Les
peoxydases sont des enzymes qui oxydent un substrat défini en
présence du peroxyde d'hydrogène (H2O2). Elles sont considérées
comme des antioxydants enzymatiques.
Ces oxydoréductases reconnaissent des substrats naturels et synthétiques.
L'une des réactions utilisées pour tester leur activité
est celle utilisant le gaiacol comme substrat, générant
le tétraguaiacol absorbant la lulière à 470 nm
(voir TP S5 peroxydases, structure et fonction).
L'oxydation
de l'o-dianisidine, un substrat synthétique conduit à un produit oxydé selon la réaction:
Ecotypes du cactus Opuntia ficus indica ? الأنماط البيئية للصبار
Plusieurs
recherches scientifiques sont actuellement au Maroc
pour identifier les meilleurs écotypes. Cela a commencé
par la collecte d'écotypes de cactus dans différentes
régions marocaines, la constitution d'arboretums pour leur
évaluation et la détermination des idiotypes adaptés
à chaque situation de culture. Les meilleurs clones seront
multipliés végétativement.
Les
Peroxydases (EC 1.11.1.7), enzymes d'oxydoréduction, ont été utilisées par Khales et Baaziz comme marqueurs pour l'identification précoce des écotypes d'O. ficusindica poussant Maroc (lire d'autres détails sur: peroxydases des plantes et leurs applications).
Les écotypes à maturation tardives 'Haddaouia' and 'Moussa'
se sont montrés riches en peroxydases solubles (unités par gramme de matière fraîche) extraites des raquettes.
Les analyses de peroxydases par éléctrophorèse
sur gel de plyacrylamide suivies d'une révélation
in situ par le gaiacol (substrat) ont montré que les
écotypes d'Opuntia collectés dans les localités
proches de Marrakech (Sud du Maroc) sont caractérisés
par des isoformes de peroxydases acides liées aux parois
migrant rapidement sur les gels (voir résumé en Anglais
ci dessous)
Peroxidases(EC 1.11.1.7) were used by Khales and
Baaziz (2005, Scientia horticulturae 103, 209-218) as markers for the
identification of Opuntia ficus indica moroccan ecoyypes. Peroxidases
were extracted from fresh cladodes harvested from 9 ecotypes of the
cactus species Opuntia ficus indica Mill. growing as a collection, in
Marrakech (South Morocco).
Two enzyme fractions were obtained by a progressive solubility method
leading to soluble peroxidases (S) and ionically wall-bound peroxidases
(I). The preferred substrate for cladode peroxidases was determined
to be o-dianisidine over 4-chloro-1-naphtol and guaiacol. Contrarily
to roots, no guaiacol-based activity was found in cladodes. The late
ecotypes 'Haddaouia' and 'Moussa' showed a relatively high peroxidase
ratio S/I (units.g-1 fresh weight). When subjected to electrophoresis
on polyacrylamide gels, S and I peroxidase fractions each exhibited
two enzyme forms based on their electric charges in basic and acidic
gel media. Acidic peroxidase forms, well separated on basic gels, showed
two principal migration zones with great differences in their enzyme
activities depending on the fraction types S and I. Acidic ionically
wall-bound peroxidases exhibited fast, highly active isoforms with Rf
values 0.42 to 0.58. Basic forms, represented essentially in fractions
S and resolved on acidic gels, were typified by slow and fast isoforms
with a double banded pattern in most ecotypes. Opuntia ecotypes collected
from localities surrounding Marrakech exhibited fast basic soluble isoforms,
while distant ecotypes from Marrakech, including late ones, were typified
by a fast acidic ionically bound isoperoxidase of Rf 0.58. Possible
roles of Opuntia ficus indica peroxidases in growth and their evaluation
as markers in this cactus species are discussed in this study. For details see published work: Khales & Baaziz. 2005. Scientia horticulturae 103, 209-218.
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تبادل المعلومة عبر المواقع الإجتماعية
New investigations on the guaiacol peroxidase of Opuntia ficus indica L. and its modulation by ascorbic acid and copper. Towards an optimization of quantitative and qualitative tests.
Abstract
When compared with o-dianisidine peroxidase, guaiacol peroxidase
extracted from acetone powder prepared from cladodes of 10 ecotypes
of Opuntia ficus indica L., was found to be less active in
90% of total ecotypes. In addition, lag time kinetics altered seriously
guaiacol peroxidase tests. A part of 80% of ecotypes showed lag
time kinetics with guaiacol substrate versus 10% for o-dianisidine.
Filtered extracts on Sephadex G-25, containing low amounts of small
molecules (such as antioxidants), did not show lag phases. This
data support the hypothesis of involvement of small size antioxidants
in the appearance of lag time. At this semi-purification level,
the enzyme showed low affinity towards guaiacol. In addition, adding
ascorbic acid (AsA) as exogenous antioxidant, to the guaiacol peroxidase
reaction medium, led to the reappearance of lag time, where its
increase follows increases in AsA concentrations. A positive correlation
(coefficient 0.86) between lag times and AsA concentrations was
established. The addition of copper (known as oxidant of ascorbic
acid) in the reaction mixture affected the lag phase behaviour.
At a final concentration of 0.5-1.0 mM, Copper abolished completely
a lag time of about 120 s, allowing guaiacol peroxidase reaction
to start immediately. Electrophoretic analysis of acidic peroxidases
on 6-15% gradient polyacrylamide gels followed by in situ revelation
using reaction mixtures containing 0.5-1.0 mM Cu++, allowed the
appearance of new fast isoforms (Rf 0.66-0.68) which are absents
in controls. Contrarily, Staining gels with substrate solutions
containing 0.10 mM and 0.15 mM AsA, led to the disappearance of
isoperoxidases at zones A1 (Rf 0.29), A2 (Rf 0.52) and A4 (Rf 0.72).
A time-dependant and zone-dependant reappearances of isoperoxidases
were noted after 30 min incubation. These results are discussed
in respect of optimizing quantitative and qualitative aspects of
guaiacol peroxidase in Opuntia ficus indica L.
KOHAICH,
K. & BAAZIZ, M. 2015. New investigations on the guaiacol
peroxidase of Opuntia ficus indica L. and its modulation
by ascorbic acid and copper. Towards an optimization of quantitative
and qualitative tests. Journal of Molecular Catalysis B, Enzymatic
119, 26-32. See video
KHALES,
A. & BAAZIZ, M. 2005. Quantitative and qualitative aspects
of peroxidases extracted from cladodes of Opuntia ficus indica.
Scientia horticulturae 103, 209-218.
KHALES,
A. & BAAZIZ, M. 2004. Peroxydases d'Opuntia ficus indica
L. Activités catalytiques et propriétés
biochimiques. . In: Baaziz, M. et Hakkou, A. Actes du congrès
International de Biochimie-Forum des Jeunes Chercheurs, Marrakech,
Maroc, 142-145.
KHALES,
A. & BAAZIZ, M. 2003. Marquage des ressources génétiques
du figuier de Barbarie (Opuntia ficus indica) par le polymorphisme
des peroxydases extraites des raquettes. 3ieme congrès National
de génétique et Biologie Moléculaire, 18-20
Décembre, Tanger, Maroc, pp. 127.
KHALES,
A. & BAAZIZ, M. 2002. Isoperoxydases du cactus, Opuntia
ficus-indica.Premier congrès National de Biochimie, Faculté
des Sciences Aïn-Chock, Casablanca, 09-11 Mai 2002
Palmier dattier نخيل التمر
Date palm
Olivier الزيتون Olive
Pin d'Alep الصنوبر الحلبي
Aleppo pine
Atriplex القطف Atriplex
Céréales الحبوب
Cereals
Cactus الصبار Cactus
