Nutrient composition, digestion and rumen fermentation in sheep of wheat straw treated with calcium oxide, sodium hydroxide and alkaline hydrogen peroxide

Animal Feed Science and Technology 74 (1998) 315±328

Nutrient composition, digestion and rumen fermentation in sheep of wheat straw treated with calcium oxide, sodium hydroxide and alkaline hydrogen peroxide A.S. Chaudhry* Australian Tropical Dairy Institute, School of Veterinary Science and Animal Production, University of Queensland, Gatton, 4345, Australia Received 2 July 1997; accepted 23 January 1998

Abstract This study tested the effect of calcium oxide (CaO), sodium hydroxide (NaOH) and NaOH plus hydrogen peroxide (H2O2; AHP) on cell wall composition, digestion and fermentation of wheat straw (straw) in sheep. Treated straws were prepared by mixing straw either with water followed by dusting with CaO at 160 g kgÿ1 DM or with a NaOH solution alone at 3 l kgÿ1 DM to supply 80 g NaOH kgÿ1 DM (Na) or pre-soaked with Na exactly as in the previous treatment for 27 h followed by mixing with 130 g H2O2 kgÿ1 DM (AHP) for 6 h. After 14 days of storage, the treated straws and an untreated straw (U) were fed automatically every 2 h to four individually housed sheep together with a supplement in a 44 latin square experiment. Each kilogram supplement DM contained 422 g CP and 10.8 MJ ME. NDF ( p<0.001) and hemicellulose ( p<0.01) contents were significantly reduced whereas cellulose was increased ( p<0.001) in treated compared to untreated straw. ADL was reduced in Ca ( p<0.05) but increased ( p<0.05) in Na and AHP compared with U. The rumen and total tract digestibility were significantly ( p<0.001) greater in sheep fed treated compared with untreated straw. Significant differences ( p<0.05) between treatments for pH, NH3 and VFA were also observed. All treatments improved the nutritive value of straws compared with untreated through modification of cell wall with a subsequent increase in digestibility by sheep. Although the digestibility for Ca was lower than that for Na despite reduction in cell wall, its use to treat straws may be more safe and cost effective than Na. AHP was the most effective and could also improve the energy value of other low quality forages for ruminants. However, the need of AHP for a large amount of NaOH to achieve highly alkaline pH limits its farm scale application.

* Tel.: +61 754601251; fax: +61 7546011444; e-mail: [email protected] 0377-8401/98/$19.00 # 1998 Elsevier Science B.V. All rights reserved PII S 0 3 7 7 - 8 4 0 1 ( 9 8 ) 0 0 1 7 8 - 3

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Therefore, further studies should either consider reducing the amount of NaOH or finding alternative alkalis that are cost effective and user-friendly. # 1998 Elsevier Science B.V. Keywords: Calcium oxide; Sodium hydroxide; Alkaline hydrogen peroxide; Wheat straw; Sheep; Digestion; Rumen fermentation

1. Introduction Chaudhry (1998b) reported significant increases in in vitro dry matter digestibility (IVDMD) of wheat straw (straw) when it was treated separately with either calcium oxide (CaO) or sodium hydroxide (NaOH) or alkaline hydrogen peroxide (AHP). However, AHP was only effective in improving in vitro digestibility when NaOH was used as a source of alkali to regulate its pH around 11.50.2. In contrast, AHP was ineffective or depressive when CaO was used to regulate its pH around 11.5. However, CaO alone increased IVDMD to a similar extent as did NaOH and therefore, its independent use as an alternative to NaOH was considered safer and cost-effective to upgrade straw. Another series of in vitro experiments suggested that IVDMD of straw could effectively be improved if alkalis and AHP were applied under specific conditions (Chaudhry, 1997). Therefore, another study was conducted to investigate the farm-scale application of the most successful in vitro treatments as reported by Chaudhry (1998b) to improve digestion and rumen fermentation of straw in sheep. The farm-scale treatments included CaO (160 g plus 2 l water kgÿ1 DM), NaOH (80 g plus 3 l water kgÿ1 DM) and AHP (NaOH plus 130 g H2O2 kgÿ1 DM). 2. Materials and methods 2.1. Straw, chemicals and treatments Norman variety of wheat straw (straw), chopped through 8 mm sieve, was used in the following treatments. NaOH (32% w/w, specific gravity 1.35) and H2O2 (27.5% w/w, specific gravity 1.1) as solutions (Ellis and Everrard Chemical, Cheshire, UK) and CaO (BDH Chemicals) as a fine powder were used to treat straws as described below. The solution of NaOH was diluted with tap water when required. Fresh samples of about 30 kg straw DM were prepared for each of the following treatments in each of the four periods. 2.1.1. Control Untreated chopped straw served as a control. 2.1.2. CaO Chopped straw was directly blown into a Martin-Markham mixer, 60 l of water was slowly added with a hand garden sprayer and mixed with straw for 15 m. 4.8 kg CaO was dusted onto the moist straw and mixed for 45 min to distribute CaO evenly into the straw.

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The treated straw was then filled into plastic bags which were then tied up and kept at room temperature for 15 days to complete the reaction. The final concentration of CaO was 160 g kgÿ1 straw DM with a liquid:straw ratio of 2:1 and an initial pH of >13. 2.1.3. NaOH About 90 l of diluted solution containing 2.4 kg NaOH was added to the straw as described in Section 2.1.2. The straw was then mixed with the solution for about 1 h, filled into plastic bags and kept for 15 days as described in Section 2.1.2. The added amount of NaOH was 80 g kgÿ1 DM and the liquid:straw ratio was 3:1. The pH gradually rose to >12 during 1 h mixing. NaOH-treated straw served as a control for that treated with AHP. 2.1.4. AHP Another batch of treated straw was prepared as described in Section 2.1.3 by using the same amounts of straw and solutions. After 27 h pre-soaking, 14.4 l of H2O2 solution containing 3.96 kg H2O2 was added and mixed for 5 h. The pH was regularly monitored at 30 m intervals and was found to remain around 11.50.2. The remaining procedure was as described above. The amounts of added NaOH and that of H2O2 were 80 g and 130 g kgÿ1 straw DM, respectively. After 14 days of storage, all treated and untreated straws were individually mixed with molasses, Molaferm-50 (British Sugar PLC), as described by Chaudhry and Miller (1996). 2.2. Preparation of a supplement A supplement was prepared to fill the gap between nutrients required by the wethers and those supplied by the straws. Each kilogram supplement of DM contained 422 g CP and 10.8 MJ of metabolizable energy (ME) and consisted of 670, 235, 35, 35, 10 and 15 g of soybean meal, unmolassed sugar beat pulp, urea, dicalcium phosphate, sodium chloride and vitamin±mineral premix, respectively. The vitamin±mineral premix included 0.21 g of Na2SO4 for each gram of added urea to provide sufficient sulphur (S) for microbial growth in the rumen. The daily allowance of 360 g supplement DM supplied 117 g rumen degradable CP which was above the requirement as recommended by the ARC (1984) for wethers of this size. 2.3. Preparation of Cr-mordanted fibre Cr-mordanted fibre was prepared as described by Uden et al. (1980). However, the following modifications were made to suit the large scale preparation. Each kilogram of chopped straw was filled into a polyester bag and boiled for 1 h in 25 l solution containing 200 g sodium lauryl sulphate. Solution was drained off, the bag was rinsed with water using 15 m washing cycles and the residue was dried at 708C. The procedure was repeated until sufficient fibre was prepared. The dried material was then mordanted as described by Uden et al. (1980), re-dried and ground through 1 mm sieve. The chemically estimated Cr in mordanted fibre was 6.8% as opposed to 13% Cr initially added with a final binding percentage of Cr of about 53%.

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2.4. Animals, housing and feeding Four ruminal- and duodenal-cannulated SuffolkMule wethers with an initial liveweight (meanSE) of 62.52.2 kg were individually housed in metabolism crates equipped with 2-hourly feeders. The treated straws and the supplement prepared in Sections 2.2 and 2.3, respectively, were used as experimental diets. Daily DM intake (1080 g) of each wether consisted of 710 g of straw and 360 g of the supplement and matched the daily DM requirements to maintain sheep (ARC, 1984). A 44 latin square design (Plan 4.1 Type 3) as described by Cochran and Cox (1957) was used by allocating 4 diets to 4 wethers in 4 different periods. Each period lasted for 21 days with 13 days for adaptation and 8 days for collection of samples. 2.5. Marker administration and sampling procedure Steady state conditions were maintained by feeding each wether daily a dose of 20 g Cr-mordanted fibre by mixing with the daily allowance of the supplement and distributing into 12, 2-hourly feeders. After 13 days' adaptation in each period total faeces was collected for 4 days in polyester cloth bags fastened over the anus of the each sheep. Six samples of duodenal digesta representing 20 min intervals of 2-hourly feeding were collected over three days from each sheep during each period. Only two samples were collected 4 hours apart on each of the three days. Bladder catheters were used to divert the flow of digesta from duodenum to self-sealed plastic bags tied to duodenal cannula. Four samples of rumen fluid were collected from each sheep on the last day of each period at 30 min intervals during 2-hourly feeding. Faecal and duodenal samples were stored at ÿ208C until required for analysis. Rumen fluid was tested for pH, strained through 4 layers of muslin and subdivided into sterlin bottles for NH3 and volatile fatty acids (VFA). Subsamples of rumen fluid were preserved by mixing each with 1 M HCl at 1:1 ratio for NH3 and with 25% metaphosphoric acid at 5:1 ratio for VFA. 2.6. Chemical analysis Samples of untreated and treated straws, supplement, faeces and digesta collected for each diet in each period were composited and analyzed for DM, OM, NDF, ADF and ADL as described by Van Soest et al. (1991). Ash free NDF was that which disappeared on ashing NDF at 5508C. Cellulose and hemicellulose contents were calculated, respectively, by subtracting ADL from ADF and ADF from NDF. Cr in mordanted fibre, faeces and duodenal digesta were analyzed colorimetrically at 440 nm as dichromate after wet digestion as described by Owen et al. (1967). pH was determined using digital pH meter-120 (Corning Medical and Scientific Instruments, Halstead, Essex, UK). NH3 in rumen fluid was determined by nitroprusside-hypochlorite reaction on a continuous flow system using Autoanalyzer (Technicon). VFAwere determined by gas chromatography using Pye-Unicam GC-204 equipped with a flame ionization detector and a SP-4100 computing integrator. A FFAP-CB wall coated open tubular column (25 m0.45 mm o.d.) supplied by Chromopack (Holland) was used. The column oven was operated at 1208C. VFA were calculated from peak areas using the basic data system.

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2.7. Digestibility and total digesta flow Ruminal and total tract digestibilities of DM, OM, NDF, ash free NDF, ADF, cellulose and hemicellulose were calculated by the following equation and the results were expressed as g kgÿ1.   100  %Cr in feed %nutrient in faeces or digesta %digestibility ˆ 100 ÿ  % Cr in faeces or digesta %nutrient in feed Total digesta flow was calculated as described below: TDcr ˆ DDcr  DD=WD where TDcr ; mg Cr gÿ1 wet digesta; DDcr ; mg Cr gÿ1 dry digesta; DD; g dry digesta and WD; g wet digesta; TDF …g dayÿ1 † ˆ 1=TDcr  Crin where TDF; is the Total digesta flow and Crin ; the mg Cr dosed dayÿ1 : 2.8. Statistical analysis Data on OM, ash free-NDF, NDF, ADF, ADL, cellulose and hemicellulose for all straws were statistically analyzed by using analysis of variance (ANOVA) on GENSTAT. The main effect of the chemical treatments was subdivided into three sets of orthogonal contrasts to compare (i) CaO and AHP with untreated and NaOH respectively, (ii) AHP and NaOH with untreated and CaO respectively and (iii) NaOH and AHP with untreated and CaO, respectively. The contrasts were according to the orthogonal arrangement on GENSTAT. One wether in 1st, 3rd and 4th period of digestibility trial did not eat the offered food so the missing values for that wether were automatically calculated by GENSTAT during statistical analysis. The data on digestibility and digesta flow were first analyzed as a 44 latin square design to examine the effect of diet, sheep and period. Since the period effect was non-significant the data were re-analyzed as a randomized block design having 4 diets blocked over four sheep with four periods as replicates using different sheep for different period. The degrees of freedom for diet were subdivided into three sets of orthogonal contrasts as described above to study the effect of straw-based diets on the digesta flow and digestibility in rumen and total tract. The data on pH, NH3 and VFA were analyzed as described above with an additional factor of four collection times for each diet. 3. Results 3.1. General No signs of mould growth were observed in any of the treated straws. Similar softness and swelling as reported by Chaudhry (1997) for all treated straws compared with the untreated was observed. These physical changes were further enhanced on storage for AHP- followed by NaOH- and CaO-treated straws. The colours of CaO-, NaOH and

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AHP-treated straws were changed to whitish yellow, light brown and bright golden yellow, respectively, following chemical treatments. CaO-treated straw maintained its colour whereas NaOH- and AHP straws turned into dark brown and brownish black colours, respectively, by the end of each period. pH of CaO straw rose to >13 during treatment but was reduced to 11 before feeding started for digestibility trial. In contrast, pH for NaOH rose to 13 during 27 h pre-soaking and then stabilized at 9 during further storage of 10 days and thereafter. When H2O2 was added to the straw after pre-soaking with NaOH for 27 h, the pH fell from 13 to 11.3 during 6 h of mixing with a further reduction to <9 during 10 days of storage and then stabilized around 8.5 during rest of the experiment. 3.2. Chemical composition Means of DM (g kgÿ1), OM (g kgÿ1 DM) and g kgÿ1 OM of CP, NDF, ash-free NDF, ADF, ADL, cellulose and hemicellulose for samples pooled for each of the treated straws and averaged over all the periods, are presented in Table 1. DM of various straws prepared in different periods were consistent with the volumes of liquids used during their preparation and the data on DM were not statistically analyzed. DM and CP for different straws were not statistically analyzed. All other components were significantly affected ( p<0.001) by the chemical treatments. OM, NDF and ash free-NDF decreased ( p<0.001) with all chemical treatments compared with untreated straw. AHP decreased OM more ( p<0.05) than NaOH but less ( p<0.05) than CaO. NDF, Ash free-NDF and hemicellulose were significantly reduced ( p<0.001) whereas ADF and cellulose were increased ( p<0.001) in treated straws compared with untreated. ADL was reduced in CaO ( p<0.01) but increased ( p<0.01) in NaOH and AHP treated straws compared with untreated.

Table 1 Chemical composition (g kgÿ1OM, or as below) of molassed wheat straw, untreated or treated with CaO, NaOH and alkaline hydrogen peroxide (AHP) (means with standard error of difference, SED) Compositiona

Straws

ÿ1

Dry matter (g kg ) OM (g kgÿ1DM) CP (N6.25) Ash free-NDF NDF ADF ADL Cellulose Hemicellulose

SED

Untreated

CaO

NaOH

AHP

798 927a 58 841a 858a 600c 104c 451c 259a

404 790d 65 676c 692c 670b 92d 528b 29b

279 830b 63 781b 792b 734a 129a 533b 67b

248 809c 74 776b 791b 767a 115b 562a 23b

ND 7 ND 16 16 17 4 12.4 26

ND: not determined. Means with different superscripts in the same row differ significantly ( p<0.05). a OM: organic matter; CP: crude protein; NDF: neutral detergent film; ADF: acid detergent film; ADL:acid detergent lignin.

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3.3. Animal health All sheep maintained good health. However, one sheep showed signs of inappetence in periods I, III and IV without any apparent reasons. Consequently, no observation could be made on that sheep and instead missing values were fitted using least square technique. 3.4. Digestibility Mean DM, OM, NDF, ash free-NDF, ADF and cellulose digestibilities in the rumen and total tract with their SED are presented in Table 2. 3.4.1. Ruminal Significant differences between diets for ruminal DM ( p<0.02), OM and ADF ( p<0.01) and NDF, ash free-NDF and cellulose ( p<0.001) digestibility were observed. All treated straw-based diets significantly increased ( p<0.001) digestibilities. The extent of increase was more in cell wall fractions than those of DM and OM. Compared with untreated, the increase in ruminal digestibility by CaO treatment was significant for OM Table 2 Total tract and ruminal digestion in wethers of untreated and CaO-, NaOH- and alkaline hydrogen peroxidetreated (AHP) straw based diets (means with standard error of difference, SED) Straw based dietsa Parameters

Untreated

Amount of chemical (g kgÿ1DM) CaO NaOH H2O2

0 0 0

Amount of water (l kgÿ1 DM)

0.1‡

SED CaO

NaOH

AHP

160 0 0

0 80 0

0 80 130

2

3

3

Ruminal digestibility (g kgÿ1) Dry matter (DM) Organic matter (OM) Neutral detergent fibre (NDF) Ash free-NDF Acid detergent fibre (ADF) Cellulose

310a 360a 402a 406a 354a 525a

368ab 441b 584b 604b 499b 597b

396b 458b 669c 682c 539b 724c

489c 539c 710c 714c 623c 802d

44.0 30.1 28.0 24.1 38.3 35.0

Total tract digestibility (g kgÿ1) DM OM NDF Ash free-NDF ADF Cellulose

574a 597a 494a 502a 464a 588a

550a 667b 639b 659b 558b 743b

682b 699bc 718c 740c 581b 810c

715b 734c 761c 798c 670c 854c

28.3 25.9 32.8 34.4 35.7 29.3

a

Each straw based diet consisted of (kgÿ1 DM dayÿ1) 710 g of an untreated or treated straw and 360 g of a supplement. Means with different superscripts in the same row differ significantly ( p<0.05).

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( p<0.05), ADF ( p<0.01), NDF and ash free-NDF ( p<0.001) but was non-significant for DM and cellulose ( p>0.05). NaOH alone increased DM ( p<0.05), OM and ADF ( p<0.003) and NDF, ash free-NDF and cellulose ( p<0.001) digestibility compared with untreated. AHP also significantly increased ( p<0.001) digestibilities compared with untreated. Compared with CaO, the increase in digestibility by NaOH was significant for NDF and ash free-NDF ( p<0.05) and cellulose ( p<0.01) but non-significant for other components ( p>0.05). AHP also improved digestibility for DM ( p<0.05), OM and ADF ( p<0.02), NDF and ash free-NDF ( p<0.004) and cellulose ( p<0.001). Compared with NaOH, the increase in digestibility by AHP (NaOH plus H2O2) was significantly greater for DM ( p<0.09), OM ( p<0.05) and ADF and cellulose ( p<0.07) but not for NDF or ash free-NDF ( p>0.0.05). 3.4.2. Total tract Significant differences between diets for total tract DM, OM and ADF ( p<0.01) and NDF, ash-free NDF and cellulose ( p<0.001) digestibilities were noticed. There was no sheep effect ( p>0.05). All treatments were effective in improving digestibilities. Compared with untreated, CaO improved OM and ADF ( p<0.05), NDF, ash free-NDF and cellulose ( p<0.004) digestibility. However, DM digestibility was slightly reduced ( p>0.05). All digestibilities for NaOH plus H2O2 were considerably greater ( p<0.001; DM and OM at p<0.004) than those of untreated. NaOH also increased digestibility of DM and OM ( p<0.01), ADF ( p<0.017), NDF, ash free-NDF and cellulose ( p<0.001) compared with that of untreated. Compared with CaO, the increase in total tract digestibility by NaOH was significant for DM ( p<0.004), NDF, ash free-NDF ( p<0.06) and cellulose ( p<0.063) but non-significant for OM and ADF ( p>0.05). AHP also improved digestibility of DM ( p<0.001), NDF, ash free-NDF and cellulose ( p<0.01), OM and ADF (p<0.05). Compared with NaOH, the increase in digestibility by AHP was significant for ADF only (p<0.05). 3.5. Rumen fermentation characteristics Means of pH, ammonia±N and VFAs, averaged over all collection times, are given in Table 3. Significant differences ( p<0.05) in ruminal pH for diets were observed. CaO gave increased ( p<0.01) ruminal pH compared with NaOH. Differences between rest of the treatments were not significant ( p>0.05). There was no sheep effect ( p<0.05), however, time effect was significant ( pˆ0.016). Mean pH (SED 0.034) for 1000, 1230, 1500 and 1730 h were 6.92, 6.95, 6.89 and 6.83, respectively. Significant differences ( p<0.001) between diets for ammonia±N were observed. Ammonia±N for untreated-straw based diet was higher ( p<0.001) than those of treated-straw based diets. The difference between treated-straw based diets for ammonia±N were not significant ( p>0.05). The concentrations were affected by time after feeding ( p<0.05). Mean ammonia±N (SED 0.231) at 1000, 1230, 1500 and 1730 h were 7.61, 8.38, 8.30 and 8.11 mmoles/l, respectively. Isoacids (iso-butyric and iso-valeric) were significantly reduced ( p<0.05) when CaO, NaOH and AHP based diets were fed to wethers compared with untreated straw. Total VFA and molar proportions of acetic acid increased non-significantly for all treated compared with untreated straw based diets ( p>0.05). The acetic:propionic ratios were

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Table 3 Ruminal pH, NH3, digesta flow and volatile fatty fatty acid (VFA) in wethers fed untreated and CaO-, NaOHand AHP-treated straw based diets (Means with standard error of difference, SED) Parameters

Dietsa Untreated

Rumen pH Ammonia±N (mmole lÿ1) Total digesta flow (kg dayÿ1) VFA (% molar) Acetic Propionic Isobutyric Butyric Isovaleric Valeric Caproic Total VFA Acetic:Propionic

ab

SED CaO

NaOH a

b

AHP

6.9 10.1a 17a

7.0 7.5b 20b

6.8 7.5b 20b

6.9ab 7.2b 17b

0.07 0.40 1.44

65a 21.0a 1.6a 8.6a 2.2a 1.0a 0.2a 102a 3.2ab

67a 21.6a 1.0b 7.8a 1.2b 0.8bc 0.3a 116a 3.1a

68a 19.8a 1.1b 8.7a 1.2b 0.7b 0.2a 129a 3.5b

67a 20.1a 1.2b 9.1a 1.5b 0.9ac 0.2a 118a 3.4ab

1.6 0.97 0.18 0.99 0.27 0.05 0.14 14.0 0.20

a

Daily straw based diet of each wether consisted of (kgÿ1 DM) 710 g of an untreated or treated straw plus 360 g of a supplement. Means with different superscripts in the same row differ significantly ( p<0.05).

also affected but only the difference between CaO and NaOH was significant ( p<0.05). Main effects of sheep or time after feeding and their interaction were non-significant ( p>0.05). 3.6. Total digesta flow Means of total digesta flow are presented in Table 3. Total digesta flow was greater for all but AHP straw based diet compared with that of untreated. CaO and NaOH significantly increased ( p<0.053) digesta flow compared with untreated and AHP. 4. Discussion 4.1. General The absence of mould in any of the treated straws was probably due to the highly alkaline pH which inhibited any microbial deterioration of the straw. These results were in line with the findings of Chaudhry et al. (1997) who did not notice any mould in Rhodes grass being conserved after NaOH-treatment as round bales for about 6 months. The possible explanation for physical changes observed in the treated straws were already discussed by Chaudhry and Miller (1996). The non-enzymic browning may be associated with high pH (Garmo, 1986). The decline in pH of treated straws may be linked with absorption of CO2 from the atmosphere or the formation of some organic acids. Although treated straws were not actually ensiled, their storage in plastic bags may have created anaerobic conditions

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similar to the ensiled materials. The formation of acetic acid due to the hydrolytic liberation of acetyl groups between lignin and xylan, by the action of alkalis may have reduced the pH of the treated straws during storage (Jung and Fahey Jr., 1983; Garmo, 1986). The decrease was more in NaOH and AHP-treated straws compared with CaO. The decrease in pH during 6 h after addition of H2O2 to the straw pre-soaked with NaOH was perhaps due to the release of certain reactive substances from H2O2 by the action of NaOH. The pH dropped from 13 to 11.5 which was then maintained around 11.5 by the buffering action of AHP (Gould, 1985) for some time. The decrease during storage was presumably due to the formation of organic acids or the release of phenolic acids during AHP reaction. A small decrease in pH of CaO-treated straws is contrary to that of Garmo (1986) where only a combination of long storage and ensiling was able to reduce the pH. 4.2. Chemical composition The variation between OM of treated straws was just a reflection of variable amounts of chemicals added during different treatments. Similarly, the low OM of CaO-treated straw was due to the greater use of CaO compared with other treatments. The decrease in NDF and ash-free NDF of wheat straw caused by CaO, NaOH and AHP was comparable to the previous reports (Chaudhry and Miller, 1996). The decrease was probably due to the solubilization of hemicellulose. This assumption is supported by the present findings (Table 1) where substantial solubilization of hemicellulose resulted in reduced NDF and ash free-NDF. Similar losses due to AHP treatment have already been demonstrated by Kerley et al. (1988). However, the extent of decrease (<50%) was smaller than (>90%) observed in this study (Table 1). Such a high decrease was perhaps due to the high (>12) pH prior to H2O2 addition. Although pH remained around 11.50.2 following addition of H2O2, the reaction at high pH with NaOH itself may have decreased hemicellulose to a greater extent. In contrast, Kerley et al. (1987) regulated pH strictly at 11.50.2 and obtained greater values for hemicellulose. The effect of NaOH in reducing NDF and hemicellulose has been discussed by Chaudhry and Miller (1996). The increase in lignin caused by AHP is consistent with the previous findings (Chaudhry and Miller, 1996) where AHP-treated straw increased in lignin. However, it was contrary to those reported by Gould (1985); Kerley et al. (1987); Brand et al. (1989) where pronounced decreases in lignin of straws treated with AHP were observed. Chesson (1986), on the other hand, manifested that the alkali treatments operated, not by reducing or destroying the phenolics, but by breaking specific lignin±carbohydrate complexes. Increases in cellulose and ADF of treated straws agreed with the previous findings (Lewis et al., 1987; Brand et al., 1989; Meeske et al., 1993; Chaudhry, 1997). The increase in ADF may largely be due to an artifact of the analysis but may be attributed to some extent to a fermentation and loss of dry matter occurring in treated straws. The increase in crude protein levels of treated straws also supports this assumption (Table 1). Reduction in NDF of straw treated with CaO (Table 1) was comparable to that reported by Zaman et al. (1994) for Ca(OH)2. However, the decrease (246 g kgÿ1 DM or 165 g kgÿ1 OM) observed in this experiment was much greater compared with 100 g kgÿ1 DM reported by those researchers. The differences in the type and level of

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chemical, method of application and other factors may have been responsible for variable results. In general, all treatments have considerably affected OM and cell wall composition of the straw. 4.3. Digestibility The effectiveness of CaO, NaOH and AHP in improving the digestibilities of straws by sheep is well reflected by the increases observed in the ruminal and total tract digestibilities (Table 2). Total tract DM digestibilities observed for all the treated straws were generally lower than in vitro values of a different variety (Avalon) of straw treated with the same levels of chemicals in earlier experiments (Chaudhry, 1998b). Over- or under-estimation of digestibilities by in vitro methods has been documented (Garmo, 1986; Nocek, 1988; Givens and Moss, 1995). However, the trend of increase in digestibility in response to different chemicals being used in this study was comparable to that of in vitro studies by Chaudhry (1998b). The absolute increase in total tract OM digestibility observed for straws treated with 160 g CaO kgÿ1 DM in this study (70 g kgÿ1 DM) was equal to that (71 g kgÿ1 DM) observed for barley straw treated with 60 g Ca(OH)2 kgÿ1 DM by Zaman et al. (1994). They concluded that 60 g Ca(OH)2 kgÿ1 DM was the most cost-effective level to achieve optimum digestibility. Almost all total tract digestibilities of straws treated with 80 g NaOH kg ÿ1 DM were greater (Table 2) than those treated with 50 g NaOH kgÿ1 DM (Chaudhry and Miller, 1996). However, the increase was much smaller when the level of NaOH was increased from 50 to 80 g compared with that from 0 to 50 g kgÿ1 DM. Garmo (1986) reported that increases in digestibility in vitro were linear upto 100 g NaOH kgÿ1 DM whereas increases in digestibility in vivo were linear only upto 60 g NaOH kgÿ1 DM. The results are in line with those of Wilson and Pigden (1964); Jackson (1977); Wilkinson (1984); Chaudhry (1997). Table 2 shows that over 60% digestion of the straw-based diets has taken place in the rumen which is understandable because feeds high in fibre are primarily digested by the rumen microbes and the fibre particles escaping ruminal digestion are only digested through hindgut fermentation (Van Soest, 1994). The ruminal and total tract digestibilities recorded in this experiment are in general agreement to those presented by Kerley et al. (1987). However, present studies suggested that similar results could be achieved by using even half the chemicals by those researchers under slightly different conditions. Despite significant variations observed (Table 3) between ruminal pH of wethers fed with different straw-based diets, all were greater than those of 6.2±6.5, being suggested as the minimum for effective fibre digestion. Generally the optimum pH reported by Mould et al. (1983) were 6.70.15 and 6.2±6.4 for all roughage and for concentrated diets, respectively. The slightly greater pH (7.00) observed for CaO straw-based diets was perhaps due to the residual CaO in the straw. However, it did not show any ill effects on the sheep. The decrease in the rumen ammonia±N (Table 3) observed in wethers fed with any of the treated straw-based diets, was presumably a consequence of an increase in available energy. The results suggested that increased energy may have stimulated more ruminal bacteria which ultimately assimilated more ammonia for their growth with a subsequent

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decrease in ammonia concentration. The lowest level of ammonia observed for wethers fed with AHP straw-based diets further strengthened the argument that the greatest energy was made available from straw for ruminal microflora through AHP treatment. The ammonia levels were, however, comparable to published values (Mehrez et al., 1977). No particular reason of inconsistency in total digesta flows (Table 3) can be given. An increase in total VFA of wethers fed any of the treated straw were probably due to an increased fermentation rate in the rumen (Andries et al., 1987) which was also reflected by the increased digestibilities observed for the same material (Table 2). The higher molar proportions of acetic acid observed for any of the treated straws were probably the result of an effective action of chemicals on acetyl linkages which released more acetic acid and consequently has affected its concentration in the rumen as well. The decrease in ruminal molar proportions (Table 3) of isobutyric (i-C4), and isovaleric (i-C5) acids (isoacids) observed for wethers fed on any of the treated straws may be linked with the increased utilization of these acids by the fibre-degrading micro-organisms present in the rumen. The similar trend being observed in ammonia and isoacids suggested that both ammonia and isoacids were utilized by the rumen microbes during enhanced degradation of straw components following chemical treatments. The association of isoacids especially the branch chain fatty acids (i-C4, i-C5 and 2-methyl butyric) with the ruminal bacterial growth and with the plant cell wall digestion is well documented (Annison and Lewis, 1959; Russel and Sniffen, 1983; Andries et al., 1987; Marounek et al., 1989; Van Soest, 1994). The increases in digestibilities achieved by chemical treatments were presumably due to the increased susceptibility of structural carbohydrates to ruminal fermentation. Maximum digestibilities observed for AHP-treated straws indicated that AHP was most effective in removing the barriers limiting the digestion of structural carbohydrates. These barriers may have been either dismantled or modified. The modification of lignin-cell wall complex, lignin molecule itself and decrystallization of cellulose polymer during alkali (Morrison, 1983) and AHP treatment (Kerley et al., 1988) has been suggested. Removal or conversion of phenolic acids was another possibility which increased digestibility (Akin et al., 1988). The association of hemicellulose polysaccharides with cellulose in plant cell walls has been postulated (Wilkie, 1979; Jung and Fahey Jr., 1983; Chaudhry, 1997). If this is correct, microbial modification of cellulose may be limited until hemicellulose is disrupted or removed. The hemicelluloses in turn are limited in their susceptibility to microbial degradation by their precipitation in covalent linkages to phenolic acids (Chaudhry, 1998a). The lowest hemicellulose content in AHP-treated straws (Table 1) supported this hypothesis. Despite its high cellulose content, the greatest digestibilities of AHP-treated straw reflected that the crystallinity of cellulose polymer may have been reduced which resulted in increased susceptibility for microbial actions. The reduced hemicellulose and potential alteration in the three-dimensional lignin structure by AHP-treatment would be expected to make cell wall more accessible for microbial attack in the rumen. It is assumed that the inhibitory effects of phenolic acids or other products of lignin±cellulose complex on the digestion of structural polysaccharides have been removed or minimized by the chemical treatments. In summary, the chemical treatments improved the digestion in sheep of wheat straw through modification of cell wall. Although the digestibility for Ca was lower than that

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for Na despite reduction in cell wall, its use to treat straws may be safer and cost effective than Na. However, the amount of CaO (160 g kgÿ1 DM) being used in this study was very high which may limit its commercial use. Therefore, further studies should explore the possibility of reducing the amount of CaO by changing treatment conditions and perhaps without affecting the increase in digestion of straw. AHP was the most effective and could be applied to improve the energy value of other low quality forages for ruminants. However, the need for a large amount of NaOH to achieve highly alkaline pH remains a constraint in the farm scale application of AHP. Therefore, further studies should either consider reducing the amount of NaOH or finding alternative alkalis that are cost effective and user-friendly. Acknowledgements Thanks to Dr. E.L. Miller for his comments during studies at the University of Cambridge, UK and to Professor Tom Cowan for reading the manuscript. References Agricultural Research Council, 1984. Nutrient requirements of ruminant livestock. Commonwealth Agricultural Bureaux. Farnham Royal, UK. Akin, D.E., Rigsby, L.L., Theodorou, M.K., Hartley, R.D., 1988. Population changes of fibrolytic bacterium in the presence of phenolic acids and plant extracts. Anim. Feed Sci. Technol. 19, 261±275. Andries, J.I., Buysse, F.X., DeBrabannder, D.L., Cottyn, B.G., 1987. Isoacids in ruminant nutrition: Their role in ruminal and intermediary metabolism and possible influences on performance ± A review. Anim. Feed Sci. Technol. 18, 169±180. Annison, E.F., Lewis, D., 1959. Metabolism in the Rumen, Mathuen, London, UK. Brand, A.A., Frank, F., Cloete, S.W.P., 1989. Preliminary note on the utilisation of alkaline hydrogen peroxide treated wheat straw by sheep. S. Afr. J. Anim. Sci. 19(3), 136±139. Chaudhry, A.S., 1998a. Chemical and biological procedures to upgrade cereal straws for ruminants. Nutr. Abst. Rev. B. 68(5), 319±331. Chaudhry, A.S., 1998b. In vitro and in sacco digestibility of wheat straw treated with calcium oxide and sodium hydroxide alone or with hydrogen peroxide. Anim. Feed Sci. Technol. 74, 299±311. Chaudhry, A.S., 1997. Washing and filtration of wheat straw treated with sodium hydroxide alone or with hydrogen peroxide to modify cell wall composition and in vitro digestibility. Aust. J. Exp. Agric. 37, 617±621. Chaudhry, A.S., Cowan, R.T., Granzin, B.C., Klieve, A.V., 1997. Nutrient composition and in sacco digestibility of Chloris gayana treated with alkalis and a microbial inoculum. Recent Adv. in Anim. Nutr., Australia, p. 233. Chaudhry, A.S., Miller, E.L., 1996. Effect of sodium hydroxide and alkaline hydrogen peroxide on chemical composition of wheat straw and voluntary intake, growth and digesta kinetics in store lambs. Anim. Feed Sci. Technol. 60, 69±86. Chesson, A., 1986. The evaluation of dietary fibre. In: Livingstone, R.M. (Ed.), Feedstuff Evaluation and Experimental Development Service Publication 1. Rowett Res. Inst. Aberdeen, Scotland, UK. Cochran, W.G., Cox, G.M., 1957. Experimental Designs, 2nd ed. Wiley, New York. Garmo, T.H., 1986. Treatment of straw with sodium, potassium and calcium hydroxide in laboratory scale. Agric. Univ. Norway, Department of Animal Nutrition. Report no. 239. Givens, D.I., Moss, A.R., 1995. The nutritional value of cereal straws for ruminants ± A review. Nutr. Abstr. Rev. 65(11), 793±811.

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