Microscopic evaluation is indispensable in the initial iden-tification of herbs, as well as in identifying small fragments of crude or powdered herbs, and in the detection of adulterants (e.g. insects, animal faeces, mold, fungi, etc.) as well as identifying the plant by characteristic tissue features.
MICROSCOPICAL EVALUATION
Microscopic evaluation is indispensable in the initial
iden-tification of herbs, as well as in identifying small fragments of crude or
powdered herbs, and in the detection of adulterants (e.g. insects, animal
faeces, mold, fungi, etc.) as well as identifying the plant by characteristic
tissue features. Every plant possesses a characteristic tissue structure, which
can be demonstrated through study of tissue arrangement, cell walls, and
configuration when properly mounted in stains, reagents, and media. Lignin
stains red or pink with a drop of phloroglucinol and concentrated hydrochloric
acid. Mucilage is stained pink with rhuthenium red, and N/50 iodine solution
stains starch and hemicellulose blue.
The characteristic features of cell walls, cell contents,
starch grains, calcium oxalate crystals, trichomes, fibres, vessels, etc. have
been studied in details. Surinam quassia is recognized by the absence of
calcium oxalate and pres-ence of uniseriate medullary rays, crystal fibres, and
wavy medullary rays of cascara bark, lignified trichomes, and plasmodesma in
nux vomica. Stone cells are absent in the frangula bark, whereas they are
present in cascara. Presence of pith in rhizomes and absence in roots, warty
trichomes of senna, and presence or absence of crystals of aloin indicates
different varieties of aloes, glandular trichomes of mint, etc. The powder of
clove stalks contains sclereids and calcium oxalate crystals, but cloves do not
contain these two. Rauwolfia micrantho,
R. densiflora, and R. perokensis
are found to serve as an adulterant for R.
serpentine. The roots of these species can be
differentiated from R. serpentine by the presence of
sclerenchyma in the above species which is absent in R. serpentine.
The techniques like microscopic linear measurements,
determination of leaf constants and quantitative microscopy are also used in this
evaluation.
Linear measurements include size of starch grains, length
and width of fibres, trichomes, etc. The diameter of starch grains present in
ipecacuanha assists in distinguishing its varieties. The diameter of starch
grains in cassia bark distinguishes from cinnamon and detects senna stalk in
powdered senna leaf. The size of the stomata in leaves of Barosma betulina distinguishes it from other species of Barosma. The diameter of phloem fibres aids the
detection of cassia in cinnamon, and the width of the vessel helps to detect
clove stalks in powdered cloves. Measurements of diameter for the
identification of commercial starches and for the detection in them of foreign
starch are few examples of linear measurements.
Determination of leaf constants include: stomatal number,
stomatal index, vein islet, vein termination number, and palisade ratios.
Stomatal number is average number of stomata per sq. mm of epidermis of the
leaf.
Stomatal index: It is the percentage which the
numbers of stomata
form to the total number of epidermal cells, each stoma being counted as one
cell. Stomatal index can be calculated by using the following formula:
Stomatal Index (S.I.) =
S / E + S × 100
where,
S = number of stomata per unit area and
E = number of epidermal cells in the same unit area.
Timmerman (1927) and Rowson (1943) were amongst the first
few to investigate leaf drugs for stomatal number and stomatal index.
Vein-islet number: It is defined as the number of vein islets per sq. mm of the leaf
surface midway between the midrib and the margin. It is a constant for a given
species of the plant and is used as a characteristic for the identification of
the allied species. Levin in 1929 determined vein-islet numbers of several
dicot leaves.
Veinlet termination number: It is defined as the number of veinlet termination per
sq. mm of the leaf surface midway between midrib and margin. A vein termination
is the ultimate free termination of veinlet. Hall and Melville in 1951
determined veinlet termination number of distinguishing between Indian and
Alexandrian Senna.
Palisade ratio: It is defined as the average number
of palisade cells beneath each epidermal cell. Unlike vein-islet number for the
determination of which an unbroken portion of the leaf is required, palisade
ratio can be deter-mined with the powdered drug. The technique of palisade
ratio determination was introduced by Zorning and Weiss (1925) in their studies
on Compositae.
One example is vein-islet number of Alexandrian senna is
25–29.5, whereas Indian senna is 19.5–22.5. Stomatal index of Alexandrian senna
is 10–15, whereas that of Indian Senna is 14–20.
Quantitative Microscopy (Lycopodium Spore
Method)
This is an important technique employed in identification of
crude drug when chemical and physical methods are inapplicable. Using this,
one can determine the proportions of the substances present by means of the
microscope, using the Lycopodium spore method.
The powdered drugs with well-defined particles which may be
counted—for example, starch grains or single-layered cells or tissues—the area
of which may be traced under suitable magnification or the objects of uniform
thickness, and the length of which, can be measured under suitable
magnification and actual area calculated are usually evaluated using this
method.
Adulterated starchy drugs can be determined by counting the
number of starch grains per mg and calculating the amount from the known number
of starch grains per mg of the pure starch or starchy material.
Thus, if spent ginger is the adulterant, one knows that
ginger contains 286,000 starch grains per mg, and the amount used as an
adulterant can be calculated by using this figure. The percentage purity of an
authentic powdered ginger is calculated using the following equation:
[ N × W × 94,000 × 100 ]/ [ S × M × P ] = % purity of drugs
where,
N = number of characteristic structures (e.g. starch grains)
in 25 fields;
W = weight in mg of lycopodium taken;
S = number of lycopodium spores in the same 25 fields;
M = weight in mg of the sample, calculated on basis of
sample dried at 105°C; and
P = 2,86,000 in case of ginger starch grains powder.
If the material is one for which a constant is not available,
it is necessary to determine one by a preliminary experiment.
Related Topics
TH 2019 - 2026 pharmacy180.com; Developed by Therithal info.