1270
M. Arman, S.A.U. Qader / Carbohydrate Polymers 88 (2012) 1264–1271
Table 6
Chemical shifts observed in the 1H NMR and 13C NMR spectrum of the HMWPF-A of H. musciformis.
Fractions
Residue
H-1
H-2
H-3
H-4
H-5
H-6endo
H-6exo
G4S
DA
4.65614
4.91040
3.62037
4.33766
3.95706
4.47883
4.75137
4.52368
3.80448
4.67547
3.84173
4.45754
3.80784
4.41322
HMWPF-A
–
C-1
C-2
C-3
C-4
C-5
C-6
–
G4S
DA
104.569
96.986
71.5699
74.2000
80.4816
81.2853
75.9639
78.8058
76.8665
78.8880
63.3753
71.8058
–
–
Peas, induced components were not quantified but a successful
HPLC method was developed and five compounds were identi-
fied as afrormosin, anhydropisatin, Pisatin, pseudobaptigenin and
Maackiain (Arman, 2011).
Appendix A. Supplementary data
Supplementary data associated with this article can be found, in
References
Question arises why polysaccharides from H. musciformis are
effective in Chickpea tissues and not for Peas tissues. To resolve this
question it is proposed that in future experiments seaweed polysac-
charides shall be used against various cash crops i.e. Tomato, Potato,
Beans, Cotton seeds, etc. to determine and establish the activity
spectrum of these polysaccharides. It is very much desirable to
study the systemic induction of phytoalexins (isoflavonoids and
their glyco-conjugates) in cell cultures of selected host inoculat-
ing with respective fungal pathogen, pathogenic derived elicitors
and elicitor preparation from seaweed under identical conditions.
The progress and induction pattern and quantification of induced
secondary metabolites in each case shall be monitored with time
and doses. This might explain what types of isoflavonoids glyco-
conjugates are induced at what time and at what concentration.
Arman, M. (2011). LC–ESI-MS characterization of phytoalexins induced in chickpea
and pea tissues in response to biotic elicitor of Hypnea musciformis (red algae).
Natural Product Research, 25, 1352–1360.
Aziza, M., Givernuad, T., Chikhaouti-khay, M.,
& Bennasser, L. (2008). Sea-
sonal variation of the growth, chemical composition and carrageenan
extracted from Hypnea musciformis (Wulfen) Lamouroux harvested
along the Atlantic coast of Morocco. Scientific Research and Essays, 2,
509–514.
Barz, W., & Mackenbrock, U. (1994). Constitutive and elicitation induced metabolism
of isoflavones and pterocarpans in chickpea (Cicer arietinum) cell suspension
cultures. Plant Cell Tissue and Organ Culture, 38, 199–211.
Bi, F., Iqbal, S., Ali, A., Arman, M.,
& Ul Hassan, M. (2008). Induction of
secondary metabolites in chickpea, carrot and potato tissues in response
to elicitor of H. musciformis. Indian Journal of Plant Physiology, 13,
101–106.
Bi, F., Iqbal, S., Arman, M., Ali, A., & Ul Hassan, M. (2010). Carrageenan as an
elicitor of induced secondary metabolites and its effects on various growth
characters of chickpea and maize plants. Journal of Saudi Chemical Society, 15,
269–273.
Bitter, T., & Muir, H. M. (1962). A modified uronic acid carbazole reaction. Analytical
Biochemistry, 4, 330–334.
4. Conclusions
Crude and purified fractions were isolated and chemically
analyzed, maximum sugar was found as 64.3–67.6% (containing
27.6–30.2% of anhydrogalactose), very small amount of protein and
uronic acids were found, sulfate content was high ranging between
14.1% and 17.4%. Acid hydrolysis and Paper Chromatography
revealed galactose as a major component of these polysaccharides.
Purified fraction was obtained by Anion-Exchange Chromatog-
raphy and provided a homogenous fraction having mass range
molecular range ≥70,000 determined by Gel Permeation Chro-
matography. Polysaccharides of red algal plants (H. musciformis)
are charged sugars, for their identification and determination of
monosaccharide composition, it is more advisable to hydrolyze
polysaccharides under control condition using various strength of
acid, at a different range of temperature and for various length
of time to provide useful characterizable fragments and absolute
composition. On the basis of IR, 1H and 13C NMR studies, puri-
fied fraction was safely characterized as k-carrageenan. Purified
fraction-A produced intense browning as compare to the Purified
fraction (PF-B) of H. musciformis. Overall browning produced by
Chickpea was comparatively higher where as browning produced
by Peas was low. On the basis of present results, It is concluded that
seaweed polysaccharide elicitor k-carrageenan of H. musciformis
can be used as a potent plant protectant as well as the good growth
promoting agent especially for the Chickpea plant.
Bradford, M. M. (1976). A rapid and sensitive method for the quantitation of
microgram quantities of protein utilizing the principle of protein–dye binding.
Analytical Biochemistry, 72, 248–254.
Charles, W. G., llan, S., Melvyn, E. G., & Wilfred, Y. (1984). Analysis of carrageenan
from Hypnea musciformis by using k and (-carrageenan and 13C NMR spec-
troscopy. Carbohydrate Research, 129, 189–196.
Chiovitti, A., Liao, M. L., Kraft, G. T., Munro, S. L. A., Craik, D. J., & Bacic, A. (1996).
Cell wall polysaccharides from Australian red algae of the family Solieriaceae
(Gigartinales, Rhodophyta): High methylated carrageenans from the genus
Rhabdonia. Botanica Marina, 39, 47–59.
Dodgson, K. S. (1961). Determination of inorganic sulphate in studies on the enzymic
and non-enzymic hydrolysis of carbohydrate and other sulphate esters. Bio-
chemical Journal, 78, 312–319.
DuBois, M., Gilles, K. A., Hamilton, J. K., Rebers, P. A., & Smith, F. (1956). Colori-
metric method for determination of sugars and related substances. Analytical
Biochemistry, 28, 350–356.
Enkerli, J., Bhatt, G., & Covert, S. F. (1998). Maackiain detoxification contributes
to the virulence of Nectria haematococca MP VI on chickpea. Molecular Plant
Microbiology Interactions, 11, 317–326.
Jimenez-Gasco, M. M., Navas-Cortes, J. A., & Jimenez-Diaz, R. M. (2004). The Fusar-
ium oxysporum f. sp. Ciceris/Cicer arietinum pathosystem: A case study of the
evolution of plant-pathogenic fungi into races and pathotypes. International
Microbiology, 7, 95–104.
Knutsen, S., Myslabodski, B., Larsen, B., & Usov, A. (1994). A modified system of
nomenclature for red algal galactans. Botanica Marina, 37, 163–169.
Liu, C. J., Deavours, B. E., Richard, S. B., Ferrer, J. L., Blount, J. W., Huhman,
D., et al. (2006). Structural basis for dual functionality of isoflavonoid O-
methyltransferases in the evolution of plant defense responses. The Plant Cell,
18, 3656–3669.
Melotto, E., & Labavitch, J. M. (1994). Biologically active cell wall materials. Revista
Brasileira de Fisiologia Vegetal, 6, 75–82.
Mou, H., Jiang, X., Liu, Z., & Guan, H. (2004). Structural analysis of kappa-carrageenan
oligosaccharides released by carrageenase from marin Cytophaga MCA-2. Journal
of Food Biochemistry, 28, 245–260.
Acknowledgments
Nicholson, R. L., & Wood, K. V. (2001). Phytoalexin and secondary products, where are
they and how can we measure them. Physiology and Molecular Plant Pathology,
59, 63–69.
Qari, R. (1988). Seasonal changes in biochemical composition of seaweeds from
Karachi coast, Pakistan. Journal of Scientific Industrial Research, 31, 94–96.
Authors are thankful to Sadaf Noureen & Mr. Muhammad Sadiq
Ali, Technical Officer for his assistance in the art work & laboratory
work.