J Sci Food Agric 1995,67,323-328

Koratikere T Sampatha, Christopher D Woodb* and Cadaba S Prasada

.National Dairy Research Institute, Bangalore 560 030, Indiab Natural Resources Institute, Chatham Maritime, Kent ME4 4TB, UK

(Received 1 September 1993; revised version received 17 August 1994; accepted 14 October 1994)

Abstract: The in-vitro fermentation characteristics of untreated and 50 g litre-Iurea-treated finger millet (Eleusine coracana) straw with four supplements (urea,rice bran, cottonseed and groundnut cakes) at three different ratios of straw:supplement were investigated. Gas production was greater from treated thanuntreated straw; groundnut cake was the most rapidly fermented supplementfollowed by cottonseed cake and rice bran. Urea incubated alone inhibited gasproduction. Untreated and treated straws were incubated with 22, 30 and 37 grumen degradable nitrogen from the supplements per kg organic matter digested.Significant (P < 0.05) positive interactive effects on gas production wereobserved with untreated straw at all three levels of groundnut cake supplementa-tion after 12, 52 and 166 h incubation. Similar interactions were observed forcottonseed cake supplementation of untreated straw and groundnut cake supple-mentation of treated straw, although statistical significance was not achieved forall supplementation levels at the three times for which data were analysed. Noconsistent significant interactive effects in gas production were observed betweencottonseed cake and treated straw. Rice bran inhibited gas production after 12 hbut, subsequently, had little effect on either type of straw. Urea inhibited the gasproduction from both straws at all three ratios of supplementation. Urea alsosignificantly reduced dry matter disappearance of treated straw at two of threelevels of urea supplementation. Interactive effects on gas production were mostpronounced in the early stages fermentation and appeared to be related to thehigh content of highly fermentable material particularly in groundnut cake butalso in cottonseed cake.

Key words: supplementation, straw, digestibility, urea treatment, gas production,fIImen ft:rmentation

INTRODUCflON

In India and many other tropical countries straw is amajor component of ruminant diets. Animal per-formance on such diets can be poor due to low intakesand digestibilities. The major contributing factors forthe low digestibility of straw diets are the unbalancedsupply of nutrients due to low nitrogen and high fibre

presence of antinutritional factors like silica,) 1993). Various methods

physical (Oje and Mowat 1978), chemical (Rai and

Mudgal 1986), and biological (Rai and Mudgal 1983)treatments have been developed to improve the nutri-tive value of straw, but each method has its own limi-tations for practical implementation.

An alternative approach is to provide the nutrientswhich are deficient in straw as supplements to stimulatethe fermentation of fibre by the rumen microbes(Preston and Leng 1987; Schiere et at 1989). Urea is arelatively inexpensive source of non-protein nitrogen,groundnut cake and cottonseed cake are by-productsavailable in many tropical countries which can be usedas protein supplements, and rice bran is a widely avail-able by-product which could be used with straw. PrasadTo whom correspondence should be addressed.

323Agric 0022-5142/95/$09.00 ~ 1995 SCI. Printed in Great Britain

324 K T Sampath, C D Wood, C S Pr~

et al (1994) described the use of an in-vitro fermentationtechnique (Merry et al 1991; Theodorou et al 1994) toevaluate supplementation of finger millet straw with dif-ferent proportions of a concentrate mixture. The objec-tive of the present experiment was to study the use ofthe same technique to compare different sources ofnitrogen together with rice bran as supplements tofinger millet (Eleusine coracana) straw.

tion of straw/supplement was chosen to give 30 g R[per kg of organic matter digested (OMO). The amOlwas increased by 25% (to 37 g RON kg-1 OMO) adecreased by 25% (to 22 g RON kg-1 OMO) to githe three levels used. The quantities of supplement usare given in Table 2. The RON provided by straw wnot taken into account for calculating the total RONthe diet.

Gas production methodEXPERIMENTAL

Gas production was measured with a pressure tranducer as descril)C(! t) Merry et al (1991) and The(dorou et al (1994) and modified by Prasad et ((1994). The volume of gas produced was measured ever3 h initially, then at lengthening intervals as the rate (production declined. Each fermentation was performe.in triplicate for 166 h and the residue recovered by fiItration into preweighed scintered glass crucibles, porosity P160 (British Standard grade 1), dried and weighed.

Materials

Computation of data and statistical analysis

Gas production data were corrected to a per g dry sub.strate basis. The cumulative gas produced after 12, 5:;'and 166 h were used for comparison between sub.strates. The data were analaysed to see if there werestatistically significant interactive effects between thestraws and the supplements, that is if the gas producedby the mixture was greater than predicted by summingthe gas produced by the two substrates individually.Pooled standard errors were calculated for the differ-ences between mean gas production values of the differ-ent treatments. Gas production assuming nointeractions was predicted by multiplying the weights ofstraw and substrate by their gas production when fer-

Finger millet straw, groundnut cake (GNC), cottonseedcake (CSC) and rice bran were obtained from com-mercial sources near Bangalore, India. Untreated and50 g litre-1 urea (ammoniated) finger millet straw(treated straw) were prepared from a single batch ofstarting material and were from the same batch ofmaterials as those used by Prasad et at (1991) in afeeding trial with concentrate supplementation. Thestraw was treated by spraying with 50 g litre-1 urea(analytical grade reagent, BDHjMerck Ltd, UK) at therate of I litre urea solution per kg straw. It was storedin pits for 14 days covered by polythene sheets (Prasadet at 1991).

Rumen nitrogen degradability of the types of supple-ments used was previously estimated by incubatingsimilar samples in nylon bags of pore size 40 Jl.m for 3,6,9, 15 and 24 h in fistulated cows (Sam path 1990). Theeffective degradability was calculated by iterative leastsquares procedure for a fractional outflow of 0.05 perhour (0rskov and McDonald 1979). The composition,nitrogen degradability values and calculated rumendegradable nitrogen (RDN) of the different supplementsand straws are presented in Table 1. The mean propor-

TABLE 1Composition of straw and supplements used for in-vitro and in-situ trials

RDNC

(g kg-I drymatter)

Etherextractable

material

(g kg-I dry

matter)

NDFG

(g kg-1 dry

matter)

Nitrogen(g kg-I dry

matter)

In-situ

nitrogendegradability

(%)b

Organic matter

(g kg-1 dry

matter)

ND"NO

705045

100

NO10"'554011

463

1210891811

NO

780765140220540ND

515798025

463

Untreated strawTreated strawGroundnut cakeCottonseed cakeRice branUrea

930927929936854998

.NOF, neutral detergent fibre.b Source: Sam path (1990).C RON, rumen degradable nitrogen, calculated from nitrogen content and degradabilityII NO, not determined.

.Nitrogen derived from urea treatment was taken as being 100% degradable.

of millet straw with supplements 325

TABLE 2Amounts of supplement (g per g substrate") used for in-vitro gas pro-duction studies (Supplements used singly, total substrate used was 1 g

for each in-vitro replicate)

Supplement Level of supplement (RDNb per kg OMDb)

22 30

0.23

0.29

0.60.034

0.270.340.650.04

Supplemented untreated strawGroundnut cake 0.18Cottonseed cake 0-23Rice bran 0-52Urea 0-025

Supplemented treated strawGroundnut cake 0-1Cottonseed cake 0-12Rice bran 0- 34Urea 0-012

0.170.220.510.024

0.230.290,60.034

Q Substrate = supplement plus straw (treated or untreated).b RON, rumen degradable nitrogen. OMO, organic matter digested.

alone. If the predicted levels of gas productionoutside the 95% confidence levels of measured gas

by supplemented straws these were taken assignificant interactive effects (P < 0,05). Dry

disappearance after 166 h incubation was simi-analysed.

RESULTS AND DISCUSSION

Gas production characteristics

The gas production curves for individually fermentedsupplements are given in Fig 1. The volumes

gas produced after 12, 52 and 166 h for 1 g (drybasis) of the supplements and straws when fermentedalone are given in Table 3. Gas production after 12 hincubation is an indicator of initial fermentation, after

h an indicator of the extent of fermentation which

~

"'~~::

~ -~ untreated straw

--0-- treated straw

.-UO1lDdDut cake

---cottonseed cake

~ rice bran

-urea

.s-~

"r50

O~66~66:6:'-616'-: :'- 16 I0 20 40 60 60 100 120 140 160 160

nme (hours)

may be found in vivo (Prasad etal1994) and after 166 han indicator of the end point of fermentation.

The gas production curves and gas production after12 h indicated that GNC, in particular, and CSC werefermented relatively quickly, presumably because theycontained relatively high amounts of readily ferment-able material. This can be inferred from the relativelylow neutral detergent fibre contents and is also reflectedin the ranking GNC > CSC > rice bran for in-situnitrogen degradability (see Table 1). The initial gas pro-duction rate of rice bran was similar to that ofuntreated straw. The gas production for urea alone wassignificantly (P < 0.05) lower than that without anyadded substrate, so urea appeared to be slightly tox.ic tothe microbes in the in-vitro system.

Urea treated straw had a higher gas production thanuntreated straw for much of the incubation period.Treated straw exhibited a marked lag phase for about9 h before the rate of gas production accelerated, the lagphase for untreated straw was similar but the sub-sequent rate of gas production increased relatively little.After 52 h cumulative gas production from treatedstraw was significantly higher (P < 0.05) than that fromuntreated straw. The final gas productions (166 hincubation) were, however, similar (differences not sta-tistically significant, P > 0.05). The in-vivo digestibilitiesof the untreated and treated straws were 52 and 69%,respectively (Prasad et al 1991). These relative digest-ibilities are reflected in the gas production data after52 h incubation, but not after 166 h.

Table 3 shows that dry matter disappearances over166 h fermentations were higher for GNC and CSCthan for untreated straw (P > 0.05), but the gas pro-duction data after 166 h were higher for untreated strawthan these two supplements (P > 0.05). There are

1. Cumulative gas produced for individual supplementsand straws at different times of incubation.

326K T Sampath, C D Wood, C S Prasad

7185

1241067111133.4

21924822320817514223.6

28628923924519715254

73.984.780.779553.4

100.0NO.

0.86

Untreated strawTreated strawGroundnut cake (GNC)Cottonseed cake (CSC)Rice bra.,UreaNo substrateStandard error

a ND, not determined.

several factors which may have caused this discrepancyacting individually or in combination with each other.The supplements may have contained a high content ofsoluble material which was not recovered by filtrationbut not degraded, hence not contributing to gas pro-duction. The stoichiometry of gas production may havebeen different for the supplements and straws; Menke etal (1979) indicated that gas production reflected morethe content of digestible carbohydrates than of proteinand fat. Changes in the proportions of different volatilefatty acids would also affect the amount of carbondioxide released from the buffer as the acids were neu-tralised as well as the amount of carbon dioxide rel-eased directly. Substrates with very differentcompositions may therefore have different in-vitro fer-mentation characteristics depending on whether theyare measured on the basis of their gas production or drymatter disappearance.

straw. The size of the interaction declined considerablyas fermentation progressed, but was statistically signifi-cant after 166 h incubation for both untreated andtreated straws at all three levels of supplementation.

Positive interactive effects were also observed foruntreated straw supplemented with CSC, these effectsbeing statistically significant (P < 0.05) for all three sup-plementation levels after 52 and 166 h, and for thehighest supplementation level after 12 h. In contrast,treated straw supplemented with CSC showed positiveinteractive effects with the highest level of supplementa-tion (but only achieving statistical significance after166 h incubation) and negative interactive effects withthe lowest supplementation level (but only statisticallysignificant after 12 h incubation).

Rice bran gave rise to negative interactive effects withboth untreated and treated straws after 12 h incubation,but this effect rapidly declined to the extent that no sta-tistically significant interactions were observed after52 h, interactions being generally positive by this time.Urea gave rise to large negative interactions, particu-larly in the early stages of gas production.

Interactive effects generally reflected the gas pro-duction characteristics of the supplements when fer-mented alone, the higher the gas production after 12 hincubation the more positive the observed interactiveeffects. The response was largely during the initial stagesof fermentation and declined as this proceeded. Gasproduction appeared to be responding to supplementa-tion with readily fermentable material which presum-ably acts as a ready energy source, possibly also asource of amino acids. This probably acts by stimu-lating the activity of the rumen microorganisms whichin turn would accelerate the digestion of the straw.Positive interactive effects generally increased with

Interactive effects between feeds on gas production

Table 4 presents data on the percentage differencebetween the observed gas production for straws plussupplements compared with that calculated from thegas production from straws and supplements fermentedseparately. Positive data indicate that gas productionfor the supplemented straws was higher than that pre-dicted from the separate fermentation of substrates,negative data indicate a lower gas production. Similardata are presented for dry matter disappearance(DMD).

The rapidly fermentable GNC greatly increased thegas production compared with predicted amounts after12 h when used to supplement untreated and treated

328 K T Sampath, C D Wood, C S Prasad

DMD suggesting that there was an increase in the pro-duction of microbial material and/or changes in thestoichiometry of gas production.

CONCLUSIONS

From the study it was concluded that supplementingstraw based diets with GNC and CSC can increase in-vitro gas production, presumably due to the stimulationof the activity of rumen microbes. The positive inter-active effects observed here between some supplementsand strav. Ulean that the benehts (of ill-vivo supplemen-tation may be potentially greater than predictable fromthe characteristics of the supplements and straws alone.The gas production technique could be a simple andrapid tool for studying the effect of various supplementson the digestion of crop residues and other roughagediets in use in India and elsewhere. It could improve thebasis on which rations are formulated. It must, however,be used with some caution as there could be importantdifferences between in-vitro and in-vivo conditionsunder certain circumstances.

ACKNOWLEDGEMENTS

The authors would like to thank Dr M Gill, NaturalResources Institute, for assistance in designing theexperiments and constructive comments on this paper;Dr M K Theodorou, Institute of Grassland andEnvironmental Research, Aberystwyth, for advice onthe use of the gas production technique; and Dr C Gayand Ms S Green, Natural Resources Institute, for sta-tistical analyses. This work was funded by the OverseasDevelopment Administration (ODA), United Kingdom.Drs Sam path and Prasad thank the Authorities of theIndo-Dutch project on Bioconversion of Crop Residues,Wageningen Agricultural University, The Netherlands,for funding their travel and subsistence in the UK.

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