Carbohydrate Polymers 209 (2019) 400–408

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Carbohydrate Polymers
journal homepage: www.elsevier.com/locate/carbpol

Thiol modified Moringa gum – A potential bioadhesive polymer T

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Pankaj Grewal, Jyoti Mundlia, Munish Ahuja
Drug Delivery Research Laboratory, Department of Pharmaceutical Sciences, Guru Jambheshwar University of Science and Technology, Hisar, 125001, India

A R T I C LE I N FO A B S T R A C T

Keywords: In the present study Moringa gum was modified by thiolation to enhance its mucoadhesive potential. Thiolation
Moringa gum was accomplished by esterification with thioglycolic acid. Thiolated Moringa gum was characterized by FT-IR,
Thiolation TGA, DSC, XRD and SEM-EDX studies. Modified gum was found to have 0.956 ± 0.024 mM of thiol groups/g as
Mucoadhesion determined by Ellman’s method. The greater force of detachment of polymer compacts of modified gum from the
Buccal tablet
mucin discs as compared to native gum indicates its enhanced mucoadhesive potential. The thiolated Moringa
gum was formulated into buccal tablets using metronidazole benzoate as the test drug. A comparative evaluation
of buccal tablets revealed 1.5-fold higher ex vivo bioadhesion time of buccal tablets of modified gum than the
native gum. The results of in vitro release studies showed that modified gum tablets provided a sustained release
of metronidazole over 24 h of the study following First-order kinetics with diffusion mechanism of release.

1. Introduction studies thiol modification of chitosan (Mahmood, Lanthaler, Laffleur,

Natural polymers and its modified derivatives have been extensively
used in many applications in the food, construction, electronics (Pandey
& Ramontja, 2016a; Pandey, 2016), biomedical (Pandey & Nanda,
2015), pharmaceutical (Pandey & Ramontja, 2016b), water purification
(Pandey, 2017) and engineering industries (Pandey & Tiwari, 2015). In
view of its potential advantage of biocompatibility, biodegradability
and easy availability, they are widely accepted in pharmaceutical in-
dustry (Pawar, Kamat, & Choudhary, 2015). The utility of these poly-
mers can further be enhanced by physically or chemically modifying
them (Makhado, Pandey, Nomngongo, & Ramontja, 2017; Makhado
et al., 2018). A number of approaches such as grafting (Bhattacharya &
Mishra, 2004), cross-linking (Banegas, Zornic, Borges, Porto, & Soldi,
2013), thiolation (Sharma & Ahuja, 2011), carboxymethylation (Rimpy,
Abhishek and Ahuja, 2017) etc. have been explored to meet tailor made
specifications. Amongst these thiolation is a promising approach which
imparts mucoadhesive characteristics on the modified derivatives. A
number of mucoadhesive approaches are based on non-covalent
bonding such as Van der’s Waal forces, hydrogen bonds and ionic in-
teraction which are weak in nature and thus doesn’t provide sufficient
residence time in the alimentary tract (Bernkop-Schnurch, 2005).
However, modification of polymers by introduction of thiol groups to
the polymer provides higher cohesive properties. The thiol groups form
disulfide linkage between the polymer and mucus layer improving its
bioadhesion (Bernkop-Schnurch, Kast & Richter 2001). During earlier
Huck, & Bernkop-Schnürch, 2017), tamarind seed polysaccharide
(Kaur, Yadav, Ahuja, & Dilbagi, 2012), pectin (Sharma & Ahuja, 2011),
alginate, hyaluronic acid (Kafedjiiski et al., 2007), xanthan gum
(Bhatia, Ahuja, & Mehta, 2015), gellan gum (Yadav, Ahuja, Kumar, &
Kaur, 2014), - Psyllium husk (Bhatia & Ahuja, 2013) etc., have been
carried out to improve their mucoadhesive characteristics.
Moringa gum is a natural gum obtained from Moringa oleifera (fa-
mily: Moringaceae). Moringa gum comprises of L-arabinose, L-glu-
coronic acid, L-galactose and L-rhamnose. The gum has actually been
investigated as binder, disintegrant, release retardant for pharmaceu-
tical applications (Panda, Choudhury, Yedukondalu, Si, & Gupta,
2008). Moringa gum has hydroxyl groups to the side chain which are
able to interact with mucus contributing to its mucoadhesion ability.
Grafting of Moringa gum with (N-vinyl-2-pyrrolidone) has already been
investigated in order to improve its bioadhesion. The present study is
the first ever report on thiolation of Moringa gum which has been car-
ried out with the objective to enhance its mucoadhesive properties.
The thiolation of Moringa gum was carried out by esterification of
the gum with thioglycolic acid. The thiolated Moringa gum was char-
acterized by FT-IR, TGA, DSC, XRD, SEM studies and for its compres-
sion nature. The thiol substitution on the Moringa gum was estimated by
Ellman’s method. The mucoadhesive property of the thiolated Moringa
gum was comparatively evaluated with native gum using texture ana-
lyzer. Thiolated Moringa gum was formulated into buccal tablets using
metronidazole benzoate as the test drug. Further, the buccal tablets

⁎
Corresponding author at: Drug Delivery Research Laboratory, Department of Pharmaceutical Sciences, Guru Jambheshwar University of Science and Technology,
Hisar, Haryana, 125001 India.
E-mail address: munishahuja17@yahoo.co.in (M. Ahuja).

https://doi.org/10.1016/j.carbpol.2018.12.100
Received 31 May 2018; Received in revised form 20 December 2018; Accepted 31 December 2018
Available online 02 January 2019
0144-8617/ © 2019 Elsevier Ltd. All rights reserved.
P. Grewal et al.
were evaluated for friability, thickness, content uniformity, average
weight, ex vivo bioadhesion and in vitro release behaviour.
2. Experimental
2.1. Materials
Moringa oleifera gum (mol wt. 190 kDa) was procured from
Nutramine Life Sciences (Delhi, India). Metronidazole benzoate was
received as gift sample from Ranbaxy Laboratories Ltd. (Gurugram,
India). Thioglycolic acid, citric acid and hydrochloric acid were pur-
chased from SD Fine-Chem Ltd. (Mumbai, India). L-Cysteine and
Ellman’s reagent (5′5′-dithiobis-(2-nitrobenzoic acid) or (DTNB),
mucin, acetone, disodium hydrogen phosphate and methanol were
procured from HiMedia Laboratories Pvt. Ltd. (Mumbai, India).
Polyethylene glycol-4000 was obtained from Merck Ltd. (Mumbai,
India).
2.2. Thiol functionalization of Moringa oleifera gum
Thiol modification of Moringa gum was carried out by the ester-
ification of Moringa gum with thioglycolic acid under acidic condition
(Dicharry et al., 2006). Briefly, 7.59 g of thioglycolic acid (80%, v/v)
and 4 ml of hydrochloric acid (7 N) were added to the aqueous dis-
persion of Moringa gum (2%, w/v) in hot water (50 ml). The tempera-
ture of the reaction mixture was raised to 80 °C, and the reaction was
continued till 150 min. The reaction mixture was then cooled and
precipitated by adding acetone (500 ml). The precipitated thiol func-
tionalized Moringa gum was further washed with2 × 200 ml of acetone
and dried in oven at 50 ± 2 °C for 1 h.
2.3. Characterization of thiolated Moringa oleifera gum
2.3.1. Estimation of thiol substitution
Ellman’s method was employed to determine the number of thiol
groups/g of the modified gum (Bernkop-Schnurch, Hornof, & Zoidl,
2003). Solutions of Moringa gum and modified Moringa gum (50 mg)
prepared in 1 N NaOH (25 ml) were diluted with an equal volume of
0.5 M phosphate buffer (pH 8.0). The above reaction mixture was al-
lowed to react with 5 ml of Ellman’s reagent (DTNB, 0.03% w/v) in
0.5 M phosphate buffer (pH 8.0) at room temperature for 2 h. The thiol
group substitution was determined by measuring the absorbance of the
reaction mixture at 450 nm, calculated using the calibration curve of L-
cysteine solution prepared in the same manner.
2.3.2. Fourier transform infra-red spectroscopy (FT-IR)
Fourier-transform infrared (FT-IR) spectrum of samples were re-
corded by FT-IR spectrophotometer (IR-Affinity-I, Shimadzu, Japan)
using KBr disc method in the range of 4000-500 cm−1. The samples of
Moringa gum and thiolated Moringa gum in powder form were tritu-
rated with KBr using agate pestle and mortar, and the blend was then
compressed into disc using IR hydraulic press (IR Hydraulic Press CAP-
15 T, PCI Analytics, Mumbai, India) at a pressure of 75 kg/cm2 for 30 s.
2.3.3. Thermogravimetric analysis
Moringa gum and thiolated Moringa gum samples were analysed
thermogravimetrically employing Mettler Toledo TGA, DSC 3 PLUS
(California, USA) instrument under nitrogen atmosphere from 25 °C to
400 °C at a heating rate of 10 °C/min.
2.3.4. Differential scanning calorimetry analysis
DSC thermogram of Moringa gum and thiolated Moringa gum were
recorded using differential scanning calorimeter (TA instruments,
DSC25, California, USA). Finely powdered sample of Moringa gum or
thiolated Moringa gum (5 mg) was sealed by crimping in standard
aluminium pan. The pan was heated at a rate of 10 °C/min over a
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Carbohydrate Polymers 209 (2019) 400–408
temperature range of 40–300 °C under the nitrogen purge of 50 ml/min.
2.3.5. Scanning electron microscopy analysis (SEM)
The shape and surface morphology of Moringa gum and thiolated
Moringa gum samples were investigated using scanning electron mi-
croscope (ZEISS, EVO18, China). The samples were coated with gold
and mounted on a sample holder and electron micrographs were cap-
tured at an accelerating voltage of 20 kV.
2.3.6. Energy-dispersive X-ray microanalysis (EDX)
The samples of Moringa gum and thiolated Moringa gum were ana-
lysed for presence of sulphur by SEM-EDX (ApreoLoVac, FEI). The
images of Moringa gum and thiolated Moringa gum were captured after
placing the samples on a sample stub and coating them with carbon.
2.3.7. Compression nature
The compressibility characteristics of Moringa gum and modified
Moringa gum were assessed by determining the crushing strength of
powder blends of Moringa gum or thiolated Moringa gum with magne-
sium stearate as reported earlier (Verma & Ahuja, 2017) using portable
digital hardness tester (VTHT 500, Vin Digital Tester, Mumbai, India).
In brief, blends of Moringa gum or thiolated Moringa gum containing
magnesium stearate (1%, w/w) were prepared. In first and second
blends Moringa gum or thiolated Moringa gum were blended with
magnesium stearate for, while in the case of third blend mixing was
carried out for 30 min. The powder blends so obtained were compressed
under the pressure of 75 kg/cm2 into tablets using hydraulic press (PCI
Analytics, Mumbai, India) keeping a dwelling time of 2 s for powder
blends first and third or 30 s for blend second. The crushing strength of
tablets was determined after 24 h storage in a dessicator.
2.3.8. Biocompatibility studies
Biocompatibility of thiolated Moringa gum was evaluated com-
paratively with Moringa gum by assessing their thrombogenic and
haemolytic potential. Thrombogenic potential was determined by
gravimetric method as reported earlier (Singh & Kumar, 2018). Briefly,
samples of Moringa gum or thiolated Moringa gum (0.5 g) were allowed
to swell in phosphate buffer saline (pH 7.2) at 37 OC for 24 h. The
hydrated samples were removed and placed in contact with citrated
blood (0.2 ml), to which 0.2 ml of 0.1 M calcium chloride was added.
After 45 min, 5 ml of distilled water was added to stop the clotting. The
clots so formed were fixed by adding 38% formaldehyde (5 ml), fol-
lowed by drying and weighing. The results were calculated as follows-
Ws − Wn
Thrombose (%) = × 100
Wp − Wn (1)
Where Ws, Wp and Wn are the weights of sample, positive and negative
controls respectively. The positive controls consisted of weight of the
clots without sample while negative control consisted of weight of re-
sidue without blood and samples.
Haemolytic potential was evaluated using the method reported
earlier (Singh & Kumar, 2018). For determining haemolytic potential,
samples (0.5 g) of Moringa or thiolated Moringa gum were placed in
phosphate buffer saline (PBS) for 24 h at 37 °C. The swollen samples
were incubated with 5 ml of PBS and 2 ml of citrated blood at 37 °C for
3 h. The positive and negative controls comprised of 2 ml of citrated
blood with 5 ml of distilled water and PBS respectively. After incuba-
tion, the samples were centrifuged and the supernatant fluid was ana-
lysed or haemolysis by detecting the absorbance at λmax 540 nm. The
haemolytic index was calculated as follows-
As − An
Haemolytic index (%) = × 100
Ap − An (2)
where As, Apand An is the absorbance of sample, positive and negative
controls respectively.
P. Grewal et al. Carbohydrate Polymers 209 (2019) 400–408

Table 1
Composition and evaluation of various batches of buccal tablets.
Batch code Quantity (mg/tablet) Average weight Thickness Drug content Friability (%) Ex vivo bioadhesion time (h)
(mg) (mm) (%)
MG TMG Lactose

F1 50 100 187.2 ± 0.36 1.31 ± 0.01 98 ± 1.40 3.7 12.1

F2 75 75 187.1 ± 0.38 1.10 ± 0.01 99 ± 0.42 0.79 11.8
F3 100 50 187.3 ± 0.37 1.31 ± 0.03 98 ± 0.20 20.96 12.2
F4 50 100 188.1 ± 0.40 1.13 ± 0.03 98 ± 0.80 5.83 7.9
F5 75 75 187.3 ± 0.21 1.11 ± 0.01 99 ± 0.84 0.86 8.0
F6 100 50 187.6 ± 0.39 1.17 ± 0.02 98 ± 0.72 6.36 8.3

MG: Moringa gum; TMG: Thiolated Moringa gum.

2.4. Formulation of metronidazole benzoate loaded buccal tablets of
Moringa and thiolated Moringa gum
Moringa gum and thiolated Moringa gum were tested as mu-
coadhesive polymeric carrier by formulating buccal tablets of me-
tronidazole benzoate. Table 1 shows the composition of various batches
of buccal tablets prepared using Moringa gum or thiol modified Moringa
gum. Powder blends of Moringa gum or thiolated Moringa gum, me-
tronidazole benzoate (16 mg), PEG-4000 (25 mg) and lactose were
prepared and compressed into tablets using IR hydraulic press (PCI
Analytics, Mumbai, India) under pressure of 75 kg/cm2 for 30 s.
2.5. Evaluation of buccal tablet
(250 mg) was compressed into tablet in an IR hydraulic press by ap-
plying the compression force of 35 kg/cm2 for a dwell time of 10 s.
Mucin tablets were adhered employing double-sided adhesive tape to
the lower and upper probes of the texture analyzer. Polymer tablets
were placed in contact with simulated gastric fluid (SGF, pH 1.2) for
5 min and allowed to hydrate. The hydrated tablets were placed on the
lower mucin tablet and the upper probe with adhered mucin tablet was
lowered to bring in contact with the surface of hydrated tablets and a
downward force (0.1 N) was applied for 1 min to provide intimate
contact between the polymer and mucin tablet. After 60 s, the upper
probe was withdrawn by moving up at a constant speed of 0.5 mm/s,
and the force of detachment of the mucin tablet from the surface of the
polymer tablet was inferred from the resultant force-time plots.

Metronidazole benzoate loaded Moringa gum or thiolated Moringa
gum buccal tablets were evaluated for uniformity of weight, thickness,
friability, in vitro release behaviour and ex vivo bioadhesion time.
2.5.1. Tablet friability
Six buccal tablets of each batch were weighed (Wpre) and placed in
the tablet friabilator (RemiEquipments, Mumbai, India). The drum of
the friability test machine was rotated for 100 revolutions at the rate of
25 rpm. After this, the buccal tablets were again weighed (Wpost). The
friability (%) was calculated as follows-
Wpre − Wpost
F= × 100
Wpre
2.5.2. Thickness
The thickness of buccal tablets was determined by using digital
vernier caliper (YAMAYO Digital Caliper, Pune, India). Ten buccal ta-
blets of each batch were selected randomly and thickness of individual
tablets was determined and the average was calculated.
2.5.3. Uniformity of weight
The weight uniformity of buccal tablets was determined by
weighing individually ten buccal tablets of each batch selected ran-
domly and calculating the average weight and standard deviation.
2.5.4. Drug content uniformity
Uniformity of drug contents in buccal tablets was determined by
measuring the contents of metronidazole in three tablets of each batch.
The tablets were dissolved in Mcllvaine buffer (pH 6.8), filtered and
diluted appropriately. The content of metronidazole benzoate in the
filtrate was determined spectrophotometrically by measuring the ab-
sorbance at 231 nm.
2.5.5. Mucoadhesion test
The mucoadhesive nature of Moringa gum and thiolated Moringa
gum tablets was investigated by determining the force of adhesion
between the tablets of mucin and polymer using texture profile analyzer
(Universal texture analyzer) (Jones, Woolfson, & Brown, 1997). Mucin
2.5.6. Ex-vivo bioadhesion time
The ex-vivo bioadhesion test was carried out to evaluate the mu-
coadhesive capability using chick buccal pouch as biological mem-
brane. Chick buccal pouch, procured within half an hour from slaughter
was washed and cleaned to remove loose tissues and other debris using
Mcllvaine buffer (pH 6.8) at 37 °C. The washed buccal pouch was tied to
the side of the paddle of USP type-II dissolution apparatus (TDL-08 L,
Electrolab, India). The buccal tablet was then pasted on the inner side
of mucosal membrane of pouch using light force for about 20 s. The
paddle was then rotated at 50 rpm in dissolution basket containing
250 ml of Mcllvaine buffer (pH 6.8), maintained at 37 °C (Jangra,
Ahuja, & Kumar, 2013). The bioadhesion time was then assessed by
(3)
finding the time required for the buccal tablets to detach or erode from
the surface of mucosa. The results were reported as average.
2.5.7. In vitro release studies
In vitro release rate of metronidazole benzoate from buccal tablets of
Moringa gum was carried out in USP type 2 dissolution apparatus at
37 ± 0.5 °C. Buccal tablets were attached to the bottom of the beaker
(250 ml) using cyanoacrylate glue. The beaker was filled with 250 ml of
Mcllvaine buffer (pH 6.8) and rpm was maintained at 50. At appro-
priate time intervals, aliquot of 5 ml of sample were withdrawn and
then replaced with 5 ml of Mcllvaine buffer (Yehia, El-Gazayerly, &
Basalions, 2008). The contents of metronidazole benzoate in sample
were analysed by measuring the absorbance in UV–vis spectro-
photometer at 231 nm.
3. Results and discussion
3.1. Thiolation of Moringa gum
Moringa gum is reported to consist of arabinogalactan moiety (Raja,
Bera, & Ray, 2016). In the present study, the reaction between the
hydroxyl groups of Moringa arabinogalactan and carboxyl group of
thioglycolic acid was carried out under acidic conditions to achieve
thiol derivatization of Moringa gum. Fig. 1 provides the schematic re-
presentation of the thiol modification of Moringa gum. The product
obtained was an off-white ordorless powder with an average yield of

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P. Grewal et al. Carbohydrate Polymers 209 (2019) 400–408

Fig. 1. Scheme for thiolation of Moringa polysaccharide.

83%. It was found to be soluble in 1 N NaOH but insoluble in water. The
thiol group content in thiolated Moringa gum as determined by Ellman’s
method was found to be 0.956 ± 0.024 mM of thiol groups/g of
Moringa gum.
3.2. Characterization of modified Moringa gum
Fig. 2 exhibits the FT-IR spectrum of Moringa gum and thiolated
Moringa gum. The IR spectrum of Moringa gum shows a broad peak at
3421 cm−1 which is attributed to eOH stretching. The CH stretching of

403
P. Grewal et al.
Fig. 2. FTIR spectra of Moringa gum and thiolated Moringa gum.
alkane shows peak at 2939 cm−1 and the glucoronic acid entity
shows COOH stretching at 1620 cm−1. In thiolated Moringa gum the
OH band of aliphatic alcohol appears at 3420 cm−1, while the eSH
stretch appears as a weak shoulder at 2554 cm−1. The carboxylic
groups of thiolated Moringa gum shows on stretch at 2938 cm−1 and
CeO stretch at 1253 cm−1. The stretch at 1074 cm−1 can be due to
CeO stretch of primary alcohol. In a recent study it was also reported
that thiol bands are weakly detectable in FTIR spectroscopy (Fernanda,
Borsagli, Carvalho, & Mansur, 2018).
Fig. 3(A) compares the weight loss of thiolated Moringa gum and
Moringa gum samples as a function of temperature. TGA curve of
Moringa gum presents 3 stage of degradation. The first stage of de-
gradation which begins around 50 °C and end at 225 °C is characterized
by weight loss of 20%. This weight loss may be ascribed to desorption of
water bound to the saccharide structure and dehydration due to the loss
of eOH groups on the polysaccharide backbone (Cozic, Picton, Garda,
Marlhoux, & Cerf, 2009). The second stage from 225 °C to 303 °C is
characterized by a very fast weight loss of 37%. The third stage is from
303 °C to 400 °C show a relatively small loss of 8%. The degradation at
Fig. 3. A) TGA curves (B) DSC thermograms of Moringa gum and thiolated Moringa gum.
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Carbohydrate Polymers 209 (2019) 400–408
such high temperature can be attributed to the depolymerization and
pyrolitic decomposition stages which result in the evolution of CO, CO2,
CH4 etc. (Zohuriaan & Shokrolahi, 2004). On the other hand thiolated
Moringa gum degrades with 12% weight loss during first stage from
50 °C to 133 °C, followed by loss of 38% from 133 °C to 270 °C and then
17% from 270 °C to 400 °C. At the end of 400 °C, the residual mass of
35% and 33% of Moringa gum and thiolated Moringa gum respectively
was left. The results show a faster thermal degradation of thiolated
Moringa gum at temperature less than 270 °C but above 270 °C there is
not much difference in the thermal degradation rate of Moringa gum
and thiolated Moringa gum.
The DSC thermogram (Fig. 3B) of Moringa gum showed a broad
endothermic peak followed by two exothermic peaks. The endothermic
peak appeared at 128.84 °C with an onset of 67.89 °C and enthalpy of
fusion 313.20 J/g. The two exothermic peaks appeared at 256.09 °C and
284.62 °C with enthalpy of fusion of 8.7857 J/g and 20.878 J/g, re-
spectively. Thermal curve of thiolated Moringa gum presented four
endothermic peaks at 108.90 °C, 144.16 °C, 150.42 °C and 156.87 °C
with respective heat flow values of 21.518 J/g, 2.6399 J/g, 0.4006 J/g
and 1.9382 J/g. Further thiolated Moringa gum curve shows one exo-
thermic peak at 242 °C with heat flow of 31.220 J/g.
Fig. 4 shows the scanning electron micrographs of Moringa gum (a,
b) thiolated Moringa gum (c, d). The particles of Moringa gum are
polyhedral while that the images thiolated Moringa gum shows the
presence of polyhedral plate shape flakes. It can be seen that surface of
Moringa gum is rough and granular and that of thiolated Moringa gum is
smooth and porous. The smooth surface of thiolated Moringa gum is
expected to provide the greater area of contact with the mucus layer as
compared to the Moringa gum.
The EDX spectrum of Moringa gum (Fig. 5a) and thiolated Moringa
gum (Fig. 5b) confirms the successful thiolation of Moringa gum. The
new X-ray intensity was observed at around 2.5 keV in thiolated Mor-
inga gum due to the presence of sulphur in thiolated Moringa gum. The
EDS layered images of Moringa and thiolated Moringa gum are pre-
sented as Supplementary material (Fig S1 and S2).
3.3. Compression behaviour
Thiol modified Moringa gum was further characterized for its com-
pression behaviour studies. The results of compressibility character-
ization study revealed that the tablets of Moringa gum prepared using
blends first, second, and third had the crushing strength of 73.3 N,
81.3 N and 19.1 N respectively. On the other hand tablets of thiolated
P. Grewal et al.
Fig. 4. Scanning electron micrographs showing (a) surface (b) shape of Moringa gum and (c) surface (d) shape of thiolated Moringa gum.
Moringa gum showed crushing strength of 120.3 N, 138.4 N and
114.8 N. It can be observed from the results that increasing the dwell
time increase the bending strength while the increase in blending time
reduces bonding which indicate that both Moringa gum and thiolated
Moringa gum show plastic behaviour. However, thiol modification re-
sults in increase in bond strength of particles.
3.4. Biocompatibility studies
Moringa gum is biocompatible and has been used in traditional
medicines in the management of diarrhoea and for its diuretic effects.
During earlier studies, radiation, cross linked Moringa gum-polyacrylic
acid hydrogels were evaluated for biocompatibility by blood compat-
ibility tests (Singh & Kumar, 2018). The comparative blood compat-
ibility tests conducted on Moringa gum and thiolated Moringa gum re-
vealed that the amount of clot formed in case of Moringa gum
(0.1258 g/2 ml citrated blood) and thiolated Moringa gum (0.1939 g/
2 ml of citrated blood) was less that the clot formed in case of positive
control (0.2471 g/2 ml of citrated blood). Thus, the percentage
thrombose of Moringa gum and thiolated Moringa gum was found to be
50.91% and 78.47% respectively. As the thrombus formed was lower in
case of Moringa gum and thiolated Moringa gum as compared to control,
Moringa gum and thiolated Moringa gum can be categorized as non-
thrombogenic. The results of haemolytic study revealed that the hae-
molytic index of Moringa gum and thiolated Moringa gum was 3.86%
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Carbohydrate Polymers 209 (2019) 400–408
and 1.20% respectively. Since the haemolytic index value is less than
5%, they can be regarded as safe and suitable for drug delivery appli-
cations.
3.5. Mucoadhesion studies
Thiolation of polymers imparts the mucoadhesive characteristics on
the polymer. So, thiolated Moringa gum was tested for mucoadhesive
properties using texture analyzer. Fig. 6 displays the force time curve of
Moringa gum and thiolated Moringa gum compacts. The force of de-
tachment of polymer compacts of Moringa gum and thiolated Moringa
gum from mucin tablets was found to be 804 mN and 1070 mN re-
spectively. The greater force of detachment of thiolated Moringa gum
indicates its mucoadhesive nature.
3.6. Formulation and characterization of buccal tablets
Considering the mucoadhesive potential of thiolated Moringa gum,
it was decided to formulate mucoadhesive buccal tablets.
Metronidazole benzoate a commonly used anti gingivitis drug
(Mitchell, 1984) was used as the model drug. Buccal tablets were for-
mulated using lactose as the diluent, PEG-4000 as the plasticizer. The
results of the characterization of buccal tablets indicate that the average
weight, drug content and thickness were within the range. However, as
the buccal tablets prepared using lower and higher levels of Moringa
P. Grewal et al. Carbohydrate Polymers 209 (2019) 400–408

Fig. 5. EDX-ray analysis of (a) Moringa gum (b) thiolated Moringa gum.

Fig. 6. Texture profile analysis of Moringa gum and thiolated Moringa gum tablet.

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P. Grewal et al. Carbohydrate Polymers 209 (2019) 400–408

Fig. 7. In vitro release profile of metronidazole benzoate from thiolated Moringa gum buccal tablet and Moringa gum buccal tablet.

gum or thiolated Moringa gum showed higher friability while the buccal
tablet containing equal proportion of polymer and lactose passed the
friability test. The ex vivo bioadhesion time study reveal about 1.5-fold
higher bioadhesion time of buccal tablets of modified gum compared to
that of native gum.
3.7. In vitro release of buccal tablets
possibility of use of metronidazole loaded thiolated Moringa gum tablets
in such case, the in vitro release study was also conducted using
Mcllvaine buffer (pH 1.2). It was observed that at acidic pH, the release
of drug from gum followed Higuchi’s square root kinetics with diffusion
through the matrix as primary mechanism of release, except batch F4
which showed anomalous transport. The results of which are presented
as Supplementary material (Figs. S3–S15).

Fig. 7 compares the results of in vitro release study carried out in

Mcllvaine buffer (pH 6.8). It can be observed that almost the entire drug 4. Conclusions
was released within 6 h from the buccal tablets formulated using Mor-
inga gum while buccal tablets of thiolated Moringa gum were able to Moringa gum was modified by esterification of its hydroxyl groups
sustain the release till 24 h. Slower release can be attributed to the with thioglycolic acid to synthesize thiolated Moringa gum. The mod-
disulfide bond formation between glycoproteins of mucus layer and ified Moringa gum was characterized by FTIR, TGA, DSC, XRD and SEM
polymer which are much stronger (Bernkop-Schnurch et al., 2001). studies. The thiolation of Moringa gum improved its mucoadhesion
Further the release data was subjected to model fitting, the results ability as evaluated by tensile studies using texture analyzer. The
(Table 2) of which unveil that the release of metronidazole benzoate thiolated Moringa gum was further explored for pharmaceutical appli-
from Moringa gum buccal tablet followed Higuchi’s square root kinetics cations by formulating metronidazole benzoate loaded tablets. The
with the anomalous release mechanism i.e. combination of diffusion thiolated Moringa gum containing tablets showed higher bioadhesion
and erosion. On the other hand release from thiolated Moringa gum time as compared to the unmodified gum. The in vitro release studies
buccal tablet followed first order kinetics with the primary mechanism indicate that thiolation of Moringa gum imparts sustained release
of release being diffusion through the matrix. Metronidazole is com- characterization. Thus, it can be inferred that thiolation of Moringa gum
monly used in the treatment of intestinal infections. To explore the is a feasible approach to improve its mucoadhesion and release re-
tardant property. However, further in vivo studies in animals are needed
Table 2 to exploit its potential for use in buccal drug delivery in human beings.
Modeling and release kinetics of metronidazole benzoate tablet formulations.
Batch R2 n
Code
Acknowledgements
Zero-order First-order Higuchi Korsmeyer-Peppas
square- root The authors express gratitude to Department of Science &
Technology, Govt. of India for providing financial assistance to Jyoti
F1 0.717 0.943 0.901 0.976 0.256
F2 0.628 0.940 0.901 0.959 0.334
Mundlia under DST-PURSE programme. The authors also express gra-
F3 0.749 0.886 0.800 0.967 0.191 titude to the Department of Textile Technology, IIT, Delhi and Materials
F4 0.807 0.948 0.939 0.963 0.777 Science Laboratory, Department of Physics, GJUS&T, Hisar for pro-
F5 0.964 0.923 0.949 0.983 0.691 viding scanning electron microscopy and X-ray diffraction facility, re-
F6 0.965 0.960 0.968 0.994 0.799
spectively.

407
P. Grewal et al.
Appendix A. Supplementary data
Supplementary material related to this article can be found, in the
online version, at doi:https://doi.org/10.1016/j.carbpol.2018.12.100.
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