Maillard reaction products in pet foods

Pet dogs and cats around the world are commonly fed processed commercial foods throughout their lives. Often heat treatments are used during the processing of these foods to improve nutrient digestibility, shelf life, and food safety. Processing is known to induce the Maillard reaction, in which a reducing sugar binds to a free reactive amino group of an amino acid. In intact proteins, the ε-amino group of lysine is the most abundant free amino group. The reaction reduces the bioavailability of lysine and results in the formation of advanced Maillard reaction products. The aim of this thesis was to determine the occurrence and progression of the Maillard reaction during the manufacturing of pet foods, the subsequent impact on nutritive value of the food, and the bioavailability of Maillard reaction products in cats. In Chapter 2, the scientific literature was reviewed to investigate the current state of knowledge on the Maillard reaction and its potential effect on the nutritive value of pet foods and on pet health. Determination of the difference between total and reactive lysine by chemical methods provides an indication of the Maillard reaction in pet foods. Studies reported that the proportion of reactive lysine is on average 73% (range 39 – 100%) of total lysine, and that foods for growing dogs may be at risk of supplying less lysine than the animals require. The endogenous analogues of Maillard reaction products, advanced glycation end-products, have been associated with age-related diseases in humans, such as diabetes and impaired renal function. In dogs, data indicate higher advanced glycation end-product contents in plasma from dogs suffering from canine diabetes mellitus compared with healthy control animals. In addition, elevated levels of advanced glycation end-products in tissue proteins in dogs were observed for a number of diseases. To date it was unknown to what extent Maillard reaction products were present in pet foods, and whether dietary Maillard reaction products can be associated with the development of diseases such as diabetes and impaired renal function in pet animals. As the Maillard reaction is induced by processing, changing processing conditions should have an influence on the severity of the reaction. However, effects of processing conditions on the difference in total and reactive lysine contents in pet foods were inconsistent and did not always correspond to model systems. Processing temperature was reported to be the most important factor followed by moisture level. In addition, differences between total and reactive lysine were observed in several ingredients commonly used in pet foods. Reviewing the literature indicates that it is unknown to which extent the Maillard reaction occurs and whether Maillard reaction products are present in pet foods. There might be a risk for certain foods not meeting minimal lysine requirements. It is also unknown what the exact effect of processing on the Maillard reaction is in pet foods. The experiment described in Chapter 3 was designed to evaluate whether commercial pet foods meet minimal lysine requirements. Sixty-seven extruded, canned and pelleted commercially available dog and cat foods formulated for growth and maintenance were analysed using conventional amino acid analysis and <em>O-</em>methylisourea as reagent for reactive lysine. Sixty out of the 67 foods in this study, regardless of the type of processing technology used, contained a lower reactive lysine than total lysine content. On average, pelleted and extruded foods contain lower reactive to total lysine ratios compared to canned foods (0.85, 0.89, and 0.93, respectively). All cat foods and foods for adult dogs met minimal lysine requirements. However, eight dry foods for growing dogs contained reactive lysine contents between 96 and 138% of the minimal lysine requirement, indicating that reactive lysine has to be between 62 and 104% digestible to meet minimal requirement. Considering the variability in reactive lysine digestibility, these foods could be at risk of not meeting minimal lysine requirements for growing dogs. In Chapter 4, the foods from Chapter 3 were used to quantitate the Maillard reaction products fructoselysine (FL), carboxymethyllysine (CML), hydroxymethylfurfural (HMF), and the cross-linked amino acid lysinoalanine (LAL) using UPLC-MS. In all foods, Maillard reaction products and LAL were found but in highly variable amounts. Type of processing seems to be a key factor for the concentration of FL, CML and HMF, with on average higher amounts in canned foods than pelleted and extruded foods (on a dry matter basis). The contents of CML and HMF found in commercial pet foods are, on average, within the range reported in processed human food products. Average daily intake (mg/kg body weight0.75) of HMF was 122 times higher for dogs and 38 times higher for cats than the calculated average intake for adult humans. Average daily intake of CML was comparable to the intake of adult humans. As Chapters 3 and 4 indicated that pelleted foods contain more Maillard reaction products than extruded foods, despite the less severe production process, an experiment was designed to gain insight in the effect of steam pelleting on the Maillard reaction in a dog food (Chapter 5). The aim was to examine the effect of conditioning temperature (65 and 90°C) and die hole length (ø 5 × 45, 65, and 80 mm) during pelleting processing of a standard dry dog food on selected indicators of the Maillard reaction (total lysine, reactive lysine, FL, CML, HMF, LAL), browning development and CIE-Lab colour. Steam pelleting did not cause a significant loss of reactive lysine and change of absorbance values. This indicates that the effect of steam pelleting on the nutritive value of the foods is low. However, steam pelleting did increase the content of Maillard reaction products. The formation of the Maillard reaction products was associated with an increase in temperature and die hole length during the steam pelleting process. The unprocessed ingredient mix already contained a larger difference between reactive and total lysine, and contents of Maillard reaction products than was induced during steam pelleting. Therefore, the choice of the ingredients used in this study mainly determines reactive lysine content and Maillard reaction products in the pet food formulation. As it is unknown to which extent extrusion processing influences the Maillard reaction in pet foods, the effect of extrusion processing on selected indicators of the Maillard reaction was determined (Chapter 6). The extrusion parameters temperature (140 and 165°C), moisture content (200 and 300 g/kg) and screw speed (100 and 200 rpm) were applied to two dry dog foods formulated using either intact or hydrolysed proteins. Extrusion processing in general results in a decrease in total and reactive lysine and an increase in FL, CML, HMF and LAL content. However, this effect appeared more pronounced in the diet containing hydrolysed protein. Decreasing temperature and moisture content led to higher total and reactive lysine contents, and less Maillard reaction products in the dog foods. Increasing screw speed had a positive influence on total and reactive lysine, but a negative influence on Maillard reaction products. As was found in Chapter 5, the unprocessed ingredient mixtures in this experiment contained already more Maillard reaction products than was induced during extrusion processing. Whether the Maillard reaction products reported in pet foods are physiologically relevant in pet animals depends on the bioavailability of these components. Therefore, urinary excretion was studied in adult cats fed commercial moist and dry foods containing varying amounts of FL, CML and the amino acid LAL (Chapter 7). A pilot study was first conducted to determine the adaptation time required for stable urinary excretion of the Maillard reaction products when changing diets with contrasting contents of Maillard reaction products. An adaptation time of 1 d was deemed sufficient in adult cats. The short adaptation time indicates an effective urinary excretion of Maillard reaction products. In the main study, six commercially processed dry and six moist diets were fed to 12 adult female cats in two parallel randomized, 36-day, balanced Latin square designs. Urine was collected quantitatively and FL, CML and LAL were analysed in foods and collected urine using HPLC-MS. Daily urinary excretion of FL and CML showed a positive relationship with daily intake in the dry and moist foods. For LAL, no significant relationship was observed. The observed increase in urinary excretion with increasing dietary intake indicates that dietary Maillard reaction products are absorbed from the gastro-intestinal tract of cats and excreted in the urine. Minimum apparent absorption based on urinary excretion (assuming 100% of the excreted component originates from the diet) of FL, CML and LAL was found to range between 8 to 23%, 25 to 73% and 6 to 19%, respectively. Urinary recovery (% ingested) showed a negative relationship with daily intake for FL, CML and LAL in the dry foods and for CML and LAL in the moist foods. The observed decrease in urinary recovery with increasing intake suggests a limiting factor in digestion, absorption, metabolism or urinary excretion. The studies reported in this thesis are one of the first to determine Maillard reaction products in pet foods and the bioavailability of FL, CML and LAL in cats. In addition, the results highlight the importance of reactive lysine measurement in foods for growing dogs used as weaning diets. Contribution of the absorption of dietary Maillard reaction products to the pathogenesis of various health conditions requires further study, as well as the potential role of restriction of dietary Maillard reaction products in prevention and treatment of long-term health implications. Extrusion and pelleting processing do increase the Maillard reaction, however, choice of ingredients appears to have a larger effect on the content of Maillard reaction products and can, therefore, be a useful strategy for pet food manufacturers that want to decrease the content of Maillard reaction products in their pet foods.