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Journal of Animal Science logoLink to Journal of Animal Science
. 2019 Jul 20;97(9):3984–3993. doi: 10.1093/jas/skz243

Effects of abomasally infused amylase and increasing amounts of corn starch on fecal excretion of starch, total and microbial nitrogen, and volatile fatty acids in heifers1

Kristina Robbers 1, Edwin Westreicher-Kristen 1,✉, Arnulf Troescher 2, Andreas Susenbeth 1,✉
PMCID: PMC6736103  PMID: 31325356

Abstract

The aim of the present study was to study the effect of exogenous amylase on postruminal disappearance of increasing amounts of corn starch being infused into the abomasum of heifers, and to detect a possible limitation of starch digestion in the small intestine. Four rumen-fistulated heifers (2 German Black Pied and 2 Jersey × German Black Pied) with an initial BW of 565 ± 6 kg were fed 5.6 kg DM/d of a diet targeted to contain only a negligible amount of starch. Animals were assigned randomly to a crossover trial with 2 experimental periods lasting 35 d each with 10 d of diet adaption followed by 25 d of sample collection. During the sampling period, each animal was abomasally infused with native corn starch at 5 levels (953, 1,213, 1,425, 1,733, and 1,993 g DM/d) each for a 5-d period with and without exogenous amylase, respectively. At days 6 to 10 the heifers received an abomasal infusion of starch in amounts of 724 g/d. Feces were sampled 4 times a day during the collection periods. Titanium dioxide was ruminally administered (10 g/d) to estimate fecal excretion. Purine bases in feces were determined and used as a marker for microbial N excretion. Fecal excretion of microbial N increased linearly with increasing level of starch infusion (P < 0.001), indicating a constant proportion of the infused starch being fermented in the hindgut. In contrast, the apparent digestibility of starch from the total postruminal tract decreased linearly from 90% to 80% (P < 0.001) when the intestinal starch supply increased from 1 to 2 kg/d. There is strong evidence based on the increasing starch excretion with feces and the indication of a constant proportion of infused starch being fermented in the hindgut for a decreasing efficiency of starch digestion in the small intestine with increasing intestinal supply. Amylase administration increased fecal excretion of butyrate (P = 0.04) and tended to increase isovalerate excretion (P = 0.06). However, amylase did not affect fecal excretion of microbial N or starch, suggesting that pancreatic amylase activity may not be the primarily limiting factor of postruminal starch digestion in heifers when corn starch is abomasally infused in amounts up to 2 kg/d.

Keywords: amylase, cattle, fecal composition, hindgut fermentation, small intestinal digestion, starch

INTRODUCTION

Previous studies with diets containing high proportion of grain (Karr et al., 1966) or with corn starch infused into the abomasum (Little et al., 1968; Kreikemeier et al., 1991; Richards et al., 2002) or duodenum (Matthé, 2001; Brake et al., 2014) have shown a limited digestion of starch in the small intestine of cattle. Possible limiting factors are well reviewed by Owens et al. (1986) and Huntington (1997) and include the capacity for glucose absorption, the time for starch hydrolysis, the activity of hydrolyzing enzymes, adequate working conditions, and the absence of enzyme inhibitors as well as the access of enzymes to starch granules. Thus, the supply of exogenous amylase could be seen as a strategy to improve starch digestibility in the small intestine. It was shown that up to 880 g/d corn starch can be digested in the small intestine of dry cows (Matthé, 2001) and 830 g/d in beef steers (Branco et al., 1999). However, intestinal digestibility of 880 g/d of corn starch infused into the abomasum of heifers was not improved when exogenous amylase was administered (Westreicher-Kristen et al., 2018). Missing effect of amylase administration was suggested to be due to a sufficient activity of endogenous amylase at this low level of starch infusion. Thus, further studies are required to clarify whether the activity of endogenous amylase might be a limiting factor at higher levels of abomasal starch infusion. The herein study aimed to investigate the postruminal digestion of different levels of starch infused into the abomasum of heifers without and with exogenous administration of amylase. We hypothesized that intestinal digestibility of starch might be limited by endogenous amylase activity with amounts of infused starch higher than 1 kg/d and that the disappearance of starch in the small intestine of adapted animals can be improved by exogenous amylase.

MATERIALS AND METHODS

The animal study reported herein was performed in accordance with the German Animal Welfare Act (Federal Republic of Germany, 2014) and approved by the Animal Welfare Commission of the Ministry of Energy, Agriculture, Environment, and Rural Areas of the Federal State of Schleswig – Holstein, Germany (V 244-7224.121-25).

Animals and Diets

Four fully grown heifers (2 German Black Pied and 2 Jersey × German Black Pied) with an initial BW of 565 ± 6 kg and fitted with rumen cannulas (#2C, 10 cm i.d.; Bar Diamond, Inc., Parma, ID) were used. The heifers were fed 5.6 kg/d of a diet targeted to contain only a negligible amount of starch and consisting of 65.4% grass hay (primary growth of Lolium perenne-dominated sward), 32.5% dried beet pulp (J. August Plambeck GmbH, Brügge, Germany), 0.9% feed urea (Piarumin; Stickstoffwerke Piesteritz GmbH, Wittenberg, Germany), and 1.2% of a mineral and vitamin premix (Panto-Mineral; Hamburger Leistungsfutter GmbH, Hamburg, Germany) on DM basis in 2 equal meals at 0700 and 1500 h. The ingredients and chemical composition of the experimental diet are shown in Table 1. The diet was formulated to meet maintenance requirements according to the German feeding standards (GfE, 2001).

Table 1.

Ingredients and chemical composition of the diet

Item % of dietary DM
Ingredient
 Grass hay 65.4
 Dried beet pulp 32.5
 Feed urea 0.90
 Mineral and vitamin mix1 1.20
Nutrient2
 Crude ash 6.5
 Crude protein 8.5
 Ether extract 0.6
 Starch 0.2
 NDForg 58.5
 ADForg 33.0
 ADL 3.4
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DM = dry matter; NDForg = neutral detergent fiber (expressed exclusive residual ash); ADForg = acid detergent fiber (expressed exclusive residual ash); ADL = acid detergent lignin.

1The mineral and vitamin mixture contained 210 g calcium, 100 g sodium, 40 g magnesium, 30 g phosphorus, 1,000 mg copper, 8,000 mg zinc, 5,000 mg manganese, 60 mg iodine, 50 mg selenium, 40 mg cobalt, 5,000 mg vitamin E, 1,000,000 IU vitamin A and 100,000 IU vitamin D3 (as fed).

2Calculated from the analyzed nutrient composition of individual ingredients.

Experimental Design and Procedure

The experiment consisted of 2 periods lasting 35 d each with 10 d for adaptation to the diet followed by 25 d for treatments application and sample collection. Between periods, a 5-wk period without treatment application was set to avoid carryover effects of adaptation to starch digestion. During the 25-d sampling periods, each animal received an abomasal infusion of corn starch at 5 infusion levels increasing each 5 d from a targeted dosage of 1,000 to 1,250, 1,500, 1,750, and 2,000 g/d. The exact amounts of starch infused were determined by recording the amount of residual infusate solution. The effective dosages were 953, 1,213, 1,425, 1,733, and 1,993 g/d. Prior to these experimental periods, animals were adapted to starch infusion and digestion at an effective dosage of 724 g/d at days 6 to 10, to have similar increases of starch supply (≈250 g/d) for all experimental starch levels including the lowest. The native corn starch contained 95.7% starch, 0.4% CP, and 0.2% crude ash on DM basis (Maisita 21.000; AGRANA AG, Vienna, Austria). The starch product was mixed in 10 liters water and kept suspended by continuous stirring. The abomasal infusion was performed using a tool to insert lines into the abomasum described by Westreicher-Kristen and Susenbeth (2017). The infusion began half an hour after morning feeding and lasted 10 h/d. A peristaltic pump (rotarus Flow 50; Hirschmann Laborgeräte GmbH & Co. KG, Eberstadt, Germany) was used for infusion.

The animals were assigned randomly to a crossover design. In each experimental period, 2 animals received an additional infusion of an exogenous α-amylase (EC.3.2.1.1; 13,000 U/mL, Distizym BA-N, 1,4-α-D-Glucan-Glucanohydrolase; Erbslöh Geisenheim AG, Geisenheim, Germany). The α-amylase was mixed with distilled water and continuously infused into the line for starch infusion throughout each 10-h infusion phase at a rate of 50 mL/h using a syringe pump (540060-HP Single Syringe; TSE Systems GmbH, Bad Homburg, Germany). The amylase activity of Distizym BA-N was determined in a bicinchoninic assay as described by Westreicher-Kristen et al. (2018). A dosage of Distizym BA-N of 260,000 U/kg starch was administered. Total fecal excretion was determined using titanium dioxide (TiO2; Kronos 1171; Harold Scholz & Co. GmbH, Recklinghausen, Germany). For this purpose, 5-g TiO2 were ruminally administered via gelatin capsules (HKG size 30 × 90 mm, 59 mL, limpid-transparent, type 36; Capsula GmbH, Ratingen, Germany) to each heifer twice daily immediately after each feeding. The TiO2 administration was initiated 5 d before the phase of sample collection to reach the equilibrium of TiO2 excretion after initial administration (Glindemann et al., 2009).

Sampling, Sample Preparation, and Analytical Procedures

During each experimental period, 6 samples (~200 g fresh matter each) of the grass hay and dried beet pulp were collected and pooled by feed for chemical analyses. The feed samples (1-mm particle size) were analyzed in duplicate for concentrations of moisture, crude ash (CA), crude protein (CP), and ether extract according to the official analytical methods in Germany (VDLUFA, 2007). Feed samples (2-mm particle size) were analyzed in triplicate for NDF, ADF, both expressed on ash-free basis, and ADL as described by Van Soest et al. (1991) using an ANKOM 200 Fiber Analyzer (ANKOM Technology, Macedon, NY). Heat-stable α-amylase was added during NDF extraction. Native corn starch used for abomasal infusion was analyzed for starch concentration based on polarimetric procedure (method 7.2.1; VDLUFA, 2007). Fecal grab samples (~500 g fresh matter) were collected from the rectum during the 25-d sampling phase 4 times a day at 0900, 1200, 1500, and 1800 h. Immediately after collection, samples were kept at −20 °C for ~15 min to quickly minimize microbial activity. Afterwards, samples were placed in a cooling room at 4 °C and merged per animal at the end of each sampling day. Six subsamples per day (minimum 100 g per sample) were taken and stored at −20 °C for analyses. Subsamples were thawed and pooled by animal and level of starch infusion at the end of each experimental period. Samples of the first day of each infusion level were excluded because they were assumed to be possibly affected by the previous level of starch infusion due to delayed excretion. The pooled fecal samples were analyzed in duplicate for moisture, CA, N, and NDF according to VDLUFA (2007). Fecal concentration of TiO2 was determined according to Brandt and Allam (1987) as described by Westreicher-Kristen et al. (2018). Concentration of starch was determined as total α-linked glucose polymers in lyophilized samples (0.2-mm particle size) by 2-step enzymatic hydrolysis according to Brandt et al. (1987), briefly described by Westreicher-Kristen et al. (2018). Concentration of VFA in feces (acetate, butyrate, propionate, valerate, isovalerate, and isobutyrate) was determined using a gas chromatograph, and concentration of l-lactate using a Lactate-UV Test Kit as described by Westreicher-Kristen et al. (2018). The concentration of purine bases (PB) in feces was determined by HPLC according to Dickhoefer et al. (2016).

Calculations and Statistical Analysis

Fecal excretion of DM was calculated as TiO2 administration (g/d) divided by the TiO2 concentration (g/kg DM) in feces, assuming a fecal recovery of 100%. Fecal excretion of OM, total N, microbial N, starch, VFA, and lactate was calculated based on their concentrations in feces (g/kg DM) and fecal DM excretion. The OM concentration in feces was calculated as 1000 − CA (g/kg DM). The concentration of total lactate was calculated assuming a ratio of l- and d-lactate of 50:50. The apparent total tract disappearances of OM (ATTDOM) and NDF (ATTDNDF) were calculated as the quotient of the total amount that apparently disappeared (intake − fecal excretion) and the amount of intake including the amount of OM infused as starch. The apparent disappearance of starch from total postruminal tract (ATTDStarch) was calculated as the quotient of the total amount that apparently disappeared (infusion − fecal excretion) and the amount of starch infused. The concentration of microbial N in feces was calculated based on PB excretion using a PB-N:microbial N ratio of 0.116 according to Chen (1989).

All data were statistically analyzed by the MIXED procedure of SAS (SAS Institute, Inc., Cary, NC). Period, infusion level, treatment, and infusion level × treatment interactions were included as fixed effects and animal as a random factor. The LSM and SEM of each parameter were calculated and compared using the LSMEANS statement. Linear and quadratic orthogonal contrasts were tested for the effects of starch infusion level using the CONTRAST statement. Unequal spacing between starch infusion levels was considered by generating the coefficients for linear and quadratic contrasts using the IML procedure of SAS. Treatment effects were considered significantly different at P < 0.05 and tendencies with P values between 0.05 and 0.10. Simple linear regression analyses were performed (PROCREQ) to determine the fecal excretion of microbial N and starch as well as starch disappearance from the small intestine as a function of the amount of starch infused.

RESULTS

Fecal Excretions and ATTD

Fecal excretion of DM, OM, N, microbial N, starch, lactate, and VFA as well as ATTDOM, ATTDNDF, and ATTDStarch at increasing levels of starch infusion with or without exogenous amylase are presented in Table 2. Fecal excretion of DM and OM increased linearly with increasing level of starch infusion (P < 0.001 for both). The ATTDOM decreased linearly with infusion level (P = 0.02) with values ranging between 72.9% and 70.1%. The administration of amylase did neither affect fecal excretion of DM (P = 0.98) and OM (P = 0.75) nor ATTDOM (P = 0.69). The ATTDNDF was neither affected by level of starch infusion (linear: P = 0.80; quadratic: P = 0.39) nor by amylase administration (P = 0.95) and averaged 69.2% for both treatments.

Table 2.

Fecal excretion of DM, OM, N, VFA, starch, and lactate in heifers infused abomasally with increasing amounts of starch with or without exogenous amylase administration

Treatment1
Starch Starch + amylase
Amount of starch infused, g/d P-value2
Item 953 1213 1425 1733 1993 953 1213 1425 1733 1993 SEM L Q α
DM, kg/d 1.97 1.96 2.06 2.17 2.34 1.95 1.97 2.05 2.22 2.32 0.77 <0.001 0.04 0.98
OM, kg/d3 1.70 1.69 1.80 1.92 2.09 1.70 1.72 1.79 1.98 2.07 0.76 <0.001 0.04 0.75
Starch, kg/d 0.09 0.13 0.20 0.28 0.39 0.10 0.16 0.18 0.34 0.39 0.07 <0.001 0.24 0.54
Total N, g/d 46.2 45.5 46.9 47.1 47.3 46.4 44.8 46.0 46.0 47.9 1.10 0.02 0.10 0.41
 Microbial N, g/d 9.38 9.61 9.59 9.97 10.6 9.33 9.74 9.91 10.4 10.7 0.53 <0.001 0.58 0.46
 Microbial N, % of N 20.3 21.1 20.5 21.1 22.4 20.2 21.7 21.7 22.7 22.4 1.00 0.01 0.92 0.21
Lactate, g/d4 0.59 1.26 1.67 2.68 3.96 0.64 1.44 1.57 2.20 3.11 0.78 <0.001 0.40 0.50
VFA, g/d 34.0 43.2 49.0 50.9 53.0 39.1 45.0 49.3 56.0 57.2 4.52 <0.001 0.22 0.18
 Acetate, g/d 22.5 26.8 29.9 32.2 32.2 25.3 26.9 30.0 33.7 34.3 2.68 <0.001 0.37 0.34
 Propionate, g/d 2.36 2.98 2.81 2.15 2.15 2.89 2.49 2.70 2.15 2.05 0.62 0.12 0.43 0.90
 Butyrate, g/d 8.11 12.3 15.3 15.7 17.9 10.0 14.5 15.5 19.2 19.9 1.88 <0.001 0.13 0.04
 Valerate, g/d 0.31 0.31 0.18 0.16 0.17 0.28 0.18 0.26 0.14 0.16 0.07 0.02 0.57 0.53
 Isobutyrate, g/d 0.29 0.31 0.35 0.29 0.24 0.29 0.36 0.35 0.32 0.27 0.06 0.22 0.02 0.34
 Isovalerate, g/d 0.39 0.45 0.45 0.33 0.32 0.39 0.49 0.47 0.49 0.41 0.08 0.33 0.05 0.06
Acetate, %5 68.5 63.8 62.1 64.2 61.8 66.3 61.3 62.3 61.2 61.2 1.22 <0.001 0.02 0.05
Propionate, %5 7.48 6.99 6.01 4.27 4.28 8.10 5.68 5.65 4.03 3.66 1.50 <0.001 0.62 0.57
Butyrate, %5 24.0 29.2 31.9 31.5 33.9 25.6 33.0 32.1 34.8 35.1 1.60 <0.001 0.01 0.02
ATTDOM, %6 71.7 72.9 72.1 71.5 70.1 71.7 72.4 72.2 70.6 70.3 1.12 0.02 0.05 0.69
ATTDNDF, % 67.2 70.1 70.8 70.1 68.2 69.1 69.2 70.4 66.9 70.4 2.79 0.80 0.39 0.95
ATTDStarch,% 90.5 89.3 85.9 84.1 80.3 89.3 87.1 87.0 80.4 80.3 4.32 <0.001 0.61 0.39
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SEM = standard error of mean; DM = dry matter; OM = organic matter; N = nitrogen; VFA = volatile fatty acids; ATTD = apparent total tract disappearance.

1Values represent least square means (LSM) and SEM of the last 4 d of each infusion level within each treatment (n = 4).

2Linear (L) and quadratic (Q) effects of starch infusion and effect of amylase administration (α).

3OM was calculated as 1000 − CA (g/kg DM). VFA are partly included as they were captured in DM analysis.

4Lactate excretion including d- and l-isomers.

5Proportion of the sum of acetate, propionate and butyrate.

6OM infused as starch was considered as intake.

The fecal excretion of total N and microbial N increased linearly with increasing level of starch infusion (P = 0.02 and P < 0.01, respectively) from 46.3 to 47.6 g/d and 9.3 to 10.7 g/d, respectively. On average, microbial N excretion increased by 0.12 g/100 g of additionally infused starch (Figure 1). Neither total N (P = 0.41) nor microbial N excretion (P = 0.46) were affected by amylase. The proportion of microbial N in feces (as % of total N) was unaffected by amylase (P = 0.21), but increased linearly with starch infusion (P = 0.01) from 20.2% to 22.7%.

Figure 1.

Figure 1.

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Fecal excretion of microbial N as a function of the amount of starch infused.

Fecal excretion of VFA increased linearly with increasing level of starch infusion (P < 0.001), but was not affected by amylase (P = 0.18). Acetate and butyrate excretion increased with level of starch infusion (P < 0.001 for both), whereas propionate excretion was not affected by infusion level (linear: P = 0.12; quadratic: P = 0.43). Acetate excretion and propionate excretion were not affected by amylase (P = 0.34 and P = 0.90, respectively), but butyrate excretion was higher with amylase than without (P = 0.04). Molar proportion of acetate and propionate decreased (P < 0.001) and of butyrate increased (P < 0.001) linearly with increasing level of starch infusion. Additionally, molar proportion of butyrate increased (P = 0.02) and of acetate decreased due to amylase (P = 0.05), whereas molar proportion of propionate was unaffected by amylase (P = 0.57). Valerate excretion decreased linearly with starch infusion (P = 0.02), but was unaffected by amylase administration (P = 0.53). Excretions of isobutyrate and isovalerate responded quadratically (P = 0.02 and P = 0.05, respectively) with increments until starch infusion levels of 1,425 and 1,213 g/d, respectively, and decrements with further increasing levels of infusion. The administration of exogenous amylase tended to increase the excretion of isovalerate (P = 0.06), whereas isobutyrate was not affected by amylase (P = 0.34). Lactate excretion increased linearly with increasing level of starch infusion (P < 0.001), but was not affected by amylase (P = 0.50) with values ranging between 0.6 and 4.0 g/d. Starch excretion increased linearly (P < 0.001) on average with 29.2 g/100 g of additionally infused starch (Figure 2). However, the ATTDStarch decreased linearly (P < 0.001) from 89.9% to 80.3% when averaged for both treatments. Amylase affected neither starch excretion (P = 0.54) nor ATTDStarch (P = 0.39).

Figure 2.

Figure 2.

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Fecal excretion of starch as a function of the amount of starch infused.

DISCUSSION

Methodological Aspects

The lowest level of starch infusion was targeted to be 1 kg/d, because this was known to be an acceptable amount from this preceding experiment, where diarrhea was not observed. The rate of starch infusion was increased moderately to determine the capacity of starch digestion in animals adapted to starch digestion. Feces started to have a softer consistence—a few hours after beginning of abomasal starch infusion and became softer and stickier with increasing level of starch infusion. However, cases of diarrhea were not observed throughout the whole infusion periods. Additionally, feed intake was not influenced by the presence of the infusion device in the abomasum during the present trial.

Administration of each infusion level of starch during 5 consecutive days was assumed to be an acceptable sampling protocol as fecal excretion of measured parameters reached an equilibrium within 4 d of starch infusion in a pretrail under similar experimental conditions (Westreicher-Kristen et al., 2018). Nevertheless, it must be noted that fecal samples were obtained during adaptation phase to increasing amounts of starch infused and not at steady state conditions after an adaptation period. However, the animals were expected to be adapted to starch digestion by the preceding period where only moderately lower amounts of starch (250 g/d) were infused. Another critical point might be the fact that starch was infused only during 10 out of 24 h a day causing diurnal variations. However, we assumed that the diurnal flow of starch entering the large intestine does not affect the extent of starch fermentation. Fecal samples were taken frequently and can be seen as representative for the total amount excreted.

The exogenous amylase used in the herein experiment was from bacterial origin; to compensate possible reduced enzyme activity by low pH, a clearly higher enzyme concentration than the dosage recommended by the manufacturer (1,560 U/kg of grain starch) was applied.

Postruminal Disappearance of Starch

The ATTDOM decreased with increasing level of starch infusion reflecting an increased excretion of MCP, VFA, and lactate due to hindgut fermentation as well as an increased excretion of starch due to incomplete digestion in the total postruminal tract. It must be noted that the ATTDOM might be overestimated based on the fact that VFA were included in OM excretion only in the extent of which they were captured in DM analysis. The ATTDStarch decreased with increasing amounts of starch infused, from 90% to 80% with intestinal supplies of 1 and 2 kg/d, respectively. This is in agreement with results of previous experiments, where a decreasing postruminal disappearance with increasing starch flow to the small intestine was observed (Little et al., 1968; Knowlton et al., 1998; Matthé, 2001). For example, Matthé (2001) reported a decreasing disappearance of starch from the total postruminal tract from 95% to 79% with intestinal supplies of 876 and 1753 g/d in dry cows, which is comparable to the herein observed values. However, when comparing the results from different experiments, the large number of influencing factors, such as the animal, the level of feed intake, the dietary composition, the nature of the administered starch, as well as the experimental design (e.g., time for adaption to starch digestion) must be considered.

In the herein experiment, fecal excretion of microbial N and VFA was taken as indicators for starch fermentation in the hindgut as discussed previously by Westreicher-Kristen et al. (2018). The missing increase of the molar proportion of propionate with increasing starch fermentation, which is observed in the rumen, might be explained by its fast absorption from the hindgut (Bergman, 1990). The quadratic response of isovalerate and isobutyrate excretion with increments until starch infusion levels of 1,213 and 1,425 g/d, respectively, and decrements with further increasing levels might either reflect changes in the rate of absorption of these VFA or changes in the provision of branched-chain amino acids as precursor substances (Smith and Macfarlane, 1997).

Based on the linear increase of microbial N excretion, a rather constant proportion of infused starch was assumed to be fermented in the hindgut. As the ATTDNDF was not affected by level of starch infusion, an unaffected fiber fermentation in the hindgut can be assumed. Therefore, microbial N excretion increased by starch fermentation only, accounting on average by 0.12 g/100 g of additionally infused starch. Assuming an efficiency of CP synthesis in the hindgut of 1 g microbial N/100 g of starch fermented as demonstrated by starch infusion into the caecum of sheep (Ørskov et al., 1970), on average 12% of the infused starch was fermented in the hindgut. This is in close agreement with the results of our previous experiment, where 1.1 g/d microbial N was additionally excreted with 880 g/d starch infusion under similar experimental conditions, indicating that on average 12.5% of infused starch were fermented in the hindgut (Westreicher-Kristen et al., 2018). Furthermore, these values are similar to that of 14% reported by Branco et al. (1999), who infused starch hydrolysate into the abomasum of steers, which were fed fescue hay at maintenance level feeding (1.8% of BW). However, microbial N excretion was estimated based on fecal excretion of PB in the herein experiment and inaccuracies due to this method were already discussed by Westreicher-Kristen et al. (2018). Microbial N excretion averaged 9.9 g/d comprising ~21% of total fecal N output, which indeed appears to be rather low. For example, Knowlton et al. (1998) observed proportions of microbial N in feces between 45% and 57% when corn-based diets were fed to lactating dairy cows. Furthermore, according to Hungate (1966) the efficiency of hindgut CP synthesis might range between 0.9 and 1.7 g microbial N/100 g carbohydrates fermented. If the level of starch infusion somehow altered the efficiency of starch fermentation in the hindgut (i.e., by altering the rate of protein degradation in the hindgut), this might have masked differences in the extent of starch fermentation between infusion levels. Consequently, values of starch disappearance from the hindgut based on microbial N excretion as herein presented must be interpreted with care.

The linear increase of starch excretion at intestinal supplies greater than 953 g/d indicates a rather constant proportion of 29% of the additionally infused starch being excreted with feces. However, considering the decreasing ATTDStarch from 90% to 80% and assuming a constant proportion of 12% of additional infused starch being fermented in the hindgut, starch disappearance from the small intestine decreased from 78% to 68% at intestinal supplies of 953 and 1993 g/d, respectively (Figure 3). This is in line with the results from previous experiments reporting a decreasing efficiency of starch digestion with increasing intestinal supplies in cattle (Karr et al., 1966; Kreikemeier et al., 1991; Matthé, 2001).

Figure 3.

Figure 3.

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Apparent disappearance of starch from the small intestine as a function of the amount of starch infused.

Effects of Abomasally Infused Amylase on Postruminal Starch Digestion

Missing effects of exogenous amylase on fecal excretion of starch, MCP, and total VFA imply an equal extent of starch disappearance from the total postruminal tract as well as an equal extent of hindgut fermentation. Therefore, a similar disappearance of starch or end-products of starch hydrolysis in the small intestine can be assumed for both treatments as well. Consequently, pancreatic amylase activity does not seem to be the primarily limiting factor of starch digestion in the small intestine, when starch escapes ruminal fermentation in amounts up to 2 kg/d under conditions of the present experiment. Nevertheless, whether starch or end-products of starch hydrolysis were fermented in the hindgut and excreted via feces could not be differentiated from the herein experiment. Therefore, it cannot be excluded that starch hydrolysis in the small intestine was improved by amylase administration, but this effect could have been masked by an insufficient activity of other hydrolyzing enzymes or limited capacity for glucose absorption. It is unlikely that protein supply to the animals’ metabolism limited the secretion of enzymes at higher levels of starch infusion, since microbial protein production at energy maintenance feeding level exceeds considerably the protein requirement of the animal. However, it cannot be excluded that limited time for starch hydrolysis at higher infusion levels due to a rapid passage of digesta through the small intestine as discussed by Owens et al. (1986) was partly responsible for the incomplete digestion. Furthermore, reduced endogenous secretion of pancreatic amylase due to amylase infusion might have masked possible effects; however, this is not supported by observations of Remillard et al. (1990), whereat Swanson et al. (2002) observed a reduced secretion of pancreatic amylase due to an infusion of starch hydrolysate at rates of 20 to 40 g/h. Therefore, an indirect effect of amylase infusion on amylase secretion due to an increased amount of hydrolyzed starch in the lumen of the small intestine might exist. Additionally, as the activity of the exogenous amylase in the duodenal chyme was not determined in this experiment, some degradation of exogenous amylase in the abomasum can be expected but might not have limited possible enzyme effects, since the high dosage of amylase have compensated possible activity losses.

Higher proportion of butyrate and lower proportion of acetate with than without amylase administration might indicate differences in the proportion of α-linked glucose polymers (starch vs. end-products of starch hydrolysis) reaching the hindgut. Additionally, isovalerate excretion was higher with amylase administration, which is suggested to be caused by hindgut fermentation of nitrogenous compounds (Smith and Macfarlane, 1997). These might originate from amylase infusion or an enhanced flow of endogenous N to the hindgut as already suggested from the results of the previous experiment (Westreicher-Kristen et al., 2018). However, total and microbial N excretion as well as the proportion of microbial N in feces were not affected by amylase administration. Hence, an effect of exogenous amylase on endogenous N excretion seems unlikely. Assuming an unaffected excretion of dietary N, the additional amount of endogenous N excreted can be calculated as the difference of additional total N and microbial N excretion, which is equal to 0.03 g/100 g of additionally infused starch. In contrast, endogenous N excretion increased on average by 0.68 g/100 g starch infused in the preceding experiment with animals not completely adapted to starch digestion (Westreicher-Kristen et al., 2018). Therefore, it can be speculated that the greater endogenous N excretion in the preceding experiment was primarily based on adaptive mechanisms, in contrast to the better adapted animals in the present experiment.

Comparison of the Energetic Efficiency of Ruminal and Postruminal Starch Digestion

Calculations and observations of Harmon and McLeod (2001) indicate that the partial energetic efficiency of ruminal starch fermentation, defined as starch energy retained in body tissue divided by starch ME intake, is 70% to 75% of that of starch digested and absorbed from the small intestine. Latter reflects the sum of energy losses attributed to digestion and absorption (e.g., methane energy, urine energy, and fecal energy due to microbial residues from ruminal fermentation and endogenous secretion) as well as energy losses due to heat production by fermentation, digestion, and utilization of digestion end-products for maintenance and production. However, results from the present experiment demonstrate that the digestion of starch in the small intestine in amounts exceeding 1.0 kg/d is inevitable accompanied by an enhanced fermentation of starch in the hindgut. Based on our calculations only 68% to 78% of starch entering the small intestine disappeared from the small intestine, whereas 12% were fermented in the hindgut and up to 20% excreted with feces. Although less methane is produced by caecal compared with ruminal fermentation, the energetic efficiency is clearly lower, since the energy of the microbial matter is completely excreted with feces (McLeod et al., 2006). When the values of the energetic efficiency of starch digestion given by Harmon and McLeod (2001) are corrected by assuming a digestibility of starch in the small intestine of 68% instead of 78%, the increase in the partial energetic efficiency by shifting starch digestion from the rumen to the small intestine equals 32% to 44%, which is in line with the value of 42% reported by Owens et al. (1986).

However, previous experiments reported that only 35% to 38% of starch disappearing from the small intestine could be accounted for as net glucose absorption in steers due to an enhanced glucose utilization by the portal-drained visceral tissues (Huntington and Reynolds, 1986; Kreikemeier et al., 1991). Furthermore, Gilbert et al. (2015) reported that a noteworthy amount of starch disappearing from the small intestine underwent microbial fermentation in the distal part of the ileum in milk-fed calves, which might as well occur in adult cattle, and would explain the rather low net absorption of glucose in the previous mentioned experiments. Beside the limited net absorption of glucose from starch digestion in the small intestine, an increased excretion of endogenous N during phase of adaption (Westreicher-Kristen et al., 2018) as well as a reduced energy supply for ruminal microbes and therefore a reduced MCP supply to the host animal must be considered when shifting starch digestion from the rumen to the small intestine.

In conclusion, the results of the present experiment indicate a decreasing efficiency of starch digestion in the small intestine with increasing intestinal supplies as well as a considerably amount of MCP and starch excreted with feces. It can be speculated that these effects and the expected reduction of MCP supply by ruminal fermentation might override potential energetic advantages of postruminal starch digestion at amounts greater than 1 kg/d. Furthermore, the results did not indicate a limitation of starch digestion in the small intestine by pancreatic amylase under conditions of the present experiment. However, starch hydrolysis might have been enhanced by exogenous amylase but not detected by fecal parameters due to limited activity of disaccharidases or a restricted capacity for glucose absorption.

Footnotes

1

Many thanks to the Institute of Animal Nutrition of the University of Veterinary Medicine Hannover for analysis of VFA and L-lactate in feces and to Prof. Dr. Uta Dickhoefer from the Institute of Agricultural Sciences in the Tropics of Hohenheim University for analysis of purine bases. Thanks to Monika Paschke-Beese, Annette Hollmann, and Wiebke Kühl for the valuable laboratory assistance and to PD Dr. Mario Hasler for statistical advice.

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