Journal of Cereal Science 97 (2021) 103155 Contents lists available at ScienceDirect Journal of Cereal Science journal homepage: http://www.elsevier.com/locate/jcs Maize nutrient composition and the influence of xylanase addition Diego Melo-Durán a, José Francisco Pérez a, Gemma González-Ortiz b, Sandra Villagómez-Estrada a, Michael R. Bedford b, Hadden Graham b, David Sola-Oriol a, * a Servei de Nutricio i Benestar Animal (SNiBA), Department de Ciencia Animal i Dels Aliments, Universitat Autonoma de Barcelona (UAB), 08193, Spain b AB Vista, Marlborough, Wiltshire, SN8 4AN, United Kingdom A R T I C L E I N F O A B S T R A C T Keywords: This study assessed differences in nutrient composition, physical characteristics, and xylo-oligosaccharide con­ Maize genotype tent with or without xylanase treatment by maize genotype and the grain position on the cob. Ten cobs each from Grain cob position sixteen maize varieties sowed in the same field were collected and classified considering the grain’s position on Xylo-oligosaccharides the cob (basal vs apical). The majority of physicochemical characteristics were influenced by an interaction Xylanase between genetic background and grain position (P < 0.05); however, moisture, crude protein, starch, ash and soluble arabinose:xylose ratio differed between maize varieties and grain on cob position, without interaction. Xylanase addition increased the concentration of soluble compounds and xylotriose content in the aqueous phase following incubation in vitro (P < 0.05) and in the case of xylotriose the amounts released varied with grain position and variety. In conclusion, maize genotype and grain position on the cob significantly influenced chemical composition and oligosaccharide content when treated with xylanase, which may contribute to nutrient variability between maize samples. livestock nutrition. Xylanase is commonly added, particularly when viscous cereals such as wheat is used in poultry diets, in order to reduce 1. Introduction intestinal viscosity of birds by degradation of soluble arabinoxylans (AX) (Choct et al., 2004). However, the benefits obtained by degradation of Maize is the most common energy source used in commercial endosperm cell walls, which includes the release of encapsulated nu­ monogastric diets, especially in the American, Southern Europe and trients (“cage effect”) and/or through a prebiotic effect as a result of most Asian countries where maize grain is the primary cereal for poultry generation of short-chain oligosaccharides from polysaccharide AX feeds (Dei, 2017). However, the nutritional value of maize for livestock (Bedford and Apajalahti, 2000; Khadem et al., 2016). However, these feeding can vary (Cowieson, 2005). Genotype, agronomic conditions effects also contribute to the responses noted but these effects can be and pre- and post-harvest processing can affect the chemical charac­ heterogeneous and result in variable animal responses, depending on the teristics of cereal grains (Gehring et al., 2013). Indeed genetics has been quantity and nature of the cereal fiber polysaccharides present. demonstrated as an important source of biochemical and nutrient vari­ It was hypothesized that maize genetic background and grain posi­ ability (Rodehutscord et al., 2016; Uribelarrea et al., 2004). The nutrient tion on the cob might influence chemical composition, physical char­ composition of maize can also be affected by the position of the grain on acteristics, and response to xylanase supplementation. The present study the cob and grain size (Nadeem et al., 2014). Consequently, the smaller, aimed to investigate the differences in nutrient composition, physical extreme apical grains can produce nutritional variability, which could characteristics and short-chain xylo-oligosaccharide (XOS) content of affect the final composition of animal diets. Little is known about how basal or apical grains obtained from different maize hybrids harvested grain position on the cob and their interaction with genetic factors from the same field and under similar environmental growing and contribute to variations in the chemical composition and physical fertilizing conditions. characteristics of maize for poultry feeds. Characterization of the vari­ ation in nutritional value of maize grains that result from these factors may help define appropriate breeding objectives and improve the ac­ curate use of additives that depend on nutrient composition, such as enzymes, in order to improve the feeding value of cereal grains for * Corresponding author. E-mail address: David.Sola@uab.cat (D. Sola-Oriol). https://doi.org/10.1016/j.jcs.2020.103155 Received 22 September 2020; Received in revised form 14 December 2020; Accepted 15 December 2020 Available online 18 December 2020 0733-5210/© 2020 The Author(s). Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). D. Melo-Durán et al. Journal of Cereal Science 97 (2021) 103155 during the day. No pesticide was applied during cultivation, the fertil­ Abbreviations izer scheme was 170 kg of N/ha before sowing, 50 kg of liquid N/ha at 8 leaves, and 50 kg of liquid N/ha plus 5 L of organic fertilizer, through the NIRS near infrared spectroscopy irrigation system, at 12 leaves. Maize hybrids were sown in April 2018 SolDM solubility of dry matter and harvested in October 2018. The size of the land area and the harvest WRC water retention capacity weights of each variety were used for the calculation of maize yield per NDF neutral detergent fiber hectare (Table 1). The plot for each variety consisted of eight rows with ADF acid detergent fiber 17 cm of separation between plants within the row, and 70 cm of sep­ PSI protein solubility index aration between rows of the same variety. The cobs used in the present NSP total non-starch polysaccharides study were collected at random from the experimental field; briefly, for AX total arabinoxylans each variety a total of 50 cobs were obtained from groups of five cobs in AME apparent metabolizable energy ten sampling points that followed a diagonal pattern along the four XOS xylo-oligosaccharides central rows from each variety plot. Ten cobs were then randomly HPLC high performance liquid chromatography selected from these 50 for each hybrid. For each cob, all grains were ELSD evaporative light scattering detector collected according to their position. Briefly, grain cob position was A:X arabinose:xylose defined as apical or basal, with the line of differentiation between cat­ P phosphorus egories established subjectively considering the shape and size of the AA amino acids grains for each individual cob. The differences in the shape and size of DAD diode array detector the grains are shown in Fig. 1. Total grains from each position were oven HILIC hydrophilic interaction liquid chromatography dried at 60 ◦ C for 6 h, weighed (portion weight, PW), and stored in a cold SLM supported liquid membrane room at 4 ◦ C. The proximate and physiochemical analyses were obtained PROC GLM general linear model procedure for each hybrid and cob position using near infrared spectroscopy (NIRS), with results expressed on a fresh weight basis. Eight maize samples (variety 1, 2, 7, 11, 12, 13, 14 and 16) were selected, based mainly on starch, protein and non-starch polysaccharide (NSP) content and were subsequently analyzed for solubility of dry Table 1 matter (solDM), water retention capacity (WRC) and xylo- Maize samples. oligosaccharides (XOS, including X2: xylobiose; X3: xylotriose and X4: Varieties Trial Code Maize Yield (kg/ha) xylotetraose) release when incubated with xylanase in vitro. KWS-7661 Variety 1 19,621 KWS-16772 Variety 2 15,890 2.2. Maize proximate and physio-chemical analysis KWS-kxb9391 Variety 3 <8000 Kontigos Variety 4 15,746 The fresh weight proximate and physiochemical characteristics of KWS-7651 Variety 5 18,415 maize samples (16 varieties & 2 grain positions) viz: apparent metabo­ Kelindos Variety 6 15,856 KWS-6553 Variety 7 16,260 lizable energy (AME), crude protein, starch, crude fat, crude fiber, KWS-4565yg Variety 8 15,481 neutral detergent fiber (NDF), acid detergent fiber (ADF), protein solu­ Kefieros Variety 9 18,276 bility index (PSI), vitreousness, phytic acid phosphorus, total non-starch KWS-9393 Variety 10 <8000 polysaccharides (NSP), total arabinoxylans (AX), soluble AX, total KWS-7569 Variety 11 16,568 KWS-7562 Variety 12 18,656 arabinose:xylose ratio (A:X) and soluble A:X were predicted by NIRS Kefrancos Variety 13 14,897 (Foss DS2500, Hilleroed, Denmark) using calibrations provided by the KWS-7554 Variety 14 17,221 Feed Quality Service of AB Vista (Marlborough, England, United KWS-kbx9392 Variety 15 <8000 Kingdom). KWS-2679yg Variety 16 18,332 2.3. Dry matter solubility and water retention capacity 2. Materials and methods Dry matter solubility and WRC (Anguita et al., 2006) for five cobs 2.1. Maize samples from the eight selected varieties (variety 1, 2, 7, 11, 12, 13, 14 and 16), with two portions (apical and basal) and two levels of xylanase inclusion Sixteen maize varieties were sown in the same field and environ­ (with and without) were determined following an in vitro procedure mental growing conditions (Gimenells, Catalunya, Spain), including which simulates gastric pH (n = 160). In short, the samples were milled fertilizer and harvesting. The rainfall of the area was 207 mm in the and 0.5 g of each sample was weighed into a 10 mL screw cap tube and cultivation period and the average minimum/maximum temperatures incubated with 5 mL of 0.1 M sodium phosphate buffer and 2 mL of 0.2 varied between 15.2 and 29.3 ◦ C (AEMET, 2020). The planting density M hydrochloric acid (pH = 2.5). Additionally, 0.5 mL of a solution of was 92,000 seeds/ha. Irrigation was with fixed cane sprinkling and, liquid xylanase (Econase XT, 16,000 BXU/mL; one BXU is defined as the from sowing until an average plant height of 50 cm, at a rate of 1 h of amount of enzyme that produces one nmol reducing sugars from watering per day, applied at night. From 50 cm until the plants had birchwood xylan in 1 min at 50 ◦ C and pH 5.3.) was added to half of the 12-14 leaves, they were watered according to the need for soil moisture, samples at a dose of 16 BXU/mL. Tubes were kept at 41 ◦ C for 2 h in a based on rainfall and visual assessment of the plants. From 12 to 14 horizontal shaking water bath. The amount of sample submitted to leaves until the flowering spike is lost, water was applied for 30 min analysis was recorded (W0) as well as the weight of the screw cap tube daily, while from flowering to harvest, plants were watered for 1 h daily plus the sample (W1). After incubation, the tubes were centrifuged for at a rate of 50 min at night and 10 min at the time of maximum heat 20 min at 2000×g. The supernatant was carefully removed and tubes were kept upside down for 10 min to ensure that the non-retained water 2 D. Melo-Durán et al. Journal of Cereal Science 97 (2021) 103155 Fig. 1. From the top left to the bottom right, in both images, samples of cobs (A) and the corresponding grain category (upper: apical and lower: basal) (B) of maize 1, 2, 7, 11, 12, 13, 14 and 16 used in the in vitro assay are presented. was drained. The supernatant was used for XOS determination. Tubes O-XBI), xylotriose (X3; Prod Code. O-XTR) and xylotetraose (X4; Prod with sample were then weighed (W2), dried in the oven at 103 ◦ C for 16 Code. O-XTE) were obtained from Megazyme (Wicklow, Ireland). h to ensure the complete drying of the insoluble residue, and then Standards were used in a 250 ppm solution concentration with milli Q weighed again (W3). The solubility of the DM was calculated as follows: water to optimize the instrument parameters, and 0, 1, 2, 3, 4, and 10 mg/L solutions were prepared with milli Q water to obtain the cali­ W1 − W3 SolDM = { } bration line. The samples and standards were filtered through 22 μm W0 syringe filters. Analysis of the 162 standards and samples was carried Water retention capacity determined after centrifugation is out on an Agilent 1100 HPLC equipped with a DAD Agilent detector and expressed as grams of water retained by the total amount of sample an Agilent 1260 ELSD infinity detector. The HPLC column used was a incubated: Luna® 3u HILIC 200 A (150 × 2 mm), Phenomenex. The ELSD tem­ W2 − W3 perature was set at 80 ◦ C and air was used as nebulizer gas with a flow WRCDM = { W0 } rate of 1 SLM. The injection volume was 1 μL. Column temperature was set at 35 ◦ C and flow rate was 1.0 mL/min. Despite high sensitivity, good precision, simple operation and rapid analysis by the HPLC-ELSD 2.4. Xylo-oligosaccharides determination method, and being validated with purified samples of XOS in other studies (Li et al., 2016; Pu et al., 2017), the values obtained from this Determination of XOS was performed by high-performance liquid method are considered semi-quantitative, since the non-purified sam­ chromatography with an evaporative light scattering detector analysis ples could contain compounds that could interfere with the results. (HPLC-ELSD), adapting the method described by Pu et al. (2017). Briefly, supernatant collected after centrifugation (20 min at 2000×g) in 2.5. Statistical analysis the SolDM and WRC analysis were used in these determinations. Xylose (X1; Merck Life Science S.L., Madrid, Spain), xylobiose (X2; Prod Code. The cob was considered the experimental unit for all variables. The 3 D. Melo-Durán et al. Journal of Cereal Science 97 (2021) 103155 Fig. 2. Effects of maize genotype on physicochemical components. Only parameters without significant interaction are shown. Moisture, protein, ash, and soluble A: X ratio (Soluble arabinose:xylose ratio). The physicochemical components are expressed in as is basis. Each maize variety is a mean of 20 samples obtained from 10 cobs with two portions (n = 320). physicochemical characteristics were analyzed by two-way ANOVA to to each variety. The contents of crude fiber, total NSP and AX were identify genetic and grain cob position effects and the interaction be­ higher in the apical cob, while vitreousness and crude fat were higher in tween them. The solDM, WRC and XOS were analyzed by three-way basal grains in most of the hybrids. In spite of the interaction, total NSP ANOVA to identify genetic, grain cob position and xylanase effects (range: 55.6–81.3 g/kg), soluble NSP (range: 1.0–8.5 g/kg), total AX and the interaction between them. The PROC GLM procedure was per­ (range: 38.8–50.0 g/kg), and soluble AX (range: 2.2–5.3 g/kg) showed formed for both statistical analyses using SAS software (SAS, 2014). the highest variability due to maize genotype. Significantly different means were separated using Tukey’s HSD test. Moisture, protein, starch, ash, and soluble A:X ratio differed (P < Significance was declared at a probability P ≤ 0.05 and tendencies were 0.005) by genetic and grain cob position as individual factors. Except for considered when P-values were between >0.05 and < 0.10. Spearman’s ash, these factors were higher in basal grains than in those from the non-parametric correlation analysis was used to explore the associations apical portion (moisture: 104 vs 108 g/kg; crude protein: 78 vs 81 g/kg; between NIRS nutrient predictions, solDM, WRC, and XOS starch: 672 vs 676 g/kg; ash: 10.2 vs 9.7, g/kg and soluble A:X: 0.86 vs semi-quantitative estimation, using corrplot package of R 3.6.1. 0.88 g/g). The average and standard deviation of these components considering the genetics were; moisture: 106 ± 11.6 g/kg; crude protein: 3. Results 80 ± 5.6 g/kg; starch: 674 ± 13.3 g/kg; ash: 10 ± 1.0 g/kg and soluble A: X 0.87 ± 0.05. 3.1. Maize physicochemical analyses 3.2. In vitro analyses The effects on physicochemical composition of the maize genotype and the grain cob position are showed in Figs. 2 and 3, as well as their Results from the SolDM, WRC and semi-quantitative determination interaction are shown in Tables 2 and 3. Predicted poultry AME, crude of XOS in maize samples after the in vitro incubation with xylanase are fat, crude fiber, NDF, ADF, total NSP, soluble NSP, total AX, soluble AX, shown in Table 4. An interaction between variety and grain cob position phytic phosphorus, PSI, vitreousness, total A:X ratio, and portion weight was observed for solDM, WRC, xylobiose, xylotriose and xylotetraose. were influenced by an interaction between genetics and grain cob po­ The solDM and WRC were highest in the apical grains in six and five of sition (P < 0.05). The poultry AME, phytic P, A:X ratio, PSI, and portion the eight varieties, respectively. The contents of xylobiose and xylote­ weight were higher in grains from the basal position compared to those traose were highest in the apical grains in four and five of the eight from the apical position; however, these differences changed according varieties, respectively, while xylotriose content was higher in apical 4 D. Melo-Durán et al. Journal of Cereal Science 97 (2021) 103155 Fig. 3. Effect of grain cob position on physicochemical components. Only parameters without significant interaction are shown. Moisture, protein, ash, and soluble A:X ratio (Soluble arabinose:xylose ratio). The physicochemical components are expressed on an as is basis. Each portion is a mean of 160 samples obtained from 10 cobs of the 16 maize varieties (n = 320). grains in seven of the eight hybrids (Fig. 4). Furthermore, two in­ soluble A:X ratio (r = 0.37), PSI (r = 0.32), and vitreousness (r = 0.49). teractions were detected for xylotriose, the first between the genetic Additionally, significant correlations (P < 0.05) between physico­ variety and xylanase, and the second between the grain portion and chemical maize values and in vitro analysis were observed. The solDM in xylanase. The xylotriose content was increased in all hybrids when samples was negative correlated with phytic P (r = − 0.60). Xylobiose xylanase was added (Figs. 5 and 6). Similarly, higher xylotriose content was negatively correlated with PSI and crude fat (r = − 0.42 & r = was observed when xylanase was added in both grain cob positions; − 0.60, respectively). Xylotriose was positively correlated with WRC (r however, the response was greater in the grains of the apical portion = 0.60). The xylotetraose was positively correlated with soluble NSP compared those from the basal portion. and soluble AX (r = 0.54 & 0.63, respectively), and negatively correlated with poultry AME (r = − 0.61), crude fat (r = − 0.56), soluble A:X ratio (r = − 0.40), and PSI (r = − 0.54). 3.3. Correlations 4. Discussion Fig. 7 shows the correlation matrix and significance for the re­ lationships between all parameters. Significant correlations (P < 0.05) In the current study, the proximate analysis by NIRS of 16 maize were observed among physicochemical maize values. For example, varieties showed an effect of genotype on all parameters; however, most predicted poultry AME was positively correlated with PSI (r = 0.83), fat of these parameters also show an interaction with the grain position on (r = 0.71), soluble A:X ratio (r = 0.68), and negative correlated with the cob. The physicochemical characteristics which were not subject to a crude fiber (r = − 0.50), total NSP (r = − 0.51), soluble NSP (r = − 0.70), genotype:position interaction were moisture, crude protein, starch, ash, and soluble AX (r = − 0.67). Starch was negatively correlated with crude and soluble A:X ratio, but, as described, these were influenced by ge­ fiber (r = − 0.72), total NSP (r = − 0.48), soluble NSP (r = − 0.47), total notype. Rodehutscord et al. (2016) showed that genotype is an impor­ AX (r = − 0.40), and ash (r = − 0.63). Crude protein was positively tant factor influencing nutrient composition in several cereals used for correlated with crude fiber (r = 0.47), total A:X ratio (r = 0.49), phytic P animal feeding. The results obtained for moisture (97 ± 5.9 g/kg), crude (r = 0.51), and vitreousness (r = 0.82), and negatively correlated with protein (93.5 ± 9.18 g/kg DM), starch (740 ± 28.2 g/kg DM) and ash starch (r = − 0.52). Soluble AX was negative correlated with PSI (r = (13.3 ± 1.51 g/kg DM) of the 27 maize genotypes analyzed by Rode­ − 0.74), crude fat (r = − 0.76), and AME (r = − 0.60). Vitreousness was hutscord et al. (2016) are similar to those reported in the present study. positive correlated with soluble A:X ratio (r = 0.61) and PSI (r = 0.62). Similarly, physicochemical composition mean values were also in Maize yield was positive correlated with crude protein (r = 0.33), 5 D. Melo-Durán et al. Journal of Cereal Science 97 (2021) 103155 Table 2 Effects of maize variety and grain cob position on non-fiber physicochemical components a,b,c. Varieties/ Poultry AME Moisture Crude Starch Crude Fat Phytic P Ash PSI Vitreousness PW Portions Protein kJ/kg g/kg % g/cob Apical/Basal Apical/ Apical/ Apical/ Apical/ Apical/ Apical/ Apical/ Apical/Basal Apical/ Basal Basal Basal Basal Basal Basal Basal Basal Variety 1 15,181b/ 109/112 81/86 689/688 47b/50a 2.2cdef/2.4a 10.3/10.0 43b/47a 61cd/62bc 56jkl/ 15,455a 193efgh Variety 2 14,866cd/ 120/125 69/74 699/698 42cd/43c 1.9klm/ 8.8/8.3 36de/40c 55q/57mno 69ij/209cde 15,057b 2.1ghijk Variety 3 14,788cd/ 124/132 73/73 696/698 40ed/42cd 1.9ijkl/ 7.9/7.3 35def/37 d 55pq/56opq 52jkl/186gh 14,916c 2.1defgh Variety 4 14,129ijkl/ 92/97 73/72 680/682 35hijk/ 2.0hijkl/ 11.2/10.5 29ijklm/ 57klmno/ 68ijkl/182gh 14,255hij 36ghijk 2.1fghijk 31hi 58jklmn Variety 5 14,096klm/ 107/112 82/81 665/672 35ijkl/ 1.9jkl/ 10.6/9.5 31hi/33fgh 59ghi/58hij 74i/209cde 14,263hi 35hijk 2.1efgh Variety 6 13,852◦ / 111/116 80/78 665/668 31◦ /33mno 1.8n/1.9ijkl 9.3/9.1 28jklm/31hi 58ijkl/58ijkl 59ijkl/181h 13,998lmn Variety 7 13,928no/ 100/101 78/83 673/676 31◦ /32mno 2.1efghij/ 9.4/8.5 27lm/29ijkl 58no/57lmno 60ijkl/ 14,185jkl 2.2cd 200defg Variety 8 13,944no/ 118/124 82/85 652/650 35hijk/ 1.9lm/2.0ijkl 10.9/11.0 32gh/37d 58ijkl/60fgh 60ijkl/237ab 14,113jkl 37fgh Variety 9 14,125ijlk/ 95/96 84/87 663/678 36ghijk/ 2.2defg/2.3bc 11.2/10.5 31hij/ 58ijk/59efg 62ijkl/219bc 14,557e 37fghi 35defg Variety 10 14,013lmn/ 106/110 77/79 673/678 32mno/ 1.9lm/2.0klm 9.6/8.9 29ijkl/30ijk 58jklmn/ 50l/183gh 14,115jkl 32no 58ijklm Variety 11 13,963mno/ 95/98 73/80 676/677 33lmn/ 2.0klm/ 10.2/10.4 26m/34efg 57no/59ghi 62ijkl/ 14,389fgh 36ghij 2.1efghi 198defgh Variety 12 14,169jkl/ 110/116 88/93 646/660 39ef/37ghij 1.9klm/ 11.9/9.9 37d/41c 61cd/63ab 68ijk/212cd 14,464efg 2.0ijkl Variety 13 14,207jkl/ 93/94 85/90 660/668 36ghijk/ 2.2defg/ 11.5/10.8 34efg/37d 59fgh/61de 74i/190fgh 14,520ef 37fgh 2.4ab Variety 14 13,970mno/ 88/91 77/81 664/673 34klm/ 2.2defg/ 11.6/10.8 30ijk/34efg 57klmno/59hij 56ijkl/ 14,359gh 37ghij 2.4ab 208cdef Variety 15 13,882no/ 103/107 74/78 672/677 32no/35jkl 1.8mn/ 10.3/9.7 27klm/31hi 57nop/58ijkl 51kl/ 14,212ijk 2.2defg 198defgh Variety 16 14,370gh/ 92/95 83/86 679/679 33lmn/38fg 2.0hijkl/ 9.8/10.5 35def/42bc 61def/63a 66ijkl/250a 14,754d 2.2defg SEMd 50.2 1.3 1.9 3.1 0.8 0.04 0.33 0.9 0.4 6.4 P-value Variety <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 Position <0.0001 <0.0001 <0.0001 0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 Variety*Position 0.001 0.204 0.605 0.184 0.004 0.031 0.091 <0.0001 0.002 <0.0001 e Values in the same column not sharing a common letter are significantly different (P < 0.05). a Apparent metabolizable energy (AME), phosphorus (P), protein solubility index (PSI) and portion weight (PW). b The physicochemical components are expressed on an as is basis. c Data are a mean of 10 samples obtained from 10 cobs with two portions each (n = 320). d Standard error of the mean. agreement with FEDNA (2019) values reported for Spanish maize; this regard, a negative effect on digestion of starch and NDF has been however, the standard deviation for many, including crude protein and associated with high vitreous maize endosperm in dairy cows (Lopes starch (5 and 13 g/kg, respectively), could result in large differences et al., 2009). Similarly, in broiler chickens fed maize-based diets, the between random samples and thus would affect the final nutritional presence of a significant content of hard (high vitreousness) endosperm value of diets based on such grains and consequently animal perfor­ reduced the nutritional value (Kaczmarek et al., 2013). The present mance. The NIRS technology has become an important analytical tool in study identified variation in vitreousness with genotype and grain cob the field of animal nutrition, due to its practicality, ease of use, low cost, position, which suggests the proportion of hard endosperm varies with and speed in obtaining results. However, the accuracy is reliant on a these factors. detailed database of samples and the calibrations developed from this The WRC was also influenced by genetic background, which is likely database. linked to differences in fiber components between varieties (Robertson Several studies have shown that the variability contributed by ge­ and Eastwood, 1981). The high variation contributed by genotype for notype for wheat, triticale, maize, and rye for nutrient digestibility was total and soluble NSP, and AX observed in the current study suggests low with respect to ruminants (Krieg et al., 2017) but higher in poultry that it could be important to consider these components in plant (Zuber and Rodehutscord, 2017). The lesser effect in ruminants could be breeding and subsequently to take these into account in feed formula­ explained by the fact that their digestive process is based largely on tion. The antinutritive effect of soluble NSP is linked with increased bacterial fermentation. digesta viscosity, with negative changes in digesta transit time and Vitreousness, the ratio of hard to soft endosperm, has been showed to modification of intestinal mucosa and microbiota of the gut. Although be an indicator of nutrient digestibility in maize used in animal diets. In the quantity of soluble AX is lower in maize compare than other cereals, 6 D. Melo-Durán et al. Journal of Cereal Science 97 (2021) 103155 Table 3 Effects of maize variety and grain cob position on fiber parametersa,b,c. Varieties/ Crude Fiber NDF ADF Total NSP Soluble NSP Total AX Soluble AX Total A:X ratio Soluble A:X Portions ratio g/kg Apical/ Apical/ Apical/Basal Apical/Basal Apical/Basal Apical/Basal Apical/Basal Apical/Basal Apical/Basal Basal Basal Variety 1 19jk/21ghij 99bc/103ab 41b/46a 59nop/57op 1.0m/1.0m 40hijklmno/43efg 2.2p/2.1p 0.72ijklmno/ 1.02/1.06 0.70opqr l l cdef cd fghijkl defg mno pqr m m jklmno op mn Variety 2 14 /15 93 /95 35 /38 61 /55 1.0 /1.0 40 /39 3.1 /2.7 ◦ 0.70qr/0.69r 0.86/0.91 Variety 3 13l/14l 89efghi/95cd 33klmnop/ 57opq/54qr 1.0m/1.0m 38op/39mno 3.4klm/2.9no 0.69pqr/0.68s 0.86/0.90 bcde 38 Variety 4 20ghijk/ 84hijkl/82ijkl 29pq/30opq 76bc/72cdef 8.3abc/ 45cde/43def 4.9cdefgh/ 0.71lmno/0.70nopq 0.85/0.85 20hijk 8.2abcde 4.7gh Variety 5 20ghijk/ 80kl/86ghijk 31nopq/ 67ghijk/ 6.2ghi/5.4hij 41fghijklmn/ 4.6h/4.6h 0.73bcdefg/ 0.83/0.85 21ghijk 34hijklm 64jklmn 42fghijkl 0.72fghijkl Variety 6 20hijk/21ghi ijkl 82 / 34hijklm/ 62klmno/ 6.9 cdefg / 39nop/ 5.0cdefg / 0.74bcde/ 0.81/0.83 86ghijk 36efghij 63jklmn 6.7efgh 41ghijklmno 5.0bcdefg 0.72efghijkl fgh efgh Variety 7 22 /22 91defgh/ 35ghijkl/ 71defg/67ghijk 8.6ab/ 43efg/43efgh 5.4 /5.2abcd a 0.74defghijk/ 0.85/0.86 88efghi 36efghij 6.5fghi 0.72fghijkl Variety 8 23def/22efg 93cdef/94cde 37defghij/38defg 69defghi/ 6.9efg/ 43efghij/ 3.9ij/3.6jk 0.73efghijkl/ 0.86/0.88 66hijklm 7.1cdefg 42fghijklm 0.72mnop Variety 9 23cdef/ 95cd/79kl 33jklmno/30q 77bc/68efghij 6.9defg/ 47c/44def 5.1bcdefg/ 0.73bcdefg/ 0.87/0.87 20hijk 6.0ghi 4.7gh 0.71cdefghi Variety 10 19ijk/19k kl 79 /78 l 29q/ 67ghijk / 7.3bcdefg/ 41ijklmno / 4.9efgh/4.7gh 0.74bcdef/ 0.82/0.82 32lmnopq 61lmno 5.1ij 40klmno 0.73cdefgh Variety 11 22efg/22fgh 91defg/81jkl 32lmnopq/ 73bcde/68fghij 9.1a/ 43ef/42fghijklmn 5.2efgh/4.6gh 0.72jklmno/ 0.83/0.86 31mnopq 7.8abcdef 0.72klmno Variety 12 25a/21fgh 94cdef/82ijkl 41bc/40bcd 66ghijkl/51r 5.4hij/2.3lm 42fghijk/37p 3.4abcde/3.2h 0.73defghijk/ 0.90/0.91 0.75ab ab abcd cd defgh cdef bc defgh ghi ghi bc cd defgh Variety 13 25 /25 95 / 37 /38 76 /70 6.4 /6.2 48 /46 4.9 / 0.74bcd/0.73cdefgh 0.88/0.91 88defghi 4.8fgh ab abc Variety 14 25 /25 108a/99bc 40 bcd /38 cdef a 85 /78 b ab 8.5 /6.5 fgh a 50 /49 ab ab 5.4 / 0.73 defghij / 0.87/0.90 5.1abcdef 0.72hijklmn Variety 15 21ghijk/ 87fghij/ 33mnlopq/ 74bcd/68fghij 8.3abcd/ 43efghi/44def 5.3abc/4.8gh 0.73cdefg/ 0.81/0.85 21fgh 87fghij 35fghijkl 6.3ghi 0.72ghijklm Variety 16 23bcde/ 88efghij/76l 41b/33ijklmn 64ijklmn/60no 3.4kl/4.5jk 42fghij/40lmno 4.8gh/4.0i 0.76a/0.74abc 0.87/0.90 21fgh d SEM 0.7 2.5 1.2 1.9 0.5 0.9 0.11 0.005 0.009 P-value Variety <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 Position 0.117 0.001 0.081 <0.0001 <0.0001 0.003 <0.0001 <0.0001 <0.0001 Variety*Position 0.002 <0.0001 <0.0001 0.028 0.001 0.002 0.027 0.021 0.091 e Values in the same column not sharing a common letter are significantly different (P < 0.05). a Neutral detergent fiber (NDF), acid detergent fiber (ADF), non-starch polysaccharides (NSP), arabinoxylans (AX), ratio arabinose: xylose (A:X). b The physicochemical components are expressed on an as is basis. c Data are a mean of 10 samples obtained from 10 cobs with two portions each (n = 320). d Standard error of the mean. such as wheat, barley, rye, and triticale, the present study showed that simulation conditions used, as it has been shown that xylanase pro­ genetics can play an important role in soluble AX variability in maize. duction of XOS can be greater under more acidic conditions (Morgan The increment of soluble dry matter content when xylanase was et al., 2017). However, despite the benefits described about the included in the incubation of maize samples in the in vitro simulation HPLC-ELSD method, and the fact that the ELSD is not limited by the suggests a release of nutrients and/or soluble NSP and oligosaccharide optical properties or functional groups of the analytes and has been production. Xylanase supplementation clearly can result in significant widely used in the detection of chromophore-free compounds, such as release of oligosaccharides when used with different cereals but maize carbohydrates (Ma et al., 2014), the results from this study should be seems to be much more intransigent to such effects compared with confirmed by other methods, since maize extracts are complex and could wheat of other small grain cereals (Dale, 2020; Morgan et al., 2017). contain compounds that could interfere with the analysis. However, there is no information concerning the variability in XOS The grain cob position influenced moisture, crude protein, starch, production from maize samples when treated with xylanase; indeed Dale ash and soluble A:X ratio. The apical and basal portions in this study (2020) reported no production of XOS from maize samples when incu­ represent 23 ± 2% and 77 ± 2% of the total grain cob weight, respec­ bated with and without xylanase. However, the results of the present tively. Apical grains contained less moisture, crude protein, starch and study have shown that XOS contents increased, albeit marginally, with soluble A:X ratio, and higher total NSP, soluble NSP and other fiber xylanase addition in all maize samples, with different effects on xylo­ compounds, suggesting the differences in nutrient availability of a maize biose, xylotriose and xylotetraose release depending upon the maize sample can be due to grain position. In this regard, our results also sample itself. The current results show a marginal increment in xylo­ showed a negative correlation between crude fiber, total NSP and total triose and/or reduction in xylotetraose contents with xylanase supple­ AX with AME, SolDM and PSI, suggesting that these fiber compounds mentation. This study demonstrates that there are clear differences in influence nutrient availability. Previous studies showed that the size of XOS contents between maize samples whether they are treated or not the cereal grain could affect the physicochemical characteristics with xylanase. The amount of XOS likely is dependent upon the (Nadeem et al., 2014), possibly related to the amount of NSP, which has 7 D. Melo-Durán et al. Journal of Cereal Science 97 (2021) 103155 Table 4 Effects of maize variety, grain cob position and xylanase supplementation on dry matter solubility (SolDM), water retention capacity (WRC) and xylo-oligosaccharides (XOS) quantificationa. Main factors g/100 g WRC SolDM Xylobiose Xylotriose Xylotetraose Maize varieties Variety 1 131bc 11.0 0.26 0.12 0.66 Variety 2 127cd 11.8 0.27 0.09 0.75 Variety 7 128cd 10.7 0.32 0.07 0.94 Variety 11 139a 10.9 0.29 0.12 0.86 Variety 12 127cd 12.7 0.25 0.08 0.81 Variety 13 129bc 10.7 0.24 0.07 0.83 Variety 14 134ab 10.8 0.27 0.07 0.79 Variety 16 123d 10.5 0.23 0.06 0.77 SEMb 2.0 0.08 0.007 0.005 0.015 Position Apical 130 11.3 0.28 0.09 0.81 Basal 129 11.0 0.26 0.08 0.79 Xylanase With 131 11.6a 0.26 0.10 0.78b Without 128 10.7b 0.27 0.07 0.82a SEM 1.0 0.04 0.004 0.003 0.008 Probabilities Variety <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 Position 0.793 <0.0001 <0.0001 0.001 0.093 Xylanase 0.076 <0.0001 0.285 <0.0001 0.006 Variety*Position 0.323 <0.0001 <0.0001 0.028 0.033 Variety*Xylanase 0.201 0.403 0.802 0.004 0.990 Position*Xylanase 0.082 0.178 0.648 0.014 0.739 Variety*Position*Xylanase 0.508 0.716 0.998 0.391 0.999 abc Values in the same column not sharing a common letter are significantly different (P < 0.05). a Data for variety effect are a mean of 20 samples obtained from 5 cobs with two portions and two levels of xylanase inclusion (n = 160). b Standard error of the mean. Fig. 4. Interaction effect of maize genotype and grain cob position on xylobiose, xylotriose and xylotetraose production. Data are a mean of 10 replicates obtained from 5 cobs with two portions and two levels of xylanase inclusion each (n = 160). been shown to affect the nutritional value of the maize (Kaczmarek differences for many of the physicochemical characteristics of maize, et al., 2013). Indeed, one of the strategies to minimize such variation in including predicted poultry AME, crude fat, crude fiber, NDF, ADF, NSP, nutrient availability is the use of xylanase to reduce variability in animal soluble NSP, AX, soluble AX, total A:X ratio, phytic acid phosphorus, PSI, performance resulting from dietary AX (González-Ortiz et al., 2016). In vitreousness, and PW. As suggested previously, the interaction between the current work, xylanase application interacted with grain position, AX and other major nutrients could have a negative effect on nutrient showing a greater production of xylotriose from the apical grains availability. Arabinoxylan physico-chemical characteristics, such as compared to those from the basal, which is likely related to the higher gelling capability, depends on many characteristics, including molecular NSP values in the apical grains and/or lower soluble A:X ratio. weight, side-chain ferulic acid concentration and A:X ratio (Izydorczyk The interaction between genotype and grain position showed and Biliaderis, 1995), and AX-protein associations can also play an 8 D. Melo-Durán et al. Journal of Cereal Science 97 (2021) 103155 Fig. 5. Interaction effect of xylanase inclusion with the maize variety (A) and grain cob position (B) on xylotriose production. Data are a mean of 10 replicates for Variety*Xylanase and 40 samples for Portion*Xylanase, obtained from 5 cobs with two portions and two levels of xylanase inclusion each (n = 160). included in maize-basal diets, suggesting that the availability of the nutrients is compromised by these fiber compounds (Cordero et al., 2019). However, modulation of the microbiota by the supply of XOS produced from AX breakdown by xylanase represents an alternative method to increase the relative abundance of bacteria producing buty­ rate in the animal gut, as has been demonstrated by direct feeding of XOS (Cordero et al., 2019; Onrust et al., 2015). In this context, xylanase supplementation of maize-based diets could represent a valuable strat­ egy to control the negative effects of AX. The improvement shown in the SolDM fractions when xylanase was added may indicate an increased nutrient release and/or oligosaccharide production. The present results suggest that genetic background and the grain position on the cob, to a greater and lesser extent, contributed to dif­ ferences in nutrient composition, while the xylanase supplementation could be used to improve the nutritional value of maize through the increment of soluble compounds (nutrient releasing) and microbiota modulation by XOS production. The eight maize samples selected for detailed analysis in the present study were assessed in a broiler in vivo trial that showed differences on performance and nutrient digestibility associated to the nutritional variability produced by the maize genotype, showing the impact of this variability under practical conditions. 5. Conclusion In conclusion, maize genotype and grain position on the cob have an important influence on the physicochemical composition and oligosac­ charide contents and may contribute to nutrient variability. The NSP Fig. 6. Chromatogram of xylose (X), xylobiose (X2), xylotriose (X3) and xylo­ compounds were close related maize predicted AME for poultry. Apical tetraose (X4) semi-quantification in a maize sample. (A) Maize sample incu­ grains have lower nutritional value and higher fiber compounds, which bated without (red) and with (blue) xylanase. (B) Maize sample without could compromise the final composition of animal diets. Xylanase sup­ xylanase (blue) and enriched with standards (red). The numbers in the graphs plementation increased dry matter solubility and xylotriose content in express time retention of the peak. (For interpretation of the references to color vitro, suggesting an improvement in the nutrient availability if oligo­ in this figure legend, the reader is referred to the Web version of this article.) saccharide production also occurred within the gastrointestinal tract of broilers. important role, for example on the digestibility of the protein fraction (Méndez-Encinas et al., 2019). These factors add to the problem of the CRediT authorship contribution statement deficient levels of lysine and tryptophan in maize (Larkins et al., 2017), so variance in their digestibility could exacerbate this and influence Diego Melo-Durán: Conceptualization, methodology, investigation, animal performance significantly. The amount and packing of protein data curation and software, formal analysis, writing-original draft within the grain is partly reflected in the vitreousness (Paulsen et al., preparation. José Francisco Pérez: Conceptualization, methodology, 2003), and a potential reduction of access to digestive enzymes, with investigation, writing-original draft preparation and supervision. consequent lower digestibility. Non-starch polysaccharide Gemma González-Ortiz: Conceptualization, methodology, writing- anti-nutritional effects are considered minimal in maize, because it is review and editing and supervision. Sandra Villagómez-Estrada: believed to have a relatively low soluble AX content with a minor impact investigation and formal analysis. Michael R. Bedford: Writing-review on animal digesta viscosity. However, in this study it was demonstrated and editing. Hadden Graham: Writing-review and editing. David Sola- that soluble AX ranged from 2.2 to 5.3 g/kg, depending on genetics. Oriol: Conceptualization, methodology, investigation, writing-original These values highlight that some maize could behave similar to wheats draft preparation and supervision. in promoting viscosity issues. An improvement has been observed in poultry performance and nutrient digestibility when carbohydrases are 9 D. Melo-Durán et al. Journal of Cereal Science 97 (2021) 103155 Fig. 7. Correlations between nutrient content of maize varieties and in vitro analysis. In vitro values only include the eight varieties analyzed. The * symbol indicates a statistically significant correlation (P < 0.05). The scale colors (Spcobman’s ρ from − 1 to +1) indicate whether the correlation is positive (blue colored circles) or negative (red colored circles). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.) Acknowledgments Gehring, C.K., Cowieson, A.J., Bedford, M.R., Dozier, W.A., 2013. Identifying variation in the nutritional value of corn based on chemical kernel characteristics. World’s Poult. Sci. J. 69, 299–312. https://doi.org/10.1017/S0043933913000317. The authors acknowledge the support of the SENESCYT (Secretary González-Ortiz, G., Olukosi, O., Bedford, M.R., 2016. Evaluation of the effect of different for Higher Education, Science, Technology and Innovation of Ecuador) wheats and xylanase supplementation on performance, nutrient and energy grant ref. CZ02-000816-2018, AB Vista Laboratory Services (UK) for utilisation in broiler chicks. Anim. 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