Amino acid composition of two Spondias gums and its Elution profiles

Amino acid composition of two Spondias gums and its Elution profiles

AFINIDAD
REVISTA DE QUÍMICA TEÓRICA Y APLICADA
EDITADA POR LA ASOCIACIÓN DE QUÍMICOS E INGENIEROS
DEL INSTITUTO QUÍMICO DE SARRIÁ
Amino acid composition of two Spondias
gums and its Elution profiles
María Leal, Olga Beltrán, Gladys León de Pinto*.
Centro de Investigaciones en Química de los Productos Naturales, Facultad de Humanidades y Educación,
La Universidad del Zulia, Apartado 526, Maracaibo-Zulia state, Venezuela
Composición de aminoácidos de dos gomas de Spondias y sus perfiles de elución
Composició d’aminoàcids de dues gomes de Spondias i els seus perfils d’elució
Recibido: 21 de octubre de 2005; revisado: 5 de octubre de 2006; aceptado 19 de octubre de 2006
Afinidad (2007), 64 (527), 38-40
Amino acid composition of two Spondias
gums and its Elution profiles
María Leal, Olga Beltrán, Gladys León de Pinto*.
Centro de Investigaciones en Química de los Productos Naturales, Facultad de Humanidades y Educación,
La Universidad del Zulia, Apartado 526, Maracaibo-Zulia state, Venezuela
Composición de aminoácidos de dos gomas de Spondias y sus perfiles de elución
Composició d’aminoàcids de dues gomes de Spondias i els seus perfils d’elució
Recibido: 21 de octubre de 2005; revisado: 5 de octubre de 2006; aceptado 19 de octubre de 2006
RESUMEN
Se determinaron los perfiles de elución y la composición
de aminoácidos de las gomas de dos especies de
Spondias (Anacardiaceae). La cromatografía de exclusión por tamaño monitoreada mediante la medición de
los carbohidratos y proteínas totales, mostró la presencia de distintas fracciones. Existen algunas evidencias
que podrían indicar la presencia de un arabinogalactan
(AG) y un arabinogalactan – proteína (AGP) en las gomas
estudiadas. La composición de aminoácidos de ambas
gomas mostró la presencia de hidroxiprolina, ácido aspartico, ácido glutámico, serina, prolina, treonina y alanina.
La hidroxiprolina es el aminoácido mayoritario en ambas
gomas, tal como se ha reportado para gomas de diversas especies de Acacia dentro de la familia Leguminosae.
Palabras claves: Perfiles de elución. Spondias purpurea.
Spondias mombin. Exudados gomosos.
SUMMARY
The elution profiles and amino acid composition of two
gums from Spondias species (Anacardiaceae) have been
determined. Size exclusion chromatography, monitored
by the measurement of total carbohydrate and protein
showed diferent fractions in the gums from S. purpurea
and S. mombin. There are some evidences which may
indicate the presence of arabinogalactan (AG) and arabinogalactan-protein (AGP) in those gums. Amino acid
composition of both gums showed that hydroxyproline,
38
aspartic acid, glutamic acid, serine, proline, threonine
and alanine are present. Hydroxyproline is the main amino acid, of both gums as has commonly been associated with Acacia gums and other genera of Leguminosae.
Key words: Elution profile. Amino acid composition.
Spondias purpurea. Spondias mombin. Gum exudates.
RESUM
Es determinen els perfils d’elució i la composició d’aminoàcids de les gomes de dues espècies de Spondias
(Anacardiaceae). La cromatografia d’exclusió per mida,
monitorada mitjançant la mesura dels carbohidrats i les
proteïnes totals, mostra la presència de diverses fraccions. Hi ha algunes evidències que semblen indicar la
presència d’un arabinogalactan (AG) i d’un arabinogalactan–proteína (AGP) en les gomes estudiades. La composició d’aminoàcids d’ambdues gomes mostra la
presència d’hidroxiprolina, àcid aspàrtic, àcid glutàmic,
serina, prolina, treonina i alanina. La hidroxiprolina és l’aminoàcid majoritari en ambdues gomes, tal com s’ha
comunicat per a diverses espècies d’Acacia dins de la
família Leguminosae.
Mots clau: Perfils d’elució. Spondias purpurea. Spondias
mombin. Exsudats gomosos.
* To whom correspondence may be addressed.
INTRODUCTION
Spondias purpurea and Spondias mombin (Anacardiaceae),
species disseminated in Venezuela, produce gum exudates very clear and soluble in water. Some analytical data
and structural features of these gums have been already
(1, 2)
published . They contain galactose as a main sugar component, as has been reported for many gums from other
(3)
(4)
species included in the genera Parkia , Grevillea and
(5)
Acacia . Arabinose, the second important sugar is much
higher in S. mombin than S. purpurea. They have also mannose and rhamnose; xylose is only present in S. purpurea
gum. Both gums contain nitrogen which is higher for S.
mombin gum. It is important to investigate these gums as
a possible source of arabinogalactan (AG) and arabinogalactan-protein (AGP). The AGPs, widely distributed throughout the plant kingdom, are present in leaves, stems, roots,
(6, 7)
floral parts and seeds . There is evidence that some AGPs
are involved in cell-cell signaling and cell-matrix interactions during tissue development, cell proliferation, and soma(6, 8)
tic embryogenesis . They have also been reported real
and potential functions as emulsifiers in the food industry
(7)
and as potential immunological regulators for human health .
This communication reports the amino acid composition
and the elution profile of the two Spondias gums which
have not been published.
MATERIALS AND METHODS
TABLE I
Gradient system used in the determination of
amino acids of S: mombin and S. purpurea gums
S. mombin y S. purpurea
Time
Flow
(ml/min)
Solvent A
(%)
Solvent B
(%)
0
0.3
0
100
10
0.6
0
100
20
0.6
95
5
40
0.6
95
5
in vacuum and dissolved in phenylisothiocyanate in order
to obtain the corresponding phenylthiocarbamil amino
acids. The derivative products were injected to a NovaPak C 18 column (Water), adapted to an UV detector (254
nm). It was used as mobile phase a mixture of an aqueous
solution of trihydrate sodium acetate (0,19 g per L) and
triethylamine (0.5 mL per L) at pH 5.7 (Solvent A) and acetonitrile-water (60/40) (Solvent B). An appropriate gradient
system was used for the elution, Table I. The nitrogen contents were determined by kjeldahl method.
Origin and purification of the gum specimens
Gums specimens from Spondias purpurea, Spondias mombin, known as ciruelo de huesito and jobo in Zulia state,
Venezuela, South America, were collected by the authors
in dry season, January-June, 2004. The identification of voucher specimens was confirmed by Lic. Carmen Clamens,
a botanical taxonomist of Universidad del Zulia, Venezuela.
The gum exudate, collected two weeks after the exudation, was purified as follows. Aqueous sample solutions
(3%) were prepared and filtered through muslin and then
through whatman No. 1 and No. 42 filter papers, dialyzed
for two days against tap water, and freeze-dried.
ANALYTICAL METHODS
Elution profiles of the original gums
The original sample (150 mg) was dissolved in deionized
water (10 mL). The elution profile was determined on a glass
column (1,75 x 65 cm) packed with Sephacryl S-400-HR
(20.000-8.000.000 Da for globular proteins ), NaCl (0.1 M)
solution was used as eluent. The carbohydrate profiles
were obtained, after the application of phenol-sulfuric acid
method at 490 nm, while the protein contents were measured at 220 nm.
RESULTS AND DISCUSSION
The sample (40 mg) was hydrolyzed with 6 M HCl (24 h,
110 °C, under nitrogen atmosphere). The residue was dried
The elution profiles of the original gums from Spondias purpurea, and Spondias mombin (Fig. 1 and 2) respectively,
obtained by size exclusion chromatography (SEC) at
atmospheric pressure, were monitored by the measurement of total carbohydrate and protein.
Fraction 1 (Ve: 48-100 mL), Fraction 2 (Ve: 100-130 mL)
Fraction 1 (Ve: 48-68 mL), Fraction 2 (Ve: 68-93 mL),
Fraction 3 (Ve: 93-150 mL)
Figure 1. Elution profile of the original gum from Spondias
purpurea.
Figure 2. Elution profile of the original gum from S mombin L.
Amino acid composition
39
TABLE II
CONCLUSION
Amino acid composition of S. mombin
and S. purpurea gum exudates.
*Amino acid (%)
Aspartic acid
Glutamic acid
Hydroxyproline
Serine
Glicine
Hystidine
Arginine
Threonine
Alanine
Proline
Tyrosine
Valine
Methyonine
Cystine
Isoleucine
Leucine
Phenylalanine
Lysine
Total
Nitrogen content, (%)
+
Hence Protein content, (%)
S. mombin L.
S. purpurea
10,73
10,29
18,55
7,26
4,32
0,88
1,76
5,74
5,77
5,19
4,61
3,35
1,27
1,89
2,97
6,98
5,25
3,18
100,00
3,03
20,65
3,56
4,96
55,16
6,94
1,38
0,35
0,00
4,65
4,24
5,36
4,08
2,89
0,00
0,00
1,57
3,35
1,50
0,00
100,00
0,37
2,34
* Composition calculated on basic of total protein content.
The protein content was calculated from N% x 6.25.
+
S. purpurea gum (Fig. 1) shows two population. The first
fraction, the major one (48 – 100 mL), contains both carbohydrate and protein and it may correspond to a possible glycopeptide or glycoprotein, and the second fraction
(100 – 130 mL) with a relatively low molecular weight constituted by carbohydrate is possibly an oligosaccharide.
The elution profile of the original gum from Spondias mombin shows three fractions. The two first fractions (48 – 68
mL), (68 – 93 mL), are both constituted by carbohydrate and
protein. These populations may be represented by two complex arabinogalactan-proteins (AGP) which may have a close molecular weight according to the elution volume. It is
also possible the presence of arabinogalactan (AG). It has
(9, 10)
been reported that gums are a source of AG and AGP .
The third fraction (93 – 150 mL), of lower molecular weight
may correspond either to a glycopeptide or glycoprotein.
The amino acid composition of S. purpurea and S. mombin gums, Table I, shows that hydroxyproline, aspartic
acid, glutamic acid, serine, proline, threonine and alanine, are mainly present in the investigated gums.
Hydroxyproline is the main component in both gums but
in the case of S. purpurea gum, this amino acid is very
high (>50%) and the other amino acids shown in Table I,
are either in low proportion or absent. The presence of
hydroxyproline as the main amino acid component in S.
purpurea and S. mombin gums is an interesting feature
that has been observed in many Acacia gums, i.e. gums
from A. senegal, A. goetzzi, A. gerrardii, A. seyal and A.
(5, 11)
robusta . Although the Combretaceae and Sterculiaceae
gums contain very low proportions of hydroxyproline. The
principal amino acids ,common to these species, are aspar(12)
tic acid, alanine and glycine . It has also been demonstrated the hydroxyproline participation in the protein-car(13, 14, 5)
.
bohydrate linkage
40
The elution profiles of the gums from S. purpurea and S. mombin suggest the presence of AG and AGP in their structures.
Hydroxyproline, the main amino acid component in both
gums, is a feature that must be investigated in order to know
its possible role in the complex arabinogalactan-protein.
The profile elutions of the gums after basic and enzymatic hydrolyses are in progress in order to complete the study
of their polidispersity.
ACKNOWLEDGMENTS
The authors would like to thank the University of Zulia
Research Council (CONDES-LUZ) for financial support and
to FUNDADESARROLLO (LUZ), institution that has helped
a lot to the formation of the novel researches.
BIBLIOGRAFÍA
(1)
. Gutierrez, O; Martínez, M; Sanabria, L; León de Pinto, G;
Igartuburu, J.: «1D- and 2D NMR spectroscopy studies of
the polysaccharide gum from Spondias purpurea var. lutea».
2005. Food Hydrocolloids, 19, 37-43.
(2)
. León de Pinto, G; Martínez, M; Beltrán, O; Rincón, F;
Igartuburu, M; Rodríguez, F.: «Structural investigation of
the polysaccharide of spondias mombin gum». 2000.
Carbohydrate Polymers, 43, 105-112.
(3)
. León de Pinto, G; Martínez, M; Beltrán, O; Torres, M;
Rivas, C.: «Spectral study of Crabon-13 NMR of three Parkia
gums». 1997. Ciencia, 5 (1), 37-43.
(4)
. Anderson, D.; León de Pinto, G.: «Gum Exudate from the
Genus Grevillea (Proteaceae)». 1982. Carbohydrate
Polymers, 2, 19 – 24.
(5)
. Islam, A.; Phillips, G.; Sljivo, A.; Snowden, M.; Williams,
P. A.: «Review of recent developments on the regulatory,
structural and functional aspects of gum Arabic». 1997.
Food Hydrocolloids, 11, 4, 493-505.
(6)
. Gaspar, Y.; Johson, K.; Mc.Kena, J.; Bacic, A.; Schultz,
C.: «The complex structures of arabinogalactan-proteins
and the Journey towards understanding function». 2001.
Plant Molecular Biology, 47, 161-176.
(7)
. Showalter, A.M.: «Arabinogalactan-proteins: structure,
expression and function». 2001. CMLS Cellular and
Molecular Life Sciences, 58, 1399-1417.
(8)
. Majewska, A.; Nothnagel, E.: «The multiple Roles of arabinogalactan- proteins in Plant Development». 2000. Plant
Physiology, 122, 3-9.
(9)
. Fauconnier, M.; Blecker, Ch.; Groyne, J.; Razafindralambo, H.; Vanzeveren, E.; Marlier, M.; Paquot, M.:
«Characterization of two Acacia gums and their fractions
using a Langmuir Films Balance». 2000. Journal of
Agricultural and Food Chemestry, 48, 2709-2712.
(10)
. Siddig, N.; Osman, M.; Al Assaf, S.; Phillips, G.; Williams,
P.: «Studies on Acacia exudates gums. Part V. Distribution
of molecular componets in Acacia seyal in relation to Acacia
Senegal». 2005. Food Hydrocolloids, 19, 679-686.
(11)
. Beltrán, O.; León de Pinto, G.; Martínez, M.; Rincón F.:
«Comparación de los datos analíticos de las gomas de
Acacia macracantha, Acacia tortuosa y otras Gummiferae».
2005. Afinidad, 62 (517), 237-241.
(12)
. Anderson, D.; Howlett, J.; Mc Nab, C.: «Amino acid composition of Gum exudates from some African Combretum,
terminalia and Anogeissus species». 1987. Phytochemestry
26, 309.
(13)
. Akiyama, Y.; Shigeru, E.; Kato, K.: «Gum Arabic is a Kind
of arabinogalactan-protein». 1984. Agricultural Biological
Chemistry, 48, (1) 235-237.
(14)
. Shirley, C.; Alistar, S.: «Structural studies of an arabinogalactan-protein from the gum exudates of Acacia robusta». 1984. Carbohydrate Research, 133, 105 – 123.