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Starch Characterization of Commercial Extruded Dry Pet Foods

Starch Characterization of Commercial Extruded Dry Pet Foods Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 I. Corsato Alvarenga & C. G. Aldrich Department of Grain Science and Industry, Kansas State University, Manhattan KS Corresponding author: aldrich4@ksu.edu Acknowledgements © The Author(s) 2020. Published by Oxford University Press on behalf of the American Society of Animal Science. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non- commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 We would like to acknowledge Dr. Michael Higgins at the Statistics Department at Kansas State University for statistical support. The authors have no conflicts of interest. ABSTRACT Starches provide an effective energy source for dogs and cats and can affect health according to its inclusion and extent of digestion. The starch fraction that escapes small intestine (SI) digestion is called resistant starch (RS) and is desirable due to its prebiotic function. Starch is not an essential nutrient for dogs and cats and thus is not reported on commercial pet food labels. Hence, the objective of this work was to characterize starches in commercial pet foods. The top five pet food companies by sales were selected to represent US pet foods, which were divided into four strata with a sampling frame of 654 foods: dog grain based (372 foods), dog grain free (71 foods), cat grain based (175 foods), and cat grain free (38 foods). Five random foods within each stratum were purchased (20 total). Starch analyses (total starch, resistant starch and starch cook), as well as nutrient analyses were conducted on all foods. Total starch, RS and starch cook means were compared using a 2-group Z-test on dog vs cat and grain-based vs grain-free diets, and differences were considered significant at a P < 0.05. Total starch was higher (P < 0.05) in dog than cat food, and starch cook was greater (P < 0.05) in grain-free diets. A regression analysis showed that NFE was a good predictor of total starch. Resistant starch was low and not different among groups. A post-hoc test showed that a total sample size of at least 28 diets per group would be required to detect differences in RS between GF and GB diets, if one exists. Key words Pet food, extrusion, grain-free, starch, gelatinized, resistant starch. Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 INTRODUCTION The US pet food industry is a growing market expected to exceed USD 30.01 billion by 2022 (Zion Market Research, 2017). Most dogs and cats are fed dry food (9.2 billion US dollars in sales in 2014; Statista, 2016), and the greatest part of it is produced through extrusion. This type of processing involves cooking with steam, water and shear. It also requires some amount of structure forming ingredients like starches (Guy, 2001) to promote food particle binding, texturization, improvement in palatability, and to aid in expansion of the kibble. Starch is not an essential nutrient for dogs and cats, but it can impact health in different ways according to its inclusion, type, and processing. The more cooked or gelatinized, the more rapidly the starch is digested (Murray et al., 2001). This has implications on metabolic utilization, and (or) the amount of starch that escapes digestion. Rapidly digested starches can promote high blood glucose/insulin peaks with subsequent fat deposition (Coulston et al., 1983). Conversely, the indigestible starch, or resistant starch (RS) can serve as substrate for colonic fermentation yielding Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 short-chain fatty acids of which butyrate can be used as a direct energy source for colonocytes (Bergman, 1990; Haenen et al., 2013). Starches may be inaccessible to digestive enzymes due to their tightly packed physical conformation, or physical barriers associated with the granule like cell walls and protein bodies (Dhital et al., 2017). There are differences in the digestion profile among starch sources for a number of reasons. For example, common cereals like corn, rice or wheat can have polyhedral and (or) oval starch granules which contain pores and channels that create adhesion sites for hydrolytic enzymes (Dhital et al., 2017). Some cereals like sorghum may be more difficult to digest than corn or rice due to tight bonding of protein bodies to the starch granule. Similarly, legume seeds are known to be high in naturally occurring RS, partly because their starches are trapped inside the cotyledon cell parenchyma (Berg et al., 2012; Würsch, 1986). Tuber starches like potato may also have some resistance to enzymatic digestion because its granules are large and smooth (Martens et al., 2018; Dhital et al., 2017). However, all these reports have been conducted with starch ingredients alone, but pet foods are composed of other ingredients which are ground, mixed and then cooked or processed in some manner. Due to morphological differences in starch ingredients used in pet foods and the interference of other ingredients and processing, it would be valuable to characterize these food starches in a complete food. There is no information required on pet food labels regarding starch percentage, extent of digestion, and (or) resistant starch concentration. Starch is not required nor allowed on the guaranteed analysis by current labeling regulation (AAFCO, 2019). Typical carbohydrate levels (starches and fibers) Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 in dry extruded dog foods range from 30-60%, while starches in commercial cat foods are included up to 35% on a DM basis (Gross et al., 2010). Nutritionists and pet food scientists commonly estimate starch content using the NFE (nitrogen-free extract) calculation (NFE= 100 – moisture – crude fiber – crude protein – crude fat – ash; Gross et al., 2010). This equation may overestimate starch content, as most of the nutrient analysis in pet foods are crude estimates of their true value and may not account for their total contribution. Knowing the true starch content of pet foods, and how much of it is digested, would be valuable information for diet development and future research. No research has been published previously characterizing and comparing the various starch components and methodologies of analysis in commercial complete pet foods. Further, there are no studies comparing the digestible starch and RS of grain-based foods and those containing elevated levels of tubers and legumes as their sole starch sources. Thus, the objective of this study was to determine the total starch content and its fractions (digestible and resistant starches) in dog and cat foods, and those that are grain-free (GF) and grain-based (GB) diets sold in the USA. The hypotheses were: 1. Dog foods would contain more starch than cat diets; 2. Extruded foods would be extensively cooked to a point that resistant starch would be almost nonexistent and thus insufficient to promote colonic health; and 3. GF diets would have more resistant starch in comparison to GB diets. MATERIALS AND METHODS Sample Selection Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 The top five pet food companies by sales (Pet Food Industry, 2017) were selected to represent the majority of US pet foods in this study. These companies, in decreasing order of sales, were: Mars Petcare, Nestlé Purina Petcare, Big Heart Pet brands, Hill’s Pet Nutrition and Blue Buffalo (Pet food Industry, 2019). A list of all dry complete extruded pet foods, excluding prescription diets, of the top five companies was created. Pet foods were divided into four strata with 654 foods composing the sampling frame: dog GB (372 foods), dog GF (71 foods), cat GB (175 foods), and cat GF (38 foods). Four lists with 10 random samples within each stratum were created using a randomization program. These lists were taken to pet stores in Manhattan, KS, and 5 foods present in the list within each stratum were purchased (20 total), according to store availability (Table 1). Grain based diets contained combinations of brewers rice (8 foods), brown rice (2 foods), corn (5 foods), wheat (3 foods), barley (4 foods), oats (4 foods), and some also included peas and potato starch. The GF foods had one ingredient or a combination of some of the following: peas (5 foods), pea starch (4 foods), sweet potatoes (4 foods), potatoes (4), potato starch (1 food), tapioca starch (3 foods), chickpeas (2 foods), and lentils (1 food). Nutrient Analysis All food samples were ground to 0.5 mm in a laboratory fixed blade impact mill (Retsch, type ZM200, Haan, Germany) prior to nutrient analyses. Ash (AOAC 942.05), nitrogen (AOAC 990.03; multiplied by 6.25 factor to estimate crude protein), fat by acid hydrolysis (AOAC 954.02), and total dietary fiber (TDF; TDF- 100A kit; Megazyme International Ireland Limited, Ireland) were measured on each sample in order to determine NFE, using the following calculation: NFE = 100(%)- ash(%) – moisture(%) – protein(%) – fat(%) – TDF(%). Total starch and starch fractions (resistant and digestible starches) were analyzed with enzymatic digestion Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 followed by colorimetric assays using kits (Total Starch Assay kit & Resistant Starch Assay kit, respectively; Megazyme International Ireland Limited, Ireland). Total starch was reported as both measured (using the total starch assay kit) and calculated (sum of digestible and resistant starches quantified by the resistant starch assay kit). Starch cook was analyzed by an enzymatic procedure as described by Mason et al. (1982). Briefly, two samples were prepared. One was boiled for 20 min with DI water. The second was equilibrated with DI water at 25ºC for 20 min. Then, buffer was added along with glucoamylase enzyme solution to both samples and they were incubated for 70 min at 40ºC. Free glucose in each sample was measured using a biochemistry analyzer (YSI 2900D, Xylem Analytics, Ohia, U.S.A.), and the level of gelatinization (%) calculated as a proportion of free glucose in the tested sample (gelatinized) to the free glucose in the boiled sample (total starch). Statistical Analysis The study was conducted using stratified random sampling. The averages of each analysis within each stratum were calculated according to Lohr (2009). Treatment means were compared using a 2 group Z-test with a significance level of α = 0.05. A regression analysis was conducted between dietary starch content measured by the total starch procedure (total starch measured) vs resistant starch procedure (total starch calculated = digestible + resistant starch), and NFE vs total starch measured, using the proc reg procedure of Statistical Analysis Software (SAS, v. 9.4; Cary, NC). RESULTS & DISCUSSION Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 The premise of this work was to characterize starch in commercial pet foods to gain some understanding of what is typical regarding starch components to aid further diet development, and to conduct future research in this area. Nutrients were measured in all diets in order to determine NFE, which is a common and rapid method to estimate starch content in diets. As expected, all nutrient levels met the specified guarantees identified on the label (information not shown). Nitrogen-free extract was calculated using TDF instead of crude fiber, which consists of a more accurate measurement of fibrous components in the food. The regression analysis between total starch measured by the total starch assay kit (Megazyme International Ireland Limited, Ireland) and NFE (P < .0001) was: 𝐹𝐸𝑁 = 1.04 × 𝑇𝑆 + 3.50 The adjusted R and standard error of this regression analysis were 0.94 and 0.0601, respectively. This indicates that NFE correlates well with total starch and thus it is a good estimation of starch content. Likewise, total starch calculated (TS ) was calc also highly correlated to total starch measured (TS): 𝑇 𝑆 = 0.944 × 𝑇𝑆 + 2.08; adjusted R =0.91, standard error= 0.0675. 𝑙𝑐𝑐𝑎 The first hypothesis stated that dog foods would contain more total starch than their feline counterparts, due to cats’ obligate carnivore nature and higher requirement for protein (NRC, 2006) which would result in a lower starch concentration in their diet. This was confirmed; wherein, total starch measured and calculated in cat diets were lower (P< 0.05) than dog diets (Table 2). The total starch (measured) difference between dog and cat foods, with 95% confidence, was estimated to be between 2.26% and 9.87% within the studied sampling frame. When grouping treatments as GB vs GF, total starch levels were not different. In the present study we found that commercial diets averaged above 87.5 % starch cook, and there was a difference (P < 0.05) in starch cook between GB and GF Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 diets (87.5 vs 94.1%, respectively). Tubers compose a large fraction of GF diets, and it was expected that tuber starches would have a greater degree of cook, since they have a higher water solubility index (Nuwamanya et al., 2011; Mishra and Rai, 2006) and gelatinize at a lower temperature than cereals (Mishra and Rai, 2006). Pezzali and Aldrich (2019) found that a GF dog food with a blend of tapioca starch, potato and peas required lower extruder thermal energy to produce kibbles with similar bulk density when compared to an ancient grain diet (composed of spelt, millet and sorghum), and the degree of starch cook of the GF treatment was also high and comparable to the present study (96.8% vs 94.1%, respectively). An important premise of this study was to determine the average level of RS in commercial diets. In order to compare starch fractions of different formula diets, digestible and resistant starches were calculated as a percentage of the total starch, so they would be on the same basis. The second hypothesis stated that commercial extruded diets would be low in RS, and indeed the RS levels of all commercial diets were observed to be less than 1% of the starch content (0.945 vs 0.703% resistant starch in dog vs cat diets, respectively; Table 1). This may not be sufficient RS to promote colonic health. Peixoto et al. (2018) were able to detect positive differences in colonic fermentation with 1.46% RS as a percent of total kibble weight, which increased butyrate production and improved nutrient absorption. Another beneficial effect from resistant starch is the reduction of the glycemic index of the food (Kimura; 2013), which decreases the rate of insulin release and positively impacts health. This can help reduce the incidence of obesity and type 2 diabetes. The third hypothesis was that GF would have more RS than GB diets. Tubers and legumes are common ingredients in GF diets, and they are known to have some resistance to α-amylase digestion due in part to low or absent starch granule pores, while most cereal starches have pores and channels that increase surface area for enzyme adsorption (Martens et al., 2018; Dhital et al., 2017). Also, most legumes Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 have a protein matrix tightly bonded with starch granules, which create a physical barrier to enzymatic digestion (Dhital et al., 2017; Berg et al., 2012). Moreover, type C starch present in legumes has a lower swelling capacity than cereals or tubers (Wani et al., 2016), and a higher amylose content (Martens et al., 2018), which contribute to enzymatic resistance. Hence, one would expect GF diets to be higher in RS than a GB recipe. However, in the present study there was no difference (P > 0.05) in RS between GB and GF diets (0.83 vs 1.06%, respectively). Although RS content was numerically greater in grain-free diets, the analytical technique employed has a high degree of variation when RS levels are below 2%. This high variability could influence the ability to detect differences. A retrospective power analysis (post- hoc) using RS as the endpoint of GF vs GB diets, showed that statistical analysis of the present study (using 10 diets as sample size) resulted in a power of only 0.52. This means that the study had a 52% probability to correctly lead to rejection of a false null hypothesis (RS in GB = RS in GF diets). In order to obtain a power of 0.80, with a significance level of 0.05, it would require 28 observations per treatment to detect some difference, if one exists. It is important to note that a statistical difference in dietary RS does not necessarily mean it would have biological significance in the animal. CONCLUSION In this study all the commercial diets tested had a very low RS level (close to or less than 1% of the total starch), which is less than what would be considered sufficient to promote colonic health. The level of starch cook did not reflect the amount of RS in the foods, possibly due to the analytical procedures themselves. When grouping treatments as GF vs GB diets, there was no difference in total starch and RS levels, whereas a difference in total starch content between dog and cat foods Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 was observed. Another important conclusion from this work was that regression analysis of NFE and total starch (calculated) showed that these were good predictors of total starch measured. Although an expanded and uniform kibble is aesthetically pleasing, a less expanded, denser kibble with less gelatinized starch might yield more RS. This work would suggest that different processing considerations than currently used in commercial products would be necessary to shift starch toward greater RS and thereby benefit colonic health. Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 REFERENCES AAFCO. 2019. American Association of Feed Control Officials. Model Regulations for Pet Food and Specialty Pet Food Under the Model Bill. In: Stan Cook, section editor. Association of American Feed Control Officials, Inc. pp 138- 202. Berg, T., Singh, J., Hardacre, A. and Boland, M. J., 2012. The role of cotyledon cell structure during in vitro digestion of starch in navy beans. Carbohydrate Polymers, 87(2):1678-1688. Bergman, E. N. 1990. Energy contributions of volatile fatty acids from the gastrointestinal tract in various species. Physiol Rev. 70:567–90. Cheftel, J. C., 1986. Nutritional effects of extrusion cooking. Food Chem. 20:263– Coulston, A. M., Liu, G. C. and Reaven, G. M., 1983. Plasma glucose, insulin and lipid responses to high-carbohydrate low-fat diets in normal humans. Metabolism, 32(1):52-56. Dhital, S., Warren, F. J., Butterworth, P. J., Ellis, P. R. and Gidley, M. J., 2017. Mechanisms of starch digestion by α-amylase—Structural basis for kinetic properties. Critical reviews in food science and nutrition, 57(5):875-892. Dust, J. M., Gajda, A. M., Flickinger, E. A., Burkhalter, T. M., Merchen, N. R., and Fahey, G. C. 2004. Extrusion Conditions Affect Chemical Composition and in Vitro Digestion of Select Food Ingredients. J. Agric. Food Chem., 52: 2989-2996. Gross, K. L., R. M. Yamka, C. Khoo, K. G. Friesen, D. E. Jewell, W. D. Schoenherr, J. Debraekeleer, and S. C. Zicker. 2010. Macronutrients. In: M. S. Hand, C. D. Thatcher, R. L. Remillard, P. Roudebusch, B. J. Novotny, editors, Small th Animal Clinical Nutrition 5 edition. Topeka, KS. P. 49-106. Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 Haenen, D., Zhang, J., Souza da Silva, C., Bosch, G., van der Meer, I. M., van Arkel, J., van den Borne, J. J., Pérez Gutiérrez, O., Smidt, H., Kemp, B. and Müller, M. 2013. A Diet High in Resistant Starch Modulates Microbiota Composition, SCFA Concentrations, and Gene Expression in Pig Intestine– 3. The Journal of nutrition, 143(3):274-283. Lohr, S. 2009. Stratified random sampling. In: M. Julet, editor, Sampling: Design and Analysis. Brooks Cole, Boston, MA. p. 73- 100. Martens, B.M., Gerrits, W.J., Bruininx, E.M. and Schols, H.A., 2018. Amylopectin structure and crystallinity explains variation in digestion kinetics of starches across botanic sources in an in vitro pig model. Journal of animal science and biotechnology, 9(1), p.91. Mason, M., Gleason, B., Rokey, G., 1982. A New Method for Determining Degree of Cook. In: American Association of Cereal Chemists 67th Annual Meeting. San Antonio, TX, USA. p. 123-124. Mishra, S. and Rai, T. 2006. Morphology and functional properties of corn, potato and tapioca starches. Food Hydrocoll. 20(5):557-566. doi.org/10.1016/j.foodhyd.2005.01.001. Murray, S. M., Flickinger, E. A., Patil, A. R., Merchen, N. R., Brent Jr, J. L. and Fahey Jr, G. C., 2001. In vitro fermentation characteristics of native and processed cereal grains and potato starch using ileal chyme from dogs. Journal of animal science, 79(2):435-444. NRC. 2006. Nutrient requirements of dogs and cats. National Academies Press, Washington, DC. Nuwamanya, E., Baguma, Y., Wembabazi, E. and Rubaihayo, P. 2011. A comparative study of the physicochemical properties of starches from root, tuber and cereal crops. African Journal of Biotechnology, 10(56):12018- Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 Pet Food Industry, 2019. Top Pet Food Companies Current Data. https://www.petfoodindustry.com/directories/211-top-pet-food-companies- current-data (Accessed 27 August 2019.) Peixoto, M. C., Ribeiro, É. M., Maria, A. P. J., Loureiro, B. A., di Santo, L. G., Putarov, T.C., Yoshitoshi, F. N., Pereira, G. T., Sá, L. R. M. and Carciofi, A. C., 2018. Effect of resistant starch on the intestinal health of old dogs: fermentation products and histological features of the intestinal mucosa. Journal of animal physiology and animal nutrition, 102(1):111-121. Pezzali, J. G. and Aldrich, C. G. 2019. Effect of Ancient Grains and Grain-free Carbohydrate Sources on Extrusion Parameters and Nutrient Utilization by Dogs. Journal of animal science, 13 July 2019. In-press. Wani, I. A., Sogi, D. S., Hamdani, A. M., Gani, A., Bhat, N. A. and Shah, A. 2016. Isolation, composition, and physicochemical properties of starch from legumes: A review. Starch‐ Stärke, 68(9-10):834-845. Yadav, B. S., Sharma, A. and Yadav, R. B., 2010. Resistant starch content of conventionally boiled and pressure-cooked cereals, legumes and tubers. Journal of food science and technology, 47(1):84-88. Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 Table 1 Nutritional composition of the commercial diets used in the study. Foo Speci Categor Crud Fat, Ash TDF Star Moistu NF 2 3 d es y e acid ch re E Prote hydroly in sis 1 Dog GB 10.3 28.8 16.6 7.75 4 24.4 6.15 30.4 2 Dog GB 27.1 10.5 9.20 9.02 32.5 6.82 37.3 3 Dog GB 29.9 12.9 7.24 6.10 27.9 6.31 37.6 4 Dog GB 29.2 16.7 7.42 7.43 26.9 5.52 33.7 5 Dog GB 26.2 15.7 6.66 8.99 30.1 6.22 36.3 6 Dog GF 23.7 14.3 8.46 9.88 28.3 6.20 37.5 7 Dog GF 11.4 22.5 13.3 6.02 6 32.9 6.35 40.3 8 Dog GF 10.1 31.0 16.4 7.83 1 22.7 6.30 28.3 9 Dog GF 22.7 10.7 8.81 8.05 37.5 6.28 43.5 10 Dog GF 11.8 33.2 20.0 7.21 3 16.2 5.99 21.7 11 Cat GB 36.7 17.9 7.03 6.54 23.9 4.04 27.8 12 Cat GB 10.5 31.9 13.0 7.05 9 25.8 6.06 31.4 13 Cat GB 10.2 33.3 10.8 7.08 0 27.3 4.89 33.7 14 Cat GB 13.5 36.2 11.2 5.86 6 19.9 6.07 27.1 15 Cat GB 10.9 36.7 15.2 8.24 0 19.4 5.59 23.4 16 Cat GF 43.8 16.7 7.93 7.92 15.8 4.25 19.4 17 Cat GF 11.7 33.8 11.5 7.04 3 22.0 5.81 30.1 18 Cat GF 37.3 15.5 7.88 7.88 21.1 5.74 25.7 19 Cat GF 41.4 14.4 7.03 8.18 20.9 4.62 24.4 20 Cat GF 33.5 14.1 8.23 9.90 22.7 5.63 28.7 All nutrients reported on a percentage as-is basis. GB = grain based; GF= grain-free. NFE was the only calculated component. Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 Table 2. Total, digestible and resistant starches of dog vs cat diets, and grain-based (GB) vs grain-free (GF) diets. Item, % Dog Cat SEM T P-value n= 10 n= 10 Total starch, 30.1 24.0 1.94 3.1218 0.0018 measured Total starch, 31.4 25.1 2.16 2.9122 0.0036 calculated Resistant 0.945 0.703 0.2212 1.0950 0.2735 starch Digestible 99.0 99.3 0.22 1.1418 0.2535 starch Starch 88.3 89.2 2.51 0.3498 0.7265 cook Item, % GB GF SEM T P-value n= 10 n= 10 Total starch, 28.4 26.7 1.14 0.6361 0.5247 measured Total starch, 29.9 26.4 1.52 1.6016 0.1092 calculated Resistant 0.828 1.062 0.1602 0.6360 0.5248 starch Digestible 99.2 98.9 0.16 0.9158 0.3598 starch Starch 87.5 94.1 1.46 3.9030 <.0001 cook Resistant and digestible starches were calculated as percentages of the total starch. Starch cook calculations were based on total starch and starch gelatinized measured at a commercial laboratory (Wenger Technical Center; Wenger Mfg., Sabetha KS, USA). Accepted Manuscript http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Translational Animal Science Oxford University Press

Starch Characterization of Commercial Extruded Dry Pet Foods

Corsato Alvarenga,, I; Aldrich, C, G
Translational Animal Science , Volume Advance Article – Apr 1, 2020
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Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 I. Corsato Alvarenga & C. G. Aldrich Department of Grain Science and Industry, Kansas State University, Manhattan KS Corresponding author: aldrich4@ksu.edu Acknowledgements © The Author(s) 2020. Published by Oxford University Press on behalf of the American Society of Animal Science. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non- commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 We would like to acknowledge Dr. Michael Higgins at the Statistics Department at Kansas State University for statistical support. The authors have no conflicts of interest. ABSTRACT Starches provide an effective energy source for dogs and cats and can affect health according to its inclusion and extent of digestion. The starch fraction that escapes small intestine (SI) digestion is called resistant starch (RS) and is desirable due to its prebiotic function. Starch is not an essential nutrient for dogs and cats and thus is not reported on commercial pet food labels. Hence, the objective of this work was to characterize starches in commercial pet foods. The top five pet food companies by sales were selected to represent US pet foods, which were divided into four strata with a sampling frame of 654 foods: dog grain based (372 foods), dog grain free (71 foods), cat grain based (175 foods), and cat grain free (38 foods). Five random foods within each stratum were purchased (20 total). Starch analyses (total starch, resistant starch and starch cook), as well as nutrient analyses were conducted on all foods. Total starch, RS and starch cook means were compared using a 2-group Z-test on dog vs cat and grain-based vs grain-free diets, and differences were considered significant at a P < 0.05. Total starch was higher (P < 0.05) in dog than cat food, and starch cook was greater (P < 0.05) in grain-free diets. A regression analysis showed that NFE was a good predictor of total starch. Resistant starch was low and not different among groups. A post-hoc test showed that a total sample size of at least 28 diets per group would be required to detect differences in RS between GF and GB diets, if one exists. Key words Pet food, extrusion, grain-free, starch, gelatinized, resistant starch. Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 INTRODUCTION The US pet food industry is a growing market expected to exceed USD 30.01 billion by 2022 (Zion Market Research, 2017). Most dogs and cats are fed dry food (9.2 billion US dollars in sales in 2014; Statista, 2016), and the greatest part of it is produced through extrusion. This type of processing involves cooking with steam, water and shear. It also requires some amount of structure forming ingredients like starches (Guy, 2001) to promote food particle binding, texturization, improvement in palatability, and to aid in expansion of the kibble. Starch is not an essential nutrient for dogs and cats, but it can impact health in different ways according to its inclusion, type, and processing. The more cooked or gelatinized, the more rapidly the starch is digested (Murray et al., 2001). This has implications on metabolic utilization, and (or) the amount of starch that escapes digestion. Rapidly digested starches can promote high blood glucose/insulin peaks with subsequent fat deposition (Coulston et al., 1983). Conversely, the indigestible starch, or resistant starch (RS) can serve as substrate for colonic fermentation yielding Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 short-chain fatty acids of which butyrate can be used as a direct energy source for colonocytes (Bergman, 1990; Haenen et al., 2013). Starches may be inaccessible to digestive enzymes due to their tightly packed physical conformation, or physical barriers associated with the granule like cell walls and protein bodies (Dhital et al., 2017). There are differences in the digestion profile among starch sources for a number of reasons. For example, common cereals like corn, rice or wheat can have polyhedral and (or) oval starch granules which contain pores and channels that create adhesion sites for hydrolytic enzymes (Dhital et al., 2017). Some cereals like sorghum may be more difficult to digest than corn or rice due to tight bonding of protein bodies to the starch granule. Similarly, legume seeds are known to be high in naturally occurring RS, partly because their starches are trapped inside the cotyledon cell parenchyma (Berg et al., 2012; Würsch, 1986). Tuber starches like potato may also have some resistance to enzymatic digestion because its granules are large and smooth (Martens et al., 2018; Dhital et al., 2017). However, all these reports have been conducted with starch ingredients alone, but pet foods are composed of other ingredients which are ground, mixed and then cooked or processed in some manner. Due to morphological differences in starch ingredients used in pet foods and the interference of other ingredients and processing, it would be valuable to characterize these food starches in a complete food. There is no information required on pet food labels regarding starch percentage, extent of digestion, and (or) resistant starch concentration. Starch is not required nor allowed on the guaranteed analysis by current labeling regulation (AAFCO, 2019). Typical carbohydrate levels (starches and fibers) Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 in dry extruded dog foods range from 30-60%, while starches in commercial cat foods are included up to 35% on a DM basis (Gross et al., 2010). Nutritionists and pet food scientists commonly estimate starch content using the NFE (nitrogen-free extract) calculation (NFE= 100 – moisture – crude fiber – crude protein – crude fat – ash; Gross et al., 2010). This equation may overestimate starch content, as most of the nutrient analysis in pet foods are crude estimates of their true value and may not account for their total contribution. Knowing the true starch content of pet foods, and how much of it is digested, would be valuable information for diet development and future research. No research has been published previously characterizing and comparing the various starch components and methodologies of analysis in commercial complete pet foods. Further, there are no studies comparing the digestible starch and RS of grain-based foods and those containing elevated levels of tubers and legumes as their sole starch sources. Thus, the objective of this study was to determine the total starch content and its fractions (digestible and resistant starches) in dog and cat foods, and those that are grain-free (GF) and grain-based (GB) diets sold in the USA. The hypotheses were: 1. Dog foods would contain more starch than cat diets; 2. Extruded foods would be extensively cooked to a point that resistant starch would be almost nonexistent and thus insufficient to promote colonic health; and 3. GF diets would have more resistant starch in comparison to GB diets. MATERIALS AND METHODS Sample Selection Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 The top five pet food companies by sales (Pet Food Industry, 2017) were selected to represent the majority of US pet foods in this study. These companies, in decreasing order of sales, were: Mars Petcare, Nestlé Purina Petcare, Big Heart Pet brands, Hill’s Pet Nutrition and Blue Buffalo (Pet food Industry, 2019). A list of all dry complete extruded pet foods, excluding prescription diets, of the top five companies was created. Pet foods were divided into four strata with 654 foods composing the sampling frame: dog GB (372 foods), dog GF (71 foods), cat GB (175 foods), and cat GF (38 foods). Four lists with 10 random samples within each stratum were created using a randomization program. These lists were taken to pet stores in Manhattan, KS, and 5 foods present in the list within each stratum were purchased (20 total), according to store availability (Table 1). Grain based diets contained combinations of brewers rice (8 foods), brown rice (2 foods), corn (5 foods), wheat (3 foods), barley (4 foods), oats (4 foods), and some also included peas and potato starch. The GF foods had one ingredient or a combination of some of the following: peas (5 foods), pea starch (4 foods), sweet potatoes (4 foods), potatoes (4), potato starch (1 food), tapioca starch (3 foods), chickpeas (2 foods), and lentils (1 food). Nutrient Analysis All food samples were ground to 0.5 mm in a laboratory fixed blade impact mill (Retsch, type ZM200, Haan, Germany) prior to nutrient analyses. Ash (AOAC 942.05), nitrogen (AOAC 990.03; multiplied by 6.25 factor to estimate crude protein), fat by acid hydrolysis (AOAC 954.02), and total dietary fiber (TDF; TDF- 100A kit; Megazyme International Ireland Limited, Ireland) were measured on each sample in order to determine NFE, using the following calculation: NFE = 100(%)- ash(%) – moisture(%) – protein(%) – fat(%) – TDF(%). Total starch and starch fractions (resistant and digestible starches) were analyzed with enzymatic digestion Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 followed by colorimetric assays using kits (Total Starch Assay kit & Resistant Starch Assay kit, respectively; Megazyme International Ireland Limited, Ireland). Total starch was reported as both measured (using the total starch assay kit) and calculated (sum of digestible and resistant starches quantified by the resistant starch assay kit). Starch cook was analyzed by an enzymatic procedure as described by Mason et al. (1982). Briefly, two samples were prepared. One was boiled for 20 min with DI water. The second was equilibrated with DI water at 25ºC for 20 min. Then, buffer was added along with glucoamylase enzyme solution to both samples and they were incubated for 70 min at 40ºC. Free glucose in each sample was measured using a biochemistry analyzer (YSI 2900D, Xylem Analytics, Ohia, U.S.A.), and the level of gelatinization (%) calculated as a proportion of free glucose in the tested sample (gelatinized) to the free glucose in the boiled sample (total starch). Statistical Analysis The study was conducted using stratified random sampling. The averages of each analysis within each stratum were calculated according to Lohr (2009). Treatment means were compared using a 2 group Z-test with a significance level of α = 0.05. A regression analysis was conducted between dietary starch content measured by the total starch procedure (total starch measured) vs resistant starch procedure (total starch calculated = digestible + resistant starch), and NFE vs total starch measured, using the proc reg procedure of Statistical Analysis Software (SAS, v. 9.4; Cary, NC). RESULTS & DISCUSSION Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 The premise of this work was to characterize starch in commercial pet foods to gain some understanding of what is typical regarding starch components to aid further diet development, and to conduct future research in this area. Nutrients were measured in all diets in order to determine NFE, which is a common and rapid method to estimate starch content in diets. As expected, all nutrient levels met the specified guarantees identified on the label (information not shown). Nitrogen-free extract was calculated using TDF instead of crude fiber, which consists of a more accurate measurement of fibrous components in the food. The regression analysis between total starch measured by the total starch assay kit (Megazyme International Ireland Limited, Ireland) and NFE (P < .0001) was: 𝐹𝐸𝑁 = 1.04 × 𝑇𝑆 + 3.50 The adjusted R and standard error of this regression analysis were 0.94 and 0.0601, respectively. This indicates that NFE correlates well with total starch and thus it is a good estimation of starch content. Likewise, total starch calculated (TS ) was calc also highly correlated to total starch measured (TS): 𝑇 𝑆 = 0.944 × 𝑇𝑆 + 2.08; adjusted R =0.91, standard error= 0.0675. 𝑙𝑐𝑐𝑎 The first hypothesis stated that dog foods would contain more total starch than their feline counterparts, due to cats’ obligate carnivore nature and higher requirement for protein (NRC, 2006) which would result in a lower starch concentration in their diet. This was confirmed; wherein, total starch measured and calculated in cat diets were lower (P< 0.05) than dog diets (Table 2). The total starch (measured) difference between dog and cat foods, with 95% confidence, was estimated to be between 2.26% and 9.87% within the studied sampling frame. When grouping treatments as GB vs GF, total starch levels were not different. In the present study we found that commercial diets averaged above 87.5 % starch cook, and there was a difference (P < 0.05) in starch cook between GB and GF Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 diets (87.5 vs 94.1%, respectively). Tubers compose a large fraction of GF diets, and it was expected that tuber starches would have a greater degree of cook, since they have a higher water solubility index (Nuwamanya et al., 2011; Mishra and Rai, 2006) and gelatinize at a lower temperature than cereals (Mishra and Rai, 2006). Pezzali and Aldrich (2019) found that a GF dog food with a blend of tapioca starch, potato and peas required lower extruder thermal energy to produce kibbles with similar bulk density when compared to an ancient grain diet (composed of spelt, millet and sorghum), and the degree of starch cook of the GF treatment was also high and comparable to the present study (96.8% vs 94.1%, respectively). An important premise of this study was to determine the average level of RS in commercial diets. In order to compare starch fractions of different formula diets, digestible and resistant starches were calculated as a percentage of the total starch, so they would be on the same basis. The second hypothesis stated that commercial extruded diets would be low in RS, and indeed the RS levels of all commercial diets were observed to be less than 1% of the starch content (0.945 vs 0.703% resistant starch in dog vs cat diets, respectively; Table 1). This may not be sufficient RS to promote colonic health. Peixoto et al. (2018) were able to detect positive differences in colonic fermentation with 1.46% RS as a percent of total kibble weight, which increased butyrate production and improved nutrient absorption. Another beneficial effect from resistant starch is the reduction of the glycemic index of the food (Kimura; 2013), which decreases the rate of insulin release and positively impacts health. This can help reduce the incidence of obesity and type 2 diabetes. The third hypothesis was that GF would have more RS than GB diets. Tubers and legumes are common ingredients in GF diets, and they are known to have some resistance to α-amylase digestion due in part to low or absent starch granule pores, while most cereal starches have pores and channels that increase surface area for enzyme adsorption (Martens et al., 2018; Dhital et al., 2017). Also, most legumes Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 have a protein matrix tightly bonded with starch granules, which create a physical barrier to enzymatic digestion (Dhital et al., 2017; Berg et al., 2012). Moreover, type C starch present in legumes has a lower swelling capacity than cereals or tubers (Wani et al., 2016), and a higher amylose content (Martens et al., 2018), which contribute to enzymatic resistance. Hence, one would expect GF diets to be higher in RS than a GB recipe. However, in the present study there was no difference (P > 0.05) in RS between GB and GF diets (0.83 vs 1.06%, respectively). Although RS content was numerically greater in grain-free diets, the analytical technique employed has a high degree of variation when RS levels are below 2%. This high variability could influence the ability to detect differences. A retrospective power analysis (post- hoc) using RS as the endpoint of GF vs GB diets, showed that statistical analysis of the present study (using 10 diets as sample size) resulted in a power of only 0.52. This means that the study had a 52% probability to correctly lead to rejection of a false null hypothesis (RS in GB = RS in GF diets). In order to obtain a power of 0.80, with a significance level of 0.05, it would require 28 observations per treatment to detect some difference, if one exists. It is important to note that a statistical difference in dietary RS does not necessarily mean it would have biological significance in the animal. CONCLUSION In this study all the commercial diets tested had a very low RS level (close to or less than 1% of the total starch), which is less than what would be considered sufficient to promote colonic health. The level of starch cook did not reflect the amount of RS in the foods, possibly due to the analytical procedures themselves. When grouping treatments as GF vs GB diets, there was no difference in total starch and RS levels, whereas a difference in total starch content between dog and cat foods Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 was observed. Another important conclusion from this work was that regression analysis of NFE and total starch (calculated) showed that these were good predictors of total starch measured. Although an expanded and uniform kibble is aesthetically pleasing, a less expanded, denser kibble with less gelatinized starch might yield more RS. This work would suggest that different processing considerations than currently used in commercial products would be necessary to shift starch toward greater RS and thereby benefit colonic health. Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 REFERENCES AAFCO. 2019. American Association of Feed Control Officials. Model Regulations for Pet Food and Specialty Pet Food Under the Model Bill. In: Stan Cook, section editor. Association of American Feed Control Officials, Inc. pp 138- 202. Berg, T., Singh, J., Hardacre, A. and Boland, M. J., 2012. The role of cotyledon cell structure during in vitro digestion of starch in navy beans. Carbohydrate Polymers, 87(2):1678-1688. Bergman, E. N. 1990. Energy contributions of volatile fatty acids from the gastrointestinal tract in various species. Physiol Rev. 70:567–90. Cheftel, J. C., 1986. Nutritional effects of extrusion cooking. Food Chem. 20:263– Coulston, A. M., Liu, G. C. and Reaven, G. M., 1983. Plasma glucose, insulin and lipid responses to high-carbohydrate low-fat diets in normal humans. Metabolism, 32(1):52-56. Dhital, S., Warren, F. J., Butterworth, P. J., Ellis, P. R. and Gidley, M. J., 2017. Mechanisms of starch digestion by α-amylase—Structural basis for kinetic properties. Critical reviews in food science and nutrition, 57(5):875-892. Dust, J. M., Gajda, A. M., Flickinger, E. A., Burkhalter, T. M., Merchen, N. R., and Fahey, G. C. 2004. Extrusion Conditions Affect Chemical Composition and in Vitro Digestion of Select Food Ingredients. J. Agric. Food Chem., 52: 2989-2996. Gross, K. L., R. M. Yamka, C. Khoo, K. G. Friesen, D. E. Jewell, W. D. Schoenherr, J. Debraekeleer, and S. C. Zicker. 2010. Macronutrients. In: M. S. Hand, C. D. Thatcher, R. L. Remillard, P. Roudebusch, B. J. Novotny, editors, Small th Animal Clinical Nutrition 5 edition. Topeka, KS. P. 49-106. Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 Haenen, D., Zhang, J., Souza da Silva, C., Bosch, G., van der Meer, I. M., van Arkel, J., van den Borne, J. J., Pérez Gutiérrez, O., Smidt, H., Kemp, B. and Müller, M. 2013. A Diet High in Resistant Starch Modulates Microbiota Composition, SCFA Concentrations, and Gene Expression in Pig Intestine– 3. The Journal of nutrition, 143(3):274-283. Lohr, S. 2009. Stratified random sampling. In: M. Julet, editor, Sampling: Design and Analysis. Brooks Cole, Boston, MA. p. 73- 100. Martens, B.M., Gerrits, W.J., Bruininx, E.M. and Schols, H.A., 2018. Amylopectin structure and crystallinity explains variation in digestion kinetics of starches across botanic sources in an in vitro pig model. Journal of animal science and biotechnology, 9(1), p.91. Mason, M., Gleason, B., Rokey, G., 1982. A New Method for Determining Degree of Cook. In: American Association of Cereal Chemists 67th Annual Meeting. San Antonio, TX, USA. p. 123-124. Mishra, S. and Rai, T. 2006. Morphology and functional properties of corn, potato and tapioca starches. Food Hydrocoll. 20(5):557-566. doi.org/10.1016/j.foodhyd.2005.01.001. Murray, S. M., Flickinger, E. A., Patil, A. R., Merchen, N. R., Brent Jr, J. L. and Fahey Jr, G. C., 2001. In vitro fermentation characteristics of native and processed cereal grains and potato starch using ileal chyme from dogs. Journal of animal science, 79(2):435-444. NRC. 2006. Nutrient requirements of dogs and cats. National Academies Press, Washington, DC. Nuwamanya, E., Baguma, Y., Wembabazi, E. and Rubaihayo, P. 2011. A comparative study of the physicochemical properties of starches from root, tuber and cereal crops. African Journal of Biotechnology, 10(56):12018- Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 Pet Food Industry, 2019. Top Pet Food Companies Current Data. https://www.petfoodindustry.com/directories/211-top-pet-food-companies- current-data (Accessed 27 August 2019.) Peixoto, M. C., Ribeiro, É. M., Maria, A. P. J., Loureiro, B. A., di Santo, L. G., Putarov, T.C., Yoshitoshi, F. N., Pereira, G. T., Sá, L. R. M. and Carciofi, A. C., 2018. Effect of resistant starch on the intestinal health of old dogs: fermentation products and histological features of the intestinal mucosa. Journal of animal physiology and animal nutrition, 102(1):111-121. Pezzali, J. G. and Aldrich, C. G. 2019. Effect of Ancient Grains and Grain-free Carbohydrate Sources on Extrusion Parameters and Nutrient Utilization by Dogs. Journal of animal science, 13 July 2019. In-press. Wani, I. A., Sogi, D. S., Hamdani, A. M., Gani, A., Bhat, N. A. and Shah, A. 2016. Isolation, composition, and physicochemical properties of starch from legumes: A review. Starch‐ Stärke, 68(9-10):834-845. Yadav, B. S., Sharma, A. and Yadav, R. B., 2010. Resistant starch content of conventionally boiled and pressure-cooked cereals, legumes and tubers. Journal of food science and technology, 47(1):84-88. Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 Table 1 Nutritional composition of the commercial diets used in the study. Foo Speci Categor Crud Fat, Ash TDF Star Moistu NF 2 3 d es y e acid ch re E Prote hydroly in sis 1 Dog GB 10.3 28.8 16.6 7.75 4 24.4 6.15 30.4 2 Dog GB 27.1 10.5 9.20 9.02 32.5 6.82 37.3 3 Dog GB 29.9 12.9 7.24 6.10 27.9 6.31 37.6 4 Dog GB 29.2 16.7 7.42 7.43 26.9 5.52 33.7 5 Dog GB 26.2 15.7 6.66 8.99 30.1 6.22 36.3 6 Dog GF 23.7 14.3 8.46 9.88 28.3 6.20 37.5 7 Dog GF 11.4 22.5 13.3 6.02 6 32.9 6.35 40.3 8 Dog GF 10.1 31.0 16.4 7.83 1 22.7 6.30 28.3 9 Dog GF 22.7 10.7 8.81 8.05 37.5 6.28 43.5 10 Dog GF 11.8 33.2 20.0 7.21 3 16.2 5.99 21.7 11 Cat GB 36.7 17.9 7.03 6.54 23.9 4.04 27.8 12 Cat GB 10.5 31.9 13.0 7.05 9 25.8 6.06 31.4 13 Cat GB 10.2 33.3 10.8 7.08 0 27.3 4.89 33.7 14 Cat GB 13.5 36.2 11.2 5.86 6 19.9 6.07 27.1 15 Cat GB 10.9 36.7 15.2 8.24 0 19.4 5.59 23.4 16 Cat GF 43.8 16.7 7.93 7.92 15.8 4.25 19.4 17 Cat GF 11.7 33.8 11.5 7.04 3 22.0 5.81 30.1 18 Cat GF 37.3 15.5 7.88 7.88 21.1 5.74 25.7 19 Cat GF 41.4 14.4 7.03 8.18 20.9 4.62 24.4 20 Cat GF 33.5 14.1 8.23 9.90 22.7 5.63 28.7 All nutrients reported on a percentage as-is basis. GB = grain based; GF= grain-free. NFE was the only calculated component. Accepted Manuscript Downloaded from https://academic.oup.com/tas/advance-article-abstract/doi/10.1093/tas/txaa018/5735650 by guest on 18 February 2020 Table 2. Total, digestible and resistant starches of dog vs cat diets, and grain-based (GB) vs grain-free (GF) diets. Item, % Dog Cat SEM T P-value n= 10 n= 10 Total starch, 30.1 24.0 1.94 3.1218 0.0018 measured Total starch, 31.4 25.1 2.16 2.9122 0.0036 calculated Resistant 0.945 0.703 0.2212 1.0950 0.2735 starch Digestible 99.0 99.3 0.22 1.1418 0.2535 starch Starch 88.3 89.2 2.51 0.3498 0.7265 cook Item, % GB GF SEM T P-value n= 10 n= 10 Total starch, 28.4 26.7 1.14 0.6361 0.5247 measured Total starch, 29.9 26.4 1.52 1.6016 0.1092 calculated Resistant 0.828 1.062 0.1602 0.6360 0.5248 starch Digestible 99.2 98.9 0.16 0.9158 0.3598 starch Starch 87.5 94.1 1.46 3.9030 <.0001 cook Resistant and digestible starches were calculated as percentages of the total starch. Starch cook calculations were based on total starch and starch gelatinized measured at a commercial laboratory (Wenger Technical Center; Wenger Mfg., Sabetha KS, USA). Accepted Manuscript

Journal

Translational Animal Science – Oxford University Press

Published: Apr 1, 2020

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