Download Assiut university researches Pharmacognostical Study Of Juglans

Assiut university researches
Pharmacognostical Study Of Juglans Nigra L
Black Walnut Family Cultivated In Egypt
‫درا سة ع قاق يري ة ل ن بات ال جوز األ سود ال جوج الن س‬
‫ن يجرا ال تاب ع ل ل عائ لة ال جوزي ة ال جوج الن دي سى‬
‫وال م نزرع ف ى م صر‬
Iman Sayed Ahmed Khallaf
‫إي مان س يد أحمد خ الف‬
dawood w. bishy , ahmed a. attia , samia a. youssef
‫ سام ية ع باس ي و سف‬،‫ أحمد عاب دي ن محمد‬،‫داود ون يس ب شاي‬
Abstract:
A pharmacognostical study of Juglans nigra L. cultivated in
Egypt. Juglans nigra L. (Black walnut) family Juglandaceae
(walnut family) is a deciduous tree, growing abundantly
through north temperate zone, South Europe and mountain’s
regions of North Africa. In folk medicine the plant is used as
hypotensive, hypoglycemic, for treatment of syphilis and
eczema. The stem and root bark are use as a stringent and
anthelmentic, while the dried nut used as kidney and stomach
stimulant. Literature survey showed that nothing could be
traced concerning leaves of Juglans nigra L. However,
naphthoquinones and ellagic acid are reported to be isolated
from the fruit of the plant. The present work includes : Part I :
Botanical study : Macro and micromorphology of the leaves,
stem, root, stem bark and inflorenscence are studied in both
the entire and powdered form. Part II : Phytochemical study :
That includes: 1.Preliminary phytochemical screening of the
leaves and stem bark. 2.Isolation and identification of the
flavonoids, lipids, tannins and volatile contents to elucidate
their chemical conctitution. Part III : Biological screening :
That includes : 1.Preliminary antibacterial study of different
extracts of the leaves . 2.Pharmacological study that include
the hypotensive activity of the leaves. 3.Toxicological effects
of the total extract of the leaves. Part I : Botanical study of
Juglans nigra L. Habitat : Juglans nigra L. is an erect,
monoecious, decidious tree. The male inflorescence occurs
as drooping catkins, while female flowers are erect. The fruit
is a large drupe like nut. The plant prefers temperate
temperature, alluvial soil with moderate moisture content. I.
Macromorphological study of Juglans nigra L.: a.The leaves :
The leaves are alternate, compound, imparipinnate and
exstipulate. Each leaflet is formed of about 7-15 leaflets. The
leaflet is ovate to lanceolate in the outline, with entire sinuate
margin, acute to acuminate apex and symmetric base. Both
surfaces are hairy especially the lower one. The petide and
rachis are cylinderical is shape, green in colour with hairy
surface. b.The stem : The main trunk is solid, erect,
cylinderical in shape and monopodially branched. The
terminal and lateral branches are narrower with short
internodes, covered with brown cork except the young portion
just below the apical buds. c.The root system : It was
obtained from nurseling plant that attains about 50-60cm in
height and cultivated in a flower pot. It consists of a long
fusiform tap root, bearing few lateral roots and numerous
rootlets. d.The stem bark : It occurs in flat or curved pieces.
The outer surface is greyish black to brownish grey bearning
numerous lenticeles, while the inner surface is yellowish
green in colour and longitudinally striated. e.The flower : The
flowers are unisexual, monoecious, pedicellate, apetalous
and greenish yellow in colour. Both male and female flowers
are subtended by numerous bracts and possessing the
following floral formula : P4 A3- (male) (female) f.Fruit :
Large, fleshy, indhescent, glabrous drupe like nut. It contain
one exalbuminous hard seed. II. Micromorphological study of
Juglans nigra L. 1. The lamina : •The upper epidermis : It
consists of polygonal cells with straight beaded anticlinal
walls. Stomata of anomocytic type are rare. Non glandular
trichomes are present. •The lower epidermis : It consists of
polygonal cells with wavy beaded anticlinal walls, covered
with striated cuticle. Numerous anomocytic stomata and nonglandular hair exactly similar to the upper epidermis are
abundant. •The mesophyll : heterogenous, formed of one row
of columnar palisade cells, some are large containing cluster
crystals of calcium oxalate. The spongy parenchyma contain
numerous resinous canals, in addition to schizogenous
glands at its upper part where the pericyclic fibres form a
crown over them. The pericycle consists of fibres and
parenchyma. 2. The petiole : •The epidermis : It consists of
polygonal cells with straight beaded anticlinal walls covered
with smooth cuticle and carrying non-glandular trichomes.
•The cortical tissue : It consists of outer collenchymatous cells
followed by wide zone of parenchyma cells containing
clusters of calcium oxalate. The endodermis is indistinct. •The
pericycle : it consists of isolated groups of lignified fibers that
interrupted by parenchyma cells. •The phloem : consists of
sieve tubes, companion cells, phloem parenchyma and
phloem fibers. The phloem shows a ring of schizogenous
glands at its outer part where pericyclic fibres form an arc
over them. Also it contains numerous resinous canals. •The
cambium is a wide zone of cambiform cells. •The xylem it is
formed of lignified vessels, fibres, wood parenchyma,
tracheids, tracheidial vessels and traversed by uniserriate
medullary rays. •The pith : consists of comparatively wide
zone of parenchymatous cells containing cluster crystals of
calcium oxalate . 3. The rachis : Exactly similar to the petiole
but smaller in size. 4. The stem : •Cork : It is formed of
several layers of yellowish brown suberized tabular cells. The
phellogen originates early and subepidermally. It is collapsed
and indistinct. •The cortex : formed of several layers of thin
walled parenchyma cells, containing cluster crystals of
calcium oxalate and minute starch granules. The endodermis
is indistinct. •The pericycle : It is formed of continuos zone of
lignified fibres interrupted by parenchyma. The parenchyma
surrounding the fibres contain prisms of calcium oxalate
forming crystal sheath. •The phloem : consists of soft element
in addition to phloem fibres that form crystal sheath. At the
periphery of the phloem large schizogenous glands are
present where the phloem fibres form a crown over them .
•The cambium : wide zone of cambiform cells . •The xylem :
consists of lignified vessels, fibres, parenchyma, tracheids,
trachedial vessels and traversed by lignified uni-or sometimes
biserriate medullary rays. •The pith : wide zone of
parenchymatous cells containing prisms and large clusters of
calcium oxate, in addition to numerous schizogenous glands.
5. The root : •The cork : it originates early in the pericyclic
region. It consists of several layers of tabular cells containing
reddish brown pigment. •The phelloderm : consists of wide
zone of thin walled parenchymatous cells containing
numerous starch granules, prisms of calcium oxalate, few
schizogenous glands and isolated group of lignified fibres.
•The phloem : consists of soft elements in addition to phloem
fibres and traversed by numerous uniserriate or sometimes
biserriate medullary rays. Phloem parenchyma and
meelullary rays contain numerous strach granules and prisms
of calcium oxalate. •The cambium : it is formed of several
rows of thin walled tangentially elongated cells. •The xylem, it
is formed of lignified xylem vessels, wood parenchyma,
fibres, tracheids, trachedial vessels and traversed by uni- or
biserriate medullary rays. 6. The stem bark : •The cork : it is
formed of several layers of tangentially elongated cells. In
surface view, the cells are polygonal containing dark brown
colouring matter. •The cortex : consists of several layers of
parenchyma cells containing numerous stach granules.
Groups of the sclereids are scattered in the cortex. •The
pericycle : it is formed of continous ring of lignified fibres
interrupted by parenchyma cells. The fibres form crystal
sheath. •The phloem : consists of sieve tube, companion
cells, phloem parenchyma and phloem fibres. The fibres form
crystal sheath with the surrounding parenchyma that contain
prisms of calcium oxalate. The phloem parenchyma contains
starch granules. Tannin cells are present. The phloem is
traversed by uni or biserriate non-lignified medullary rays. 7.
The inflorescence : A. The male flower : •The bract : the inner
epidermis is polygonal with straight beaded anticlinal walls,
covered with smooth cuticle. The outer epidermis is similar to
those of the inner showing anomocytic stomata. •The perianth
: the inner epidermis, the cells are polygonal, slightly
elongated with straight beaded antclinal walls and covered
with smooth cuticle. The outer epidermis similar to the inner
and showing anomocytic stomata. •Androecium : the filament
consists of polygonal cells, with straight anticlinal walls and
covered with faintly striated cuticle. The anther : •Epidermis of
anther : it is formed of polygonal cells with straight beaded
anticlinal walls. The cells are papillosed and covered with
smooth cuticle. •The fibrous layer : it consist of a single layer
of beaded bar-like thickened walls. •The pollen grains :
spherical in sphape, yellow in color with three germ-pores . B.
The female flower : •The bract : The outer epidermis :
polygonal cells with straight beaded anticlinal walls. Hairs are
not observed. The inner epidermis : similar to those of the
outer. •The perianth : The outer epidermes : polygonal with
straight anticlinal walls, covered with smooth cuticle.
Anomocytic stomata are present. Numerous non glandular
unicellular or multicellular hairs are present, the inner
epidermis are similar to those of the outer ones. The
gynaecium : •The ovary : formed of polygonal cells with
straight or slightly wavy anticlinal walls. Stomata are not
observed. The wall is hairy carrying numerous glandular
trichomes. •The style : consists of polygonal cells with straight
anticlinal walls. Stomata are not observed. •The stigma :
consists of polygonal cells with straight anticlinal walls and
slightly papillosed at its tip. C. The peduncle : •The epidermis
: polygonal cells with more or less straight beaded anticlinal
walls, showing anomocytic stomata and carrying nonglandular hairs. •The cortex : consists of several layers of
parenchyma cells containing cluster crystals of calcium
oxalate . •The pericycle : separated groups of lignified fibres
interupted by parenchyma. The fibres form an arc ove the
schizogenous glands that present at the peripheral part of the
phloem. •The phloem : it consists of sieve tubes, companion
cells and phloem parenchyma. •The xylem : formed of
lignified xylem vessels, wood parenchyma, wood fibres,
tracheids and trachedial vessels. The medullara rays being
uni-or biserriate with pitted lignified walls. However, being not
lignified in the phloem region. •The pith : consists of wide
parenchymatous zone containing cluster crystals of calcium
oxalate . Part II : Phytochemical study of Juglans nigra L.
cultivated in Egypt : Chapter I : Preliminary phytochemical
screening of the leaves and stem bark of Juglans nigra L. The
preliminary phytochemical screening of the air dried leaves
and stem bark revealed : 1.The leaves showed the presence
of carbohydrates and/or glycosides, unsaturated sterols and
/or triterpenes, flavonoids (aglycone and glycosides), low
concentration of coumarin and volatile oil and absence of
alkaloids, cardenolides, iridoids, lactones. 2.The stem bark
showed the presence of carbohydrates and/or glycosides,
tannins, unsaturated sterols and/or triterpenes and absence
of volatile oil, alkaloids, anthraquinones, iridoids, cardenolides
and cyanogenitic glycosides. Chapter II : Extraction and
isolation of the active cons-tituents of Juglans nirgra L. leaves
: •The air-dried powdered leaves are extracted with ethanol.
The ethandic extract was evaporated under reduced pressure
till dryness. The residue was suspended in water and
extracted with n-hexane, chloroform. The aqueous layer was
applied to a column of Diaion Hp-20 and eluted with water,
methanol and acetone respectively. •The n-hexane fraction
was saponified with alc. KOH to obtain the unsaponifiable
fraction, that further applied to silica gel column to isolate two
compounds J1 and J3. •The saponifiable fraction was
methylated and subjected to GLC analysis for detection of
fatty acids. •The chloroformic fraction was subjected to silica
gel column chromatography to obtain compound J4. •The
methanolic fraction was subjected to silica gel column
chromatography where three compounds J5 , J6 and J7 were
isolated . •The air dried stem bark was extracted with ethanol
and fractionated to obtain n-hexane, chloroformic and
ethylacetate fractions. •The n-hexane fraction was subjected
to silica gel column chromatography to obtain three major
compounds J1, J2, J3 , while the fractionation of ethylacetate
fraction on silica gel column two compounds J8 and J9.
Chapter III : Identification of the isolated compounds :
Investigation of fatty acids composition. The methylesters of
fatty acids were analysed by GLC together with authentic
samples. It was found that the major fatty acids are capric
(23.8), oleic (19.3) and eicosenic (20.8) . Structure elucidation
of the isolated compounds : With the help of physical,
chemical and spectroscopic techniques . Chapter IV : Study
of the volatile contents of Juglans nigra L. The steam
distillation of the leaves of Juglans nigra L. revealed the
presence of low percentage of volatile oil. The GC/MS
revealed the presence of fourty three components, from them
thirty tree components were identified. Part III : Biological
study of Juglans nigra L. cultivated in Egypt : Chapter I :
Antibacterial activity of the different extracts : The total and
aqueous extracts show significant antibacterial activity
against gram +ve and gram –ve bacteria. Chapter II :
Pharmacological activates of the different extracts :
Hypotensive activity : the total and aqueous extract of the
leaves exhibited a significant hypotensive activity. Chapter III
: Toxicological study of the extract : The leaves extract are
found to be safe LD50 are found to be 14g/kg.