Revista Computadorizada de Producción Porcina
Vol: 11 No. 1 2004
A NOTE ON THE DETERMINATION OF METABOLITES IN HARD FAECES OF
RABBITS
H. Phimmasam1, S. Pok2 and J. Ly3
An Giang University
Long Xuyen City, Vietnam
1
Present address:
Province Agriculture and Forestry Office
Xieng Khuang, Laos
email: hongthongp@yahoo.com
2
Present address:
UTA-Cambodia
Roulos, Cambodia
email: samkolpok@yahoo.com
3
Present address:
Swine research Institute
PO Box 1, Punta Brava
Havana City, Cuba
email: iip@enet.cu and julioly@utafoundation.org
SUMMARY
A total of nine samples of hard faeces from nine growing rabbits fed broken rice and
water spinach were used to compare two distillation procedures (Wagner-Parnas and
Tekator-type apparatus) for estimation of ammonia and total short chain fatty acids
(SCFA) concentration.
Data compared by the t-test technique did not reveal any significant difference (P>0.05)
between both examined methods: ammonia, 62.6 and 66.2 mmol/100 g DM; SCFA, 73.1
and 71.3 mmol/100 g DM for Wagner-Parnas and Tekator-type apparatus, respectively.
The fast procedure of metabolite estimation in the Tekator type apparatus is highly
recommended. However, in more modest working conditions, the Warner-Parnas
apparatus should be used instead.
Key words: ammonia, short chain fatty acids, hard faeces, rabbits
Short title: Determination of metabolites in hard faeces of rabbits
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Revista Computadorizada de Producción Porcina
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NOTA SOBRE LA DETERMINACION DE METABOLITOS EN EXCRETAS DURAS DE
CONEJOS
RESUMEN
Se usaron nueve muestras de excretas duras de conejos en crecimiento alimentados
con arroz partido y espinaca acuática, para comparar dos métodos de destilación
(aparatos de tipo Wagner-Parnas y Tekator) para la determinación de la concentración
de amoníaco y ácidos grasos de cadena corta (AGCC) totales.
Los datos comparados por la técnica t de datos pareados no revelaron diferencias
significativas (P>0.05) entre ambos métodos ensayados: amoníaco, 62.6 y 66.2
mmol/100 g MS; AGCC totales, 73.1 y 71.3 mmol/100 g MS respectivamente para
resultados obtenidos en los aparatos de tipo Wagner-Parnas y Tekator.
Se recomienda la determinación de metabolitos en el aparato del tipo Tekator debido a
su rapidez.
Palabras claves: amoníaco, ácidos grasos de cadena corta, excretas duras, conejos
Título corto: Determinación de metabolitos en excretas duras de conejos
INTRODUCTION
Ammonia and total short chain fatty acids (SCFA) are end products in hard faeces of
rabbits, and these metabolites are derived from fermentative activity in the hind gut of
the animals (Carabaño and Piquer 1998; Fraga 1998). The importance of fermentation in
the rabbit is not negligeable, particularly caecal degradation of fibre by the indigenous
microflora (Gidenne et al 1998). On the other hand, N reciclying in rabbits largely occurs
through ammonia production and reutilization (Fraga 1998).
The objective of the present note is to report a comparison made of two distillation
methods conducted to determine ammonia and SCFA concentration in hard faeces of
rabbits.
MATERIALS AND METHODS
A total of nine samples of hard faeces from nine growing rabbits fed graded levels of
broken rice and fresh water spinach offered ad libitum (Hongthong Phimmasan 2003)
were used. The samples were from five-days pooled fresh hard faeces, and after
thawing and thoroughly mixed, were suspended in distilled water in a proportion of 1:4
by weight (Pok Samkol and Ly 2001). The resulting slurry was used for ammonia and
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Revista Computadorizada de Producción Porcina
Vol: 11 No. 1 2004
total SCFA determination. In the case of ammonia, a Wagner-Parnas distillation unit was
used, after treating the sample with 40% NaOH solution added at a rate of 1:2 by
volume. Then the distillate was collected into 1% boric acid solution followed by a
titration with H2SO4 solution of approximately 0.01 N until end point attained, using a
methylene blue indicator. Another aliquot from the same sample was distilled in a
Tekator-type apparatus. Distillation time was one minute.
The SCFA concentration was determined by steam distillation too, by using the WagnerParnas apparatus (vide supra). One mL of 10 N-H2SO4 and 8 g of MgSO4.7H2O were
added to 10 mL of sample for distillation following Pennington (1952) recommendations.
Data from the two compared methods were obtained in samples titrated with NaOH
solution of approximately 0.01 N until reaching end point by using phenol red as
indicator. The other apparatus was the same distillation unit of the Tekator. Only one
minute was considered for distillate collection, and titration was conducted in the same
manner as described for the Wagner-Parnas apparatus technique.
Paired data from every sample were compared by a t test methodology (Steel and Torrie
1980). Additionally, a correlation matrix of Pearson was established, by using pH and
DM values already published (Hongthong Phimmasam 2003).
RESULTS AND DISCUSSION
Table 1 lists results from both compared methods. Data compared by the t-test
technique did not reveal any significant difference (P>0.05) between both examined
methods: ammonia, 62.6 and 66.2 mmol/100 g DM; SCFA, 73.1 and 71.3 mmol/100 g
DM for Wagner-Parnas and Tekator-type apparatus, respectively. In spite of a small
number of samples used, and the logical variability due to different treatment studied, a
rather low variability was found, this more evident for ammonia than for SCFA.
Table 1. Comparison of two methods of determination of metabolites from
hard faeces of rabbits
Distillation method
Warner
SE ±
Parnas
Tekator
Metabolites, mmol/100 g DM
Ammonia
62.63
66.28
6.85
SCFA
73.17
71.36
14.23
The possible interdependence existing among different faecal metabolites was
investigated through the technique of the Pearson correlation matrix (table 2). In this
case, average values from both methods of determination of ammonia and SCFA were
used. In this connection, there were only a highly significant (P<0.001) inverse
relationship between ammonia and DM concentration in the hard faeces of the rabbits.
The other interesting inverse interdependence observed was between faecal pH and
SCFA concentration (P<0.034).
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Table 2. Pearson correlation coefficients in hard
faeces from rabbits
pH
DM
SCFA
DM
-0.167
SCFA
-0.670
-0.431
Ammonia
0.154
-0.966
0.418
SCFA and DM correspond to short chain fatty acids and
dry matter concentration respectively
P<0.05 for r> 0.500
The Tekator (Kjeltec) distillation unit of a Tekator system has been utilized in other
occasions for ammonia determination in samples from rabbit’s hindgut (Carabaño et al
1988), although several methods based on spectrophotometric analysis and automatic
procedures have been used in experiments related to microbial digestion in the rabbit
(Weatherburn 1967; Verdow et al 1977). On the other hand, SCFA concentration in
rabbit hindgut samples are often determined by gas-liquid chromatography (see for
example Vernay 1986; Carabaño et al 1988; Gidenne and Bellier 1992; Gidenne et al
1998), as compared to the simple, not expensive method of Pennington (1952).
Perhaps the advantageous approach of the two alternative methods compared in the
present evaluation could be supported by its low cost and fast determination of the endproduct metabolites of rabbit faeces. In this connection, the fast procedure of metabolite
estimation in the Tekator type apparatus is highly recommended. However, in more
modest working conditions, the Warner-Parnas apparatus should be used instead.
ACKNOWLEDGMENTS
The authors should express their gratitude to the authorities of the An Giang University,
for the use of their laboratory facilities in order to conduct the herein reported study.
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