at three different levels through the following methods: (1) blocking oxidative degradation of the Amadori intermediate of the Maillard reaction; (2) removing of toxic carbonyl products of glucose and lipid degradation; and (3) confining of reactive oxygen species.

In addition, vitamin B complexes such as pyridoxamine protects endothelial cells against high glucose-induced DNA oxidative damage. It also inhibits copper-catalyzed low density lipoprotein oxidation. It also decreases high glucose-induced free radicals generation and lipid peroxidation in human red blood cells. Furthermore, Pyridoxamine was found to inhibit AGEs/ALEs formation and hyperlipidemia. It is also good to note that daily administration of an active form of vitamin B6, pyridoxal phosphate, significantly prevented the development of  albuminuria, glomerular hypertrophy, mesangial expansion, and interstitial fibrosis in association with the reduced expression of Receptors for AGEs in the kidney 

  1. Physical Exercise 

Glycemic control and consequent reduction of AGEs accumulation in diabetic patients and during aging can be achieved through increasing physical activity. Studies have shown that people who are relatively more active such as trained athletes have generally lower circulating AGEs. They also have lower AGEs cross-links in tendons compared to people with a sedentary lifestyle. Regular exercise also has strong positive effects against the renal accumulation of AGEs. Physical activities prevent renal AGE deposition in aging animal and human subjects. Specifically, treadmill exercise reduces AGEs accumulation and has been shown to have protective effects on the retina against AGEs.

Generally, regular exercise has a wide range of benefits to physical capacity, hypertension, oxidative stress, and lipid metabolism. It effectively prevents reactive oxygen species generation and increases the enzymatic activities of antioxidants. The reduction in the reactive intermediate sources available for glycation may be caused by  higher energy demands induced by physical exercise. The inhibition of AGE formation by regular physical activity may be the primary mechanism of exercise-associated antioxidant activity because the protein glycation reaction is driven and accelerated by reactive oxygen species. Lastly, regular physical exercise also can improve glycemic control, which can reduce the formation and accumulation of AGEs in tissues.

In conclusion, Glycation is the spontaneous non-enzymatic reaction wherein there is covalent attachment of free reducing sugars such as glucose, fructose, or their derivatives with the free amino groups of proteins, DNA, and lipids in order to form the advanced glycation end products (AGEs).  There is wide scientific evidence proving that the accumulation of AGEs is a vital factor in the progression of aging and age-related diseases such as chronic disease like diabetes, Alzheimer’s disease. Hence, strategies to inhibit the glycation process and to remove existing glycation products is a potential method in order to prolong the lifespan through reducing the AGE-mediated adverse health effects. In this sense, restricting and minimizing dietary AGEs, AGEs inhibitors and supplements, and physical exercise may be distinctly advantageous in reducing the burden of AGEs in the human body.

References: 

  1. Abate, G. , Delbarba, A., Marziano, M., Memo, M., and Uberti, D. 2015. Advanced Glycation End Products (AGEs) in Food: Focusing on Mediterranean Pasta. J Nutr Food Sci 5:6 DOI: 10.4172/2155-9600.1000440
  2. Brown, M. J. Healthline. 2019. What Are Advanced Glycation End Products (AGEs)?. Retrieved from: https://www.healthline.com/nutrition/advanced-glycation-end-products#how-much-is-too-much?. Retrieved on 17 June 2020.
  3. Chen, J., Lin, X., Bu, C. et al. Role of advanced glycation end products in mobility and considerations in possible dietary and nutritional intervention strategies. Nutr Metab (Lond) 15, 72 (2018). https://doi.org/10.1186/s12986-018-0306-7
  4. Glenn, J.; Stitt, A. 2009. The role of advanced glycation end products in retinal ageing and disease”. Biochimica et Biophysica Acta (BBA) – General Subjects. 1790 (10): 1109–1116. doi:10.1016/j.bbagen.2009.04.016.
  5. Kazeem MI, Akanji MA, Hafizur RM, Choudhary MI. Antiglycation, antioxidant and toxicological potential of polyphenol extracts of alligator pepper, ginger and nutmeg from Nigeria. Asian Pac J Trop Biomed. 2012;2(9):727-732. doi:10.1016/S2221-1691(12)60218-4
  6. Kim CS, Park S, Kim J. The role of glycation in the pathogenesis of aging and its prevention through herbal products and physical exercise. J Exerc Nutrition Biochem. 2017;21(3):55-61. doi:10.20463/jenb.2017.0027
  7. Semba, R. D.; Ferrucci, L.; Sun, K.; Beck, J.; Dalal, M.; Varadhan, R.; Walston, J.; Guralnik, J. M.; Fried, L. P. 2009. Advanced glycation end products and their circulating receptors predict cardiovascular disease mortality in older community-dwelling women. Aging Clinical and Experimental Research. 21 (2): 182–190. doi:10.1007/BF03325227
  8. Thomas MC, Baynes JW, Thorpe SR, Cooper ME. The role of AGEs and AGE inhibitors in diabetic cardiovascular disease. Curr Drug Targets. 2005;6(4):453-474. doi:10.2174/1389450054021873
  9. Uribarri J, Woodruff S, Goodman S, et al. Advanced glycation end products in foods and a practical guide to their reduction in the diet. J Am Diet Assoc. 2010;110(6):911-16.e12. doi:10.1016/j.jada.2010.03.018
  10. Yamagishi, S.‐i., Nakamura, K., Matsui, T., Ueda, S., Noda, Y. and Imaizumi, T. (2008), Inhibitors of Advanced Glycation End Products (AGEs): Potential Utility for the Treatment of Cardiovascular Disease. Cardiovascular Drug Reviews, 26: 50-58.
  11. Zanteson, L. 2014. Advanced Glycation End Products. Retrieved from: https://www.todaysdietitian.com/newarchives/030314p10.shtml. Retrieved on 17 June 2020.