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Biomarkers of food intake for nuts and vegetable oils : an extensive literature search
Garcia-Aloy, Mar ; Hulshof, Paul J.M. ; Estruel-Amades, Sheila ; Osté, Maryse C.J. ; Lankinen, Maria ; Geleijnse, Johanna M. ; Goede, Janette De; Ulaszewska, Marynka ; Mattivi, Fulvio ; Bakker, Stephan J.L. ; Schwab, Ursula ; Andres-Lacueva, Cristina - \ 2019
Genes & Nutrition 14 (2019)1. - ISSN 1555-8932
Biomarker - Intake - Metabolomics - Nuts - Oils

Nuts and vegetable oils are important sources of fat and of a wide variety of micronutrients and phytochemicals. Following their intake, several of their constituents, as well as their derived metabolites, are found in blood circulation and in urine. As a consequence, these could be used to assess the compliance to a dietary intervention or to determine habitual intake of nuts and vegetable oils. However, before these metabolites can be widely used as biomarkers of food intake (BFIs), several characteristics have to be considered, including specificity, dose response, time response, stability, and analytical performance. We have, therefore, conducted an extensive literature search to evaluate current knowledge about potential BFIs of nuts and vegetable oils. Once identified, the strengths and weaknesses of the most promising candidate BFIs have been summarized. Results from selected studies have provided a variety of compounds mainly derived from the fatty fraction of these foods, but also other components and derived metabolites related to their nutritional composition. In particular, α-linolenic acid, urolithins, and 5-hydroxyindole-3-acetic acid seem to be the most plausible candidate BFIs for walnuts, whereas for almonds they could be α-tocopherol and some catechin-derived metabolites. Similarly, several studies have reported a strong association between selenium levels and consumption of Brazil nuts. Intake of vegetable oils has been mainly assessed through the measurement of specific fatty acids in different blood fractions, such as oleic acid for olive oil, α-linolenic acid for flaxseed (linseed) and rapeseed (canola) oils, and linoleic acid for sunflower oil. Additionally, hydroxytyrosol and its metabolites were the most promising distinctive BFIs for (extra) virgin olive oil. However, most of these components lack sufficient specificity to serve as BFIs. Therefore, additional studies are necessary to discover new candidate BFIs, as well as to further evaluate the specificity, sensitivity, dose-response relationships, and reproducibility of these candidate biomarkers and to eventually validate them in other populations. For the discovery of new candidate BFIs, an untargeted metabolomics approach may be the most effective strategy, whereas for increasing the specificity of the evaluation of food consumption, this could be a combination of different metabolites.

Loes Moor-Hulshof, operationeel directeur de vegetarische slager: 'Ik wil ervoor zorgen dat de bio-industrie straks niet meer nodig is'
Goot, Atze Jan van der - \ 2019
‘I want to make sure factory farming is soon superfluous’: Loes Moor-Hulshof, operational director af The Vegetarian Butcher
Goot, Atze Jan van der - \ 2019
Food Composition Tables in Southeast Asia : The Contribution of the SMILING Project
Hulshof, Paul ; Doets, Esmee ; Seyha, Sok ; Bunthang, Touch ; Vonglokham, Manithong ; Kounnavong, Sengchanh ; Famida, Umi ; Muslimatun, Siti ; Santika, Otte ; Prihatini, Sri ; Nazarudin, Nazarina ; Jahari, Abas ; Rojroongwasinkul, Nipa ; Chittchang, Uraiporn ; Mai, Le Bach ; Dung, Le Hong ; Lua, Tran Thi ; Nowak, Verena ; Elburg, Lucy ; Melse-Boonstra, Alida ; Brouwer, Inge - \ 2019
Maternal and Child Health 23 (2019)1. - ISSN 1092-7875 - p. 46 - 54.
Capacity building - Data quality - Food composition - Nutrient data - SMILING project

Objectives Food composition data are key for many nutrition related activities in research, planning and policy. Combatting micronutrient malnutrition among women and young children using sustainable food based approaches, as aimed at in the SMILING project, requires high quality food composition data. Methods In order to develop capacity and to align procedures for establishing, updating and assessing the quality of key nutrient data in the food composition tables in Southeast Asia, a detailed roadmap was developed to identify and propose steps for this. This included a training workshop to build capacity in the field of food composition data, and alignment of procedures for selecting foods and nutrients to be included for quality assessment, and update of country specific food composition tables. The SEA partners in the SMILING project finalised a country specific food composition table (FCT) with updated compositional data on selected foods and nutrients considered key for designing nutrient dense and optimal diets for the target groups. Results Between 140 and 175 foods were selected for inclusion in the country specific FCTs. Key-nutrients were: energy, protein, total fat, carbohydrates, iron, zinc, (pro-)-vitamin A, folate, calcium, vitamin D, vitamin B1, vitamin B2, vitamin B3, vitamin B6, vitamin B12 and vitamin C. A detailed quality assessment on 13 key-foods per nutrient was performed using international guidelines. Nutrient data for specific local food items were often unavailable and data on folate, vitamin B12 and vitamin B6 contents were mostly missing. For many foods, documentation was not available, thereby complicating an in-depth quality assessment. Despite these limitations, the SMILING project offered a unique opportunity to increase awareness of the importance of high quality well documented food composition data. Conclusion for Practise The self-reported data quality demonstrated that there is considerable room for improvement of the nutrient data quality in some countries. In addition, investment in sustainable capacity development and an urgent need to produce and document high quality data on the micronutrient composition of especially local foods is required.

Validation of accelerometer for measuring physical activity in free-living individuals
Agogo, George O. ; Voet, Hilko Van Der; Hulshof, Paul J.M. ; Trijsburg, Laura ; Eeuwijk, Fred A. Van; Boshuizen, Hendriek C. - \ 2018
Baltic Journal of Health and Physical Activity 10 (2018)1. - ISSN 2080-1297 - p. 7 - 21.
Background: The aim of this research was to validate a triaxial GT3X accelerometer against double labelled water for measuring total energy expenditure (TEE) in a study of free-living Dutch adults and to compare the two prediction equations used to calculate accelerometerderived activity related energy expenditure. Material/Methods: We used a measurement error model to estimate bias in the mean TEE, a correlation coefficient between measured and true TEE (a validity coefficient, which quantifies loss of statistical power to detect association) and the attenuation factor (which quantifies bias in the association), with and without conditioning on age, sex and BMI. We proposed a calibration method for the accelerometer-based TEE.
Results: The accelerometer underestimated TEE by about 500kcal/day. The validity coëfficiënt estimate conditional on age, sex and BMI was 0.8; the same value was observed for the attenuation factor estimate. With the devised calibration method, the bias in accelerometerderived mean TEE reduced to 6 kcal/day, validity coefficient estimate increased to 0.95 and attenuation factor to 0.94.
Conclusions: The GT3X accelerometer would underestimate mean TEE, lead to minimal loss in statistical power to detect significant association, and would result in biased estimate of the association between TEE and a health outcome.
Key words: attenuation, measurement error, physical activity, activity energy expenditure, validity coefficient.
Validating fatty acid intake as estimated by an FFQ : how does the 24 h recall perform as reference method compared with the duplicate portion?
Trijsburg, Laura ; Vries, Jeanne H.M. de; Hollman, Peter C.H. ; Hulshof, Paul J.M. ; ’t Veer, Pieter van; Boshuizen, Hendriek C. ; Geelen, Anouk - \ 2018
Public Health Nutrition 21 (2018)14. - ISSN 1368-9800 - p. 2568 - 2574.
Biomarker - Dietary assessment - Duplicate portion - Fatty acids - Measurement errors - Validity

Objective: To compare the performance of the commonly used 24 h recall (24hR) with the more distinct duplicate portion (DP) as reference method for validation of fatty acid intake estimated with an FFQ. Design: Intakes of SFA, MUFA, n-3 fatty acids and linoleic acid (LA) were estimated by chemical analysis of two DP and by on average five 24hR and two FFQ. Plasma n-3 fatty acids and LA were used to objectively compare ranking of individuals based on DP and 24hR. Multivariate measurement error models were used to estimate validity coefficients and attenuation factors for the FFQ with the DP and 24hR as reference methods. Setting: Wageningen, the Netherlands. Subjects: Ninety-two men and 106 women (aged 20–70 years). Results: Validity coefficients for the fatty acid estimates by the FFQ tended to be lower when using the DP as reference method compared with the 24hR. Attenuation factors for the FFQ tended to be slightly higher based on the DP than those based on the 24hR as reference method. Furthermore, when using plasma fatty acids as reference, the DP showed comparable to slightly better ranking of participants according to their intake of n-3 fatty acids (0·33) and n-3:LA (0·34) than the 24hR (0·22 and 0·24, respectively). Conclusions: The 24hR gives only slightly different results compared with the distinctive but less feasible DP, therefore use of the 24hR seems appropriate as the reference method for FFQ validation of fatty acid intake.

Comparing intake estimations based on food composition data with chemical analysis in Malian women
Koréissi-Dembélé, Yara ; Doets, Esmee L. ; Fanou-Fogny, Nadia ; Hulshof, Paul J.M. ; Moretti, Diego ; Brouwer, Inge D. - \ 2017
Public Health Nutrition 20 (2017)8. - ISSN 1368-9800 - p. 1351 - 1361.
Chemical analysis of duplicate portion - Food composition database - Food weighed record - Nutrient intake of Malian women - Validation
Objective: Food composition databases are essential for estimating nutrient intakes in food consumption surveys. The present study aimed to evaluate the Mali food composition database (TACAM) for assessing intakes of energy and selected nutrients at population level. Design: Weighed food records and duplicate portions of all foods consumed during one day were collected. Intakes of energy, protein, fat, available carbohydrates, dietary fibre, Ca, Fe, Zn and vitamin A were assessed by: (i) estimating the nutrient intake from weighed food records based on an adjusted TACAM (a-TACAM); and (ii) chemical analysis of the duplicate portions. Agreement between the two methods was determined using the Wilcoxon signed-rank test and Bland–Altman plots. Setting: Bamako, Mali. Subjects: Apparently healthy non-pregnant, non-lactating women (n 36) aged 15–36 years. Results: Correlation coefficients between estimated and analysed values ranged from 0·38 to 0·61. At population level, mean estimated and analysed nutrient intakes differed significantly for carbohydrates (203·0 v. 243·5 g/d), Fe (9·9 v. 22·8 mg/d) and vitamin A (356 v. 246 µg retinol activity equivalents). At individual level, all estimated and analysed nutrient intakes differed significantly; the differences tended to increase with higher intakes. Conclusions: The a-TACAM is sufficiently acceptable for measuring average intakes of macronutrients, Ca and Zn at population level in low-intake populations, but not for carbohydrate, vitamin A and Fe intakes, and nutrient densities.
O-methylisourea can react with the α-amino group of lysine : Implications for the analysis of reactive lysine
Hulshof, Tetske G. ; Rutherfurd, Shane M. ; Sforza, Stefano ; Bikker, Paul ; Poel, Thomas van der; Hendriks, Wouter H. - \ 2017
Journal of Agricultural and Food Chemistry 65 (2017)4. - ISSN 0021-8561 - p. 964 - 972.
Guanidination reaction - Reaction conditions - Reactive lysine - Specificity
The specificity of O-methylisourea (OMIU) to bind to the ε-amino group of Lys, an important supposition for the OMIU-reactive Lys analysis of foods, feeds, ingredients, and digesta, was investigated. Crystalline L-Lys incubated under standard conditions with OMIU resulted in low homoarginine recoveries. The reaction of OMIU with the α-amino group of Lys was confirmed by MS analysis, with double derivatized Lys being identified. None of the changes in reaction conditions (OMIU pH, OMIU to Lys ratio, and reaction time) with crystalline L-Lys resulted in 100% recovery of homoarginine. The average free Lys content in ileal digesta of growing pigs and broilers was found to be 13% of total Lys, which could result in a significant underestimation of the reactive Lys content. The reaction of OMIU with α-amino groups may necessitate analysis of free Lys to accurately quantify reactive lysine in samples containing a large proportion of Lys with a free α-amino group.
Processing beïnvloedt eiwitkwaliteit
Hulshof, Tetske - \ 2017

AnimalTetske Hulshof onderzoekt effecten hittebehandeling

Amino acid utilization and body composition of growing pigs fed processed soybean meal or rapeseed meal with or without amino acid supplementation
Hulshof, T.G. ; Poel, A.F.B. van der; Hendriks, W.H. ; Bikker, P. - \ 2017
Animal 11 (2017)7. - ISSN 1751-7311 - p. 1125 - 1135.
amino acid - body composition - nutrient retention - post-absorptive utilization - toasting with lignosulfonate

Feed ingredients used in swine diets are often processed to improve nutritional value. However, (over-)processing may result in chemical reactions with amino acids (AAs) that decrease their ileal digestibility. This study aimed to determine effects of (over-)processing of soybean meal (SBM) and rapeseed meal (RSM) on post-absorptive utilization of ileal digestible AAs for retention and on body AA composition of growing pigs. Soybean meal and RSM were processed by secondary toasting in the presence of lignosulfonate to obtain processed soybean meal (pSBM) and processed rapeseed meal (pRSM). Four diets contained SBM, pSBM, RSM or pRSM as sole protein source. Two additional diets contained pSBM or pRSM and were supplemented with crystalline AA to similar standardized ileal digestible (SID) AA level as the SBM or RSM diet. These diets were used to verify that processing affected AA retention by affecting ileal AA digestibility rather than post-absorptive AA utilization. The SID AA levels of the protein sources were determined in a previous study. In total, 59 pigs were used (initial BW of 15.6±0.7 kg) of which five were used to determine initial body composition at the start of the experiment. In total, 54 pigs were fed one of six experimental diets and were slaughtered at a BW of 40 kg. The organ fraction (i.e. empty organs plus blood) and carcass were analyzed separately for N and AA content. Post-absorptive AA utilization was calculated from AA retention and SID AA intake. An interaction between diet type, comprising effects of processing and supplementing crystalline AA, and protein source was observed for CP content in the organ fraction, carcass and empty body and for nutrient retention. Processing reduced CP content and nutrient retention more for SBM than for RSM. Moreover, processing reduced (P

BMI was found to be a consistent determinant related to misreporting of energy, protein and potassium intake using self-report and duplicate portion methods
Trijsburg, L.E. ; Geelen, M.M.E.E. ; Hollman, P.C.H. ; Hulshof, P.J.M. ; Feskens, E.J.M. ; Veer, P. van 't; Boshuizen, H.C. ; Vries, J.H.M. de - \ 2017
Public Health Nutrition 20 (2017)4. - ISSN 1368-9800 - p. 598 - 607.

As misreporting, mostly under-reporting, of dietary intake is a generally known problem in nutritional research, we aimed to analyse the association between selected determinants and the extent of misreporting by the duplicate portion method (DP), 24 h recall (24hR) and FFQ by linear regression analysis using the biomarker values as unbiased estimates.

For each individual, two DP, two 24hR, two FFQ and two 24 h urinary biomarkers were collected within 1·5 years. Also, for sixty-nine individuals one or two doubly labelled water measurements were obtained. The associations of basic determinants (BMI, gender, age and level of education) with misreporting of energy, protein and K intake of the DP, 24hR and FFQ were evaluated using linear regression analysis. Additionally, associations between other determinants, such as physical activity and smoking habits, and misreporting were investigated.

The Netherlands.

One hundred and ninety-seven individuals aged 20–70 years.

Higher BMI was associated with under-reporting of dietary intake assessed by the different dietary assessment methods for energy, protein and K, except for K by DP. Men tended to under-report protein by the DP, FFQ and 24hR, and persons of older age under-reported K but only by the 24hR and FFQ. When adjusted for the basic determinants, the other determinants did not show a consistent association with misreporting of energy or nutrients and by the different dietary assessment methods.

As BMI was the only consistent determinant of misreporting, we conclude that BMI should always be taken into account when assessing and correcting dietary intake.
Guanidination of amino acids: what is going on?
Hulshof, T.G. ; Rutherfurd, S.M. ; Bikker, P. ; Poel, A.F.B. van der; Hendriks, W.H. - \ 2016
WIAS Science Day 2016
Hulshof, Tetske - \ 2016
Processing influence on protein digestion and post-absorptive amino acid utilisation in growing pigs
Hulshof, Tetske - \ 2016
Oilseed meal processing affects whole body amino acid retention and composition in growing pigs
Hulshof, Tetske - \ 2016
Oilseed meal processing affects protein digestion kinetics and metabolic organ load of growing pigs
Hulshof, Tetske - \ 2016
Impact of free lysine on reactive lysine measurement by guanidination
Hulshof, Tetske ; Poel, A.F.B. van der; Hendriks, W.H. ; Bikker, P. - \ 2016
Oilseed meal processing affects whole body amino acid retention and composition in growing pigs
Hulshof, Tetske ; Poel, A.F.B. van der; Bikker, P. - \ 2016
In: Energy and protein metabolism and nutrition. - Wageningen Academic Publishers (EAAP publication 137) - ISBN 9789086862863 - p. 291 - 292.
Oilseed meal processing affects protein digestion kinetics and metabolics organ load of growing pigs
Hulshof, Tetske ; Poel, A.F.B. van der; Bikker, P. - \ 2016
In: Energy and protein metabolism and nutrition. - Wageningen Academic Publishers (EAAP publication 137) - ISBN 9789086862863 - p. 293 - 294.
An experiment was conducted to determine the effects of processing of soybean meal (SBM) and rapeseed meal (RSM) on crude protein (CP) digestibility along the small intestine, N solubility in chyme, and the effect of post-absorptive amino acid (AA) availability on metabolic organ load. Fiftyfour growing pigs were fed 1 of 6 experimental diets containing SBM, processed SBM (pSBM), RSM, processed RSM (pRSM), or pSBM and pRSM supplemented with crystalline AA. Pigs were
slaughtered at 40 kg, organ weight was recorded and chyme was collected at 3 locations along the small intestine. The CP digestibility was reduced due to processing, which could not be explained by an increased amount of insoluble N as fraction of total N. The weight of the kidneys, pancreas, liver, and small intestine was affected by processing and supplementation with crystalline AA.
Protein quality of pig diets : processing effects on amino acid digestibility and post-absorptive utilization
Hulshof, Tetske - \ 2016
Wageningen University. Promotor(en): Wouter Hendriks, co-promotor(en): Paul Bikker; Thomas van der Poel. - Wageningen : Wageningen University - ISBN 9789462579026 - 175
pigs - protein quality - pig feeding - feeds - feed processing - amino acids - protein digestibility - digestive absorption - protein utilization - nutrition physiology - animal nutrition - varkens - eiwitkwaliteit - varkensvoeding - voer - voedermiddelbewerking - aminozuren - eiwitverteerbaarheid - verteringsabsorptie - eiwitgebruik - voedingsfysiologie - diervoeding

The increasing world population and per capita income imposes a risk for protein scarcity. It is, therefore, necessary to use current ingredients more efficiently which includes the accurate assessment of protein quality before inclusion in animal diets. Protein quality is defined in this thesis as the capacity of a dietary protein to meet a pig’s requirement for nitrogen (N) and amino acids (AA) to meet a particular production target. Protein quality is influenced by processing applied to feed ingredients which may lead to the formation of Maillard reaction products (MRP) or cross-link products. The Maillard and cross-link reactions mainly involve lysine (Ly)s and their products may decrease ileal crude protein (CP) digestibility. During the acid hydrolysis step used to analyze AA, part of the early MRP revert back to Lys. This reverted Lys is not bioavailable for animals. Therefore, methods that specifically analyze Lys with a free ε-amino group (that is, not bound to other nutrients) have been developed. The guanidination reaction with O-methylisourea (OMIU) is one such method. The initial aim of this thesis was to evaluate the ileal digestible reactive Lys assay as a more accurate measure for protein quality of processed protein sources than the ileal digestible total Lys assay. Soybean meal (SBM) and rapeseed meal (RSM) were used as sole protein sources throughout this thesis. Processing of SBM and RSM by toasting at 95°C for 30 min in the presence of a sugar-rich lignosulfonate was used as model for over-processed protein sources.

Digestibility, post-absorptive utilization, and pig growth performance

In Chapter 2, protein quality in processed protein sources was determined using the content of AA, OMIU-reactive Lys, MRP, and lysinoalanine (LAL; as cross-link product), the standardized ileal digestibility (SID) of AA and OMIU-reactive Lys and pig growth performance. The SBM and RSM diets contained furosine and carboxymethyllysine (CML) as MRP, and LAL indicating that the Maillard and cross-link reactions had taken place in SBM and RSM, presumably during the oil extraction/desolventizing process. The amounts of furosine, CML, and LAL were elevated in the pSBM and pRSM diets due to further processing. Processing resulted in a reduction in total and OMIU-reactive Lys contents, a decreased pig growth performance as determined by the gain to feed ratio (G:F), and the SID of CP, AA, and OMIU-reactive Lys. The SID AA contents of the protein sources from Chapter 2 were used to formulate the diets of the main in vivo experiment (Chapters 3 and 4). In this experiment, six experimental diets were used of which four contained either SBM, pSBM, RSM, or pRSM as sole protein source. The remaining two experimental diets contained pSBM or pRSM and were supplemented with crystalline AA to the same SID AA levels as the SBM or RSM diet. These supplemented diets were used to verify that processing affected AA digestibility rather than post-absorptive AA utilization. The effects of processing on CP digestibility and N solubilization along the small intestine, metabolic load as assessed by organ weight, and nutrient composition of the empty body of growing pigs are described in Chapter 3. The small intestine was divided in three segments of similar length and digesta was collected from the last 100 cm of each segment. The amount of insoluble N as a fraction of N in digesta at each small intestinal segment was not affected by processing. Thus, the reduced SID of CP and AA reported in Chapter 2 was not caused by a reduced N solubility but by a general increase of N in digesta. Processing reduced the SID of CP, CP content in the empty body, and G:F. Supplementing crystalline AA to diets containing pSBM or pRSM increased the CP content and G:F to the level of the SBM and RSM diets. Processing also reduced the weight of several organs and supplementing crystalline AA restored organ weight. The effects of processing on whole body AA composition, nutrient retention, and post-absorptive utilization of AA in growing pigs are described in Chapter 4. Post-absorptive AA utilization was calculated as percentage of SID AA intake used for AA retention. Processing affected the AA composition of protein in the organ fraction (that is, empty organs and blood), carcass, and empty body. The Lys concentration in body protein was mainly reduced by processing. Supplementing crystalline AA restored the AA composition of body protein for SBM and RSM. Processing reduced AA retention and again supplementing crystalline AA restored AA retention for both SBM and RSM. Since crystalline AA were supplemented on an SID AA basis, the results indicated that processing affected AA digestibility but not post-absorptive AA utilization. Thus, correcting AA retention for SID AA intake would result in a similar post-absorptive AA utilization which was found for most AA for the RSM diets. However, the post-absorptive AA utilization was lower for the pSBM diet than for the SBM diet which might be related to an imbalanced AA supply after absorption in the first diet.

The assessment of ileal digestibility and utilization is expensive and laborious. Therefore, two alternative in vitro methods for determining protein digestibility for processed protein sources were evaluated (Chapter 5). The protein digestibility determined using the pH-STAT method and a 2-step enzymatic method was compared with the in vivo SID of CP reported in Chapter 2. Initial pH and the degree of hydrolysis assessed in the pH-STAT method were positively correlated to SID of CP. Protein digestibility determined with the 2-step enzymatic method, simulating digestion in the stomach and small intestine, tended to correlate to SID of CP. Both the 2-step enzymatic method and pH-STAT method were suitable alternatives for the assessment of SID of CP. However, only four ingredients were tested. The suitability of the methods should be further studied using multiple (processed) feed ingredients before they can be used as alternatives for in vivo assays.

Reactive Lys analysis

O-methylisourea was reported to bind specifically to the ε-amino group of Lys. The results of Chapter 2, however, cast doubt on the specificity of OMIU to react only with the ε-amino group of Lys. A series of experiments was conducted to study this specificity (Chapter 6). Incubating crystalline L-Lys with OMIU under standard conditions (OMIU pH of 10.6, OMIU to AA ratio of 1000:1, and reaction time of 7 d) resulted in a low homoarginine (that is, Lys with OMIU bound to its ε-amino group) recovery. The reaction of OMIU with the α-amino group of Lys was confirmed by mass spectrometry analysis with double derivatized Lys being identified. Several reaction conditions (OMIU pH, OMIU to Lys ratio, and reaction time) were studied but none of these resulted in 100% recovery of homoarginine. Binding of OMIU to the α-amino group of Lys could result in an underestimation of the reactive Lys content when significant levels of Lys with a free α-amino group (that is, crystalline L-Lys (HCl), free and N-terminal Lys) are present in food/feed ingredients, diets, and ileal digesta. The free Lys content in food/feed ingredients was on average 1.3% of total Lys. The free Lys content can be substantial in certain diets and was reported to be 13% of total Lys in ileal digesta. The latter might result in an overestimation of the OMIU-reactive Lys digestibility. The reaction of OMIU with α-amino groups may necessitate analysis of free Lys to accurately quantify reactive lysine in samples containing a large proportion of Lys with a free α-amino group.

The results presented in this thesis indicate that the effects of processing on SID of CP and AA, body composition, nutrient retention, post-absorptive AA utilization, and growth performance could be substantial. These effects should, therefore, be taken into account when using processed feed ingredients in diets for growing pigs. The extent of protein damage in feed ingredients can be assessed by the analysis of OMIU-reactive and total Lys, MRP, and cross-link products. However, OMIU-reactive Lys only provides accurate results when samples contain small levels of Lys with a free α-amino group (that is, crystalline L-Lys (HCl), free and N-terminal Lys). When samples contain significant levels of Lys with a free α-amino group, it is recommended to use standard guanidination conditions (OMIU pH of 10.6, OMIU to AA ratio of 1000:1, and reaction time of 7 d) to convert protein-bound Lys to homoarginine and to separately analyze such samples for free Lys.

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