4 : 277 283
ISSN-p : 2338-9427
DOI: 10.14499/indonesianjpharm25iss4pp277
Research Article
Blood samples were collected in micro mixed with 0.1% sodium carbonate solution
centrifuge tubes from individual animals by (2mL) and its optical density was measured at
retro-orbital plexus on DAY 7 for primary 680nm. The phagocytic index (K) was calculated
antibody titer and for secondary antibody titer using the equation: K= (logOD1-logOD2)/ 15
on DAY 15. Serum was separated and briefly where OD1 and OD2 are optical densities at
equal volumes of individual serum samples of 0 and 15min respectively (Shruti et al., 2009).
each group were pooled. Two fold dilutions of
pooled serum samples were made in 25L Cyclophosphamide-induced myelosupp-
volumes of normal saline in a micro titration ression
plate to which were added 25L of 1% The mice were divided into 6 groups,
suspension of SRBC in saline. After mixing, the each consisting of 6 animals. Group I: Received
plates were incubated at room temperature for 1 mL 1% SCMC for 13 days; Group II: Were
1h and examined for haemaggltination titer. given cyclophosphamide (30mg/kg BW) for
The reciprocal of the highest dilution of the 11-13 days. Group II-III: Administered
test serum giving agglutination was taken as the methanolic extract at dose 200 and 400mg/kg/
antibody titer. The mean titer values of the BW respectively for 13 days; Group IV-V:
drug and test extracts treated groups were Administered aqueous extract at dose 250 and
compared of the control (Sensitized) (Satnam 500mg/kg BW for 13 days.
Singh et al., 2012, Shinde et al., 1999). On 11th, 12th and 13th day, all the
animals of each group except control were
SRBC induced delayed type hypersen- given cyclophosphamide (30mg/kg i.p.), one
sitivity reaction hour after administration of extract. On 14th
The mice were divided into 5 groups, day blood samples were then withdrawn from
each consisting of 6 animals. Group I: Received retro-orbital plexus lysed in sodium carbonate
1mL 1% SCMC for 14 days; Group II-III: solution from all the groups and total
Administered methanolic extract at dose 200 leucocytes count was determined (Dhumal et
and 400mg/kg/ BW respectively for 14 days; al., 2013).
Group IV-V: Administered aqueous extract at
dose 250 and 500mg/kg BW for 14 days. Statistical analysis
The mice were primed with injecting Data were expressed as standard error of
20L of SRBC suspension intraperitoneally, on the means (S.E.M) of and statistical analysis
day 7 and challenged on day 14 with same was carried out employing one-way ANOVA
amount of SRBC suspension intradermally in followed by Dunnett test, which compares the
the right hind foot pad. The contra lateral paw test groups with the control groups.
received equal volume of saline, served as
control. The thickness of the foot pad was RESULTS AND DISCUSSION
measured at 24h after challenge using Phytochemical screening
speromicrometer (Gokhale et al., 2002). The preliminary phytochemical screening
of S. grandiflora flowers revealed the presence of
Phagocytic response alkaloids, saponins, terpenoids, phenolics,
The mice were divided into 5(five) flavonoids and polysaccharides as essential
groups, each consisting of 6 animals. Group I: phytochemical constituents of the methanolic
Recieved 1mL 1% SCMC for 5 days; Group II- and aqueous flowers extract. Result of
III: Administered methanolic extract at dose preliminary phytochemical screening of various
200 and 400 mg/kg/ BW. respectively for 5 extract of S. grandiflora flowers is shown in
days; Group IV-V: Administered aqueous extract table I
at dose 250 and 500mg/kg BW for 5 days.
At the end of five days, after 48h, mice Acute toxicity study
were injected via tail vein with carbon ink The LD50 of methanolic and aqueous
suspension (10L/g BW).Blood samples were extract of S. grandiflora flowers was determined.
drawn (in EDTA solution, 5L) from the retro Since no mortality was observed at 2000 and
orbital vein at 0 and 15min.; a 25L sample was 5000mg/kg respectively.
Effect of S. grandiflora on in- vivo SRBC titre was higher with methanolic extract as
specific humoral immune responses compared to aqueous extract. The production
Methanolic and aqueous extract of S. of secondary antibodies was more prominent as
grandiflora flowers on primary and secondary compare to the primary antibodies.
antibody response on H A titre are shown in Antibody molecules, a product of B
figure 1. In Contrast with control the lymphocytes and plasma cells, are vital to
methanolic and aqueous extract of S. grandiflora humoral immune responses. IgG and IgM are
increase in the primary and secondary antibody the major immunoglobulins which are involved
formation as dose dependently. Higher dose of in the complement activation, opsonization,
methanolic extract (400mg/kg BW) produced neutralization of toxins, etc. It is evidenced by
maximum enhance with 234.6721.33 and increase in the antibody titre in mice, indicated
298.6742.68 primary and secondary antibody the enhanced responsiveness of B lymphocyte
formation. Aqueous extract does not show subsets, involved in the antibody synthesis by
significant augment (apart from 500mg/kg) in the augmentation of the humoral immune
primary and secondary antibody titre. The response to SRBCs by S. grandiflora (Yadav et al.,
increase in primary and secondary antibody 2011, Gautama et al., 2009).
Figure. 1 Effect of Sesbania grandiflora on in- vivo SRBC specific humoral immune responses.
Statistical analysis was carried out employing the ANOVA followed by Dunnett test
*: <0.05, **: P<0.01 comparing with the control;
Figure 2. Effect of Sesbania grandiflora on in-vivo SRBC induced delayed type hypersensitivity
reaction. Statistical analysis was carried out employing the ANOVA followed by Dunnett test
*: P<0.05, **: P<0.01 comparing with the control;
Effect of S. grandiflora on in-vivo SRBC increased by 54.16% and 75% i.e. Most
induced delayed type hypersensitivity significantly (p<0.01) enhanced the delayed
reaction type of hypersensitive activity as compared to
Methanolic and aqueous extract of S. control (Sensitized) were observed at 24h after
grandiflora flowers on delayed type of SRBC injection in the footpad.
hypersensitive activity is shown in figure 2. Whereas at dose 500mg/kg BW,
Methanolic extract of S. grandiflora with the aqueous extract of S. grandiflora increased in
dose of 200 and 400mg/kg increased paw food pad thickness after 24h but at the dose
volume as dose dependent manner after 24 hrs, 250 mg/kg BW dose not show any significant
foot pad thickness in these group were
result. Cell-mediated immunity (CMI) involves count compare to aqueous extract which was
effectors mechanisms carried out by T lowered by cyclophosphamide, a cytotoxic
lymphocytes and their products (lymphokines). drug, indicating that the test drug can stimulate
CMI responses are critical to defense against the bone marrow activity (Damre et al., 2003).
infectious organisms, infection of foreign
grafts, tumor immunity and delayed-type CONCLUSION
hypersensitivity reactions. Therefore, increase The present investigation suggests that
in DTH reaction in mice in response to T cell methanolic extract derived from S. grandiflora
dependent antigen revealed the stimulatory flowers not only potentiates nonspecific
effect of methanolic flowers extract of S. immune response, but also improves humoral
grandiflora on T cells (Bafna and Mishra., 2004). as well as cell-mediated immunity effectively.
The effectiveness of extract can be explored for
Effect of S. grandiflora on in vivo its medical utilization in treatment of
phagocytosis immunodeficiency diseases, cancer and as
The faster removal of carbon particles combinational therapy with antibiotics.
has been correlated with the enhanced Administration of S. in human is simple as
phagocytic activity. The phagocytic activity of its seeds are used as common dietary
the eticulo-endothelium system was measured constituents in Indian household. Its reported
by the removal of carbon particles from the immunomodulatory effects warrant further
blood circulation (Miller, 1991). investigation for its use in the cases of clinical
Methanolic and aqueous extract of immunosupression.
S. grandiflora flowers on phagocytic activity
is shown in table II. Both dose 200 and ACKNOWLEDGEMENT
400mg/kg BW methanolic extract of The authors wish to thank Bansal
S. grandiflora significantly increased phagocytic College of Pharmacy, Bhopal, INDIA for
activity as dose dependent manner. These supporting research work.
group were increased by 29.03% and 46.77%
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