Sustainable Chemistry and Pharmacy 21 (2021) 100414

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Sustainable Chemistry and Pharmacy

journal homepage: http://www.elsevier.com/locate/scp

Characterization and sustainable anti-ulcer activity of hydrogels based on

acid-diol reactions
Chukwuma O. Agubata a, *, Nkemjika I. Ezeanya a, Ifeanyi T. Nzekwe b, Samuel W. Uzondu c,
Chukwuemeka C. Mbah a
a
Department of Pharmaceutical Technology and Industrial Pharmacy, University of Nigeria Nsukka, Enugu State, Nigeria
b
Department of Pharmaceutics and Pharmaceutical Technology, Nnamdi Azikiwe University, Awka, Anambra State, Nigeria
c
Department of Pharmaceutics, University of Nigeria Nsukka, Enugu State, Nigeria

A R T I C L E I N F O A B S T R A C T

Keywords: Hydrogels have been applied in biomedical field to facilitate drug delivery and wound closures. However, there is
Sustainable a need for a more sustainable chemical approach. The objective of this study was to prepare oral hydrogels based
Hydrogels on dynamic interactions between boric acid and chitosan or bioactives-rich extract of Mangifera indica leaf, and
Gastro-protective
assess their physicochemical and sustainable gastro-protective qualities. The leaves of Mangifera indica were
collected, dried, powdered and extracted with methanol using soxhlet extractor. The extract was characterized
by Gas chromatography-mass spectroscopy (GCMS). Hydrogels were prepared using varied combinations of diol-
based chitosan and extract (1:1, 3:1, 1:3, 1:0, 0:1) in mixtures with boric acid. The pH, viscosity, sedimentation
volume, redispersibility, Fourier Transform-InfraRed Spectroscopy (FTIR), release of phytoconstituents and anti-
ulcer activities of the hydrogels were investigated. The GCMS result revealed the presence of diols, quinoline
derivatives and other compounds. The formulations showed pH range of 1.5–6.2 and viscosities were mostly
greater than 400 mPa/s. The values of redispersibility number and sedimentation volumes of the formulations
expressed the relative physical stabilities of the batches and compatibility was assessed from FTIR spectra. The
hydrogels were found to have gastro-protective effects and effectively released entrapped bioactives in extract.
Test hydrogels showed significantly high anti-ulcer activities and provision of barrier functions against toxic
substances. Furthermore, preparations containing the extract showed less adverse changes in colour of gastric
mucosa. Oral hydrogels based on boric acid, chitosan and methanol extract of M. indica leaves show promising
properties in ulcer treatment.

1. Introduction properties. This characteristics of materials can also enable them to

Stimuli-responsive hydrogels have the abilities to respond to external
triggers or conditions such as pH, temperature, specific chemical com­
pounds or moieties, light etc. This unique property has been applied in
drug delivery, biomedicine (Nakahata et al., 2014), tissue engineering
and other fields. Self-healing hydrogels have also been developed as
novel biomimetic explorations of the behaviors of some materials with
abilities to regenerate or repair and heal after physical damage or tear
(Pettignano et al., 2017). Based on dynamic covalent chemistry, some
materials form hydrogels which can self-repair after injury. Boric acid
and related boronic acid have been shown to interact reversibly with
diols and the dynamic covalent bonding with these polyhydroxyl groups
creates a reversible equilibrium state that imparts self-reparation
effectively provide barrier protection for sensitive surfaces. These
reparative and barrier properties of these materials can be extended to
natural macromolecules at biointerfaces to initiate biological healing
processes or protect susceptible ones. The use of natural sustainable
medical interventions is desirable for improved efficacy, reduced
toxicity, favorable metabolic pathway, availability, patients’ compli­
ance and minimum cost.
Numerous polyhydroxy compounds of low molecular weights can
form valuable complexes with boric acid in aqueous solutions. The
complexation of boric acid with different hydroxyl compounds show
that polyols form stronger complexes with boric acid (Bai et al., 2019).
Biologically-derived polyols include chitosan. Chitosan is known to be
soluble in dilute acids with pH lower than 6.3 forming non-Newtonian

* Corresponding author. Department of Pharmaceutical Technology and Industrial Pharmacy, University of Nigeria, Nsukka, 410001, Enugu State, Nigeria.
E-mail address: chukwuma.agubata@unn.edu.ng (C.O. Agubata).

https://doi.org/10.1016/j.scp.2021.100414
Received 23 October 2020; Received in revised form 24 January 2021; Accepted 17 February 2021
Available online 26 February 2021
2352-5541/© 2021 Elsevier B.V. All rights reserved.
C.O. Agubata et al.
shear-thinning fluid (Mucha 1997). Under condition of low pH, the free
amino groups in the chitosan become protonated causing electrostatic
repulsions between adjacent polymer chains, thereby facilitating sol­
vation (Szymanska and Winnicka 2015).
There is a research need to develop barriers around gastric mucosa
that can maintain its integrity when exposed to ulcer-inducing chemical
agents and chitosan-based formulations have shown some promise.
Chitosan is the deacetylated form of chitin, and it is usually soluble in
acidic medium (Younes and Rinaudo 2015). Chitin is a polysaccharide
that occur naturally as ordered macrofibrils and form structural com­
ponents in exoskeleton of crabs, shrimps and fungal cell wall (Eli­
eh-Ali-Komi and Hamblin 2016). Chitosan is a polysaccharide obtained
from chitin and it is non-toxic, biocompatible, stable, bioadhesive and
biodegradable (Parhi 2020). Reports show that entrapment of viable
biological cells into chitosan-based hydrogels does not show any sig­
nificant undesirable effects, which implies these materials show
biocompatibility (Chenite et al., 2000).
Plant-derived extracts are also rich in polyhydroxyl groups that may
be valuable in healing and barrier functions. The plant, Mangifera indica
has been reported to contain different phytocompounds of which some
entities are polyhydric (Shah et al., 2010). Also, Leaf extract of Mangifera
indica L. has been reported to show acute anti-inflammatory activities
based on its abilities to increase the miRNA expression of peroxisome
proliferator-activated receptor alpha (PPAR-α) (Toledo et al., 2019). The
effect of leaf extract on PPAR-α and the influence of PPAR-α on fatty acid
catabolism including interactions with ligands such as arachidonic acid
explains these earlier findings.
Ulcers can occur on the stomach or intestinal mucosa. Different
models have been applied to study anti-ulcer activities of some agents
and these include ethanol-induced model (Cho and Ogle 1979; Oates
and Hakkinen 1988). In this present study, ethanol is being used to cause
gastric injury and also increase the gastric pH to levels that would
require proposed preventive and restorative qualities of formulated
borate-diol hydrogels. Disruption of the gastric mucosal barrier would
permit back diffusion of gastric acid into the submucosal sub-region
(German et al., 2008) and initiate a series of events that causes
inflammation, exposure to harsh substances and eventual injury.
Moreover, ethanol has a direct injurious effect on the gastric mucosa and
gastro-protective materials are desired for preventive purposes. An
important function of biological hydrogels (biogels) is to serve as
effective barrier against foreign or undesirable matters. Other entities
can be introduced to interact with the biogel matrix and this can
modulate or tune the functionality of these materials (Schiller and Lai,
2020).
The sustainable approach of formulation being employed in this
study mostly involve the use of natural materials that do not have
negative impact on the environment and are readily available.
The aim of research is to prepare hydrogels based on boric acid-diol
interactions and evaluate their gastro-protective capabilities.
2. Materials and methods
2.1. Materials
Boric acid (Guangdong Guanghua Chemicals, China), Chitosan
(Sigma Aldrich, USA), Hydrochloric acid (Sigma Aldrich, USA), Meth­
anol (Guangdong Guanghua Chemicals, China), Omeprazole tablet
(Shalina laboratories PVT. LTD, India), Powdered Mango leaves pre­
pared in Department of Pharmaceutical Technology and Industrial
Pharmacy Laboratory, University of Nigeria Nsukka.
2.2. Methods
2.2.1. Plant material collection
The leaves of Mangifera indica were harvested from trees on the
grounds of University of Nigeria, Nsukka. Enugu state. The leaves were
2
Sustainable Chemistry and Pharmacy 21 (2021) 100414
dried in an oven at a temperature of 38 ± 1 ◦ C and thereafter, reduced to
powder which was stored in an airtight container.
2.2.2. Extraction from Mangifera indica leaves
A 100 g quantity of the powdered leaves was transferred into a clean
soxhlet extractor. Using a 500 ml measuring cylinder, 500 ml of meth­
anol was measured out and transferred into the distillation flask. The
soxhlet extractor was set up and after extraction of the leaves, the
solvent-extract mixture was transferred to a beaker and was allowed to
cool. The solvent was allowed to evaporate from the mixture leaving
behind a sticky solid extract. This procedure was repeated four times.
2.2.3. Physicochemical characterization of Mangifera indica leaf extract
2.2.3.1. Organoleptic properties and percentage yield. Organoleptic
properties of the leaves extract were evaluated and these include colour,
odour and feel. The percentage yield was calculated using the equation;
Weight of extract
% ​ yield = × 100 1
Weight of powdered leaves
2.2.3.2. Gas Chromatography-mass spectroscopy (GC-MS). Gas
chromatography-mass spectroscopy was performed on a sample of
methanol extract of Mangifera indica leaves to elucidate its constituents.
A GC-MS equipment (Agilent, USA) was used for this study. The reten­
tion times of GC and data obtained from MS were simultaneously
correlated and compared with spectra library using computer assisted
techniques to identify some constituents or by-products. Peak height and
area were used to ascertain the relative quantities of the components.
2.2.4. Preparation of sustainable hydrogels
A 2 g amount of chitosan was dissolved in 50 ml 0.1N HCl and 2 g of
Extract was incorporated into the chitosan solution in a beaker by stir­
ring for 15 min until a homogenous mixture was obtained. A 2 ml vol­
ume of 25% boric acid was added into the mixture dropwise while
simultaneously stirring. The beaker containing the resulting hydrogel
was thereafter placed on a mechanical shaker and shaken at room
temperature for 1 h. The product contained chitosan and extract at 1:1
ratio. Other batches were prepared using the above procedure at 3:1,
1:3, 1:0 and 0:1 chitosan/extract ratios by weight.
2.2.5. Physicochemical characterization of hydrogels
2.2.5.1. pH and viscosity measurement. The pH of the different hydrogel
preparations was ascertained using a well-calibrated pH meter (HANNA
Instruments, Padova, Italy). The electrode part was immersed into 30 ml
quantities of each batch and readings obtained.
The viscosity of 30 ml of extract was measured using a DV-1 Broo­
kefield Digital Viscometer using Spindle number 2, at a speed of 60 rpm.
Viscosity is an expression of the resistance to flow of the fluids. The
viscosity measured represents internal friction and measures resistance
to deformation, change in shape or movement of ‘intimate’ portions
relative to one another. The viscosity values from the instrument are
presented as mPa/s.
Different concentrations of the components create varying layers of
dynamic resistance, therefore rotation using the instrument produces
viscous drag unique to each batch.
2.2.5.2. Sedimentation volume. Twenty millilitres (20 ml) of each
hydrogel was transferred into a 50 ml measuring cylinder and was left
undisturbed for 5 days at room temperature. The volume of sediment
was taken every 24 h for five days. The sedimentation volume was
calculated using Equation (2).
Vt
F= 2
Vo
C.O. Agubata et al.
where Vt is the volume of the sediment at time t and Vo is the entire
volume of preparation.
2.2.5.3. Redispersibility test. Redispersibility was assessed immediately
after the last Sedimentation volume was recorded by firmly sealing the
mouth of the cylinder and rotating the cylinder through 180◦ . The
number of times the cylinder was rotated to achieve complete redis­
persion of the sediment of each test sample was recorded as redis­
persibility number.
2.2.5.4. Fourier Transform infrared spectroscopy (FTIR). FTIR was used
to investigate any chemical or physical interaction between components
of the extract and chitosan in the formulations. Small quantities of
extract, chitosan and formulated hydrogel were scanned over a wave­
length range of 4000-500cm− 1 at a resolution of 4cm− 1 in a FT-IR in­
strument (Agilent Technologies, USA). The scan was operated in the
transmittance mode. The spectra generated represent functional groups
present in the samples and these were interpreted based on uniqueness
of different wavelengths to specific chemical groups.
2.2.5.5. Phytoconstituent release studies. A 10 mg% extract solution in
0.1N HCl was scanned through different wavelengths with a UV/VIS
spectrophotometer (Jenway, UK) to obtain a λmax at 220 nm and a
standard calibration was done using series of concentrations. Thereafter,
One ml of the test extract sample was enclosed in a dialysis membrane
(MWCO 5000–8000) and suspended in a beaker containing 500 ml of
0.1N Hydrochloric acid as medium. The beaker was mounted on a
magnetic stirrer set-up at 37 ◦ C and 50 rpm. A 5 ml quantity of test
solution was withdrawn and 5ml of fresh medium immediately re-
injected at 30 min interval for 3 h. The test solutions were assayed at
220 nm using UV/VIS spectrophotometer to obtain absorbances for
phytoconstituent group with λmax at 220 nm. The study was repeated
twice. This study was also done for formulations 3C/1E, 1C/3E and 2C/
2E.
2.2.5.6. In-vivo animal studies. Ethanol-induced ulcer model was used
to study the gastro-protective/anti-ulcer activity of the oral hydrogels.
The rats were fasted (for 24 h) prior to onset of the experiment but with
free access to water. The rats were divided into 6 groups (n = 5) and the
following was administered orally; Group 1 received Omeprazole (5 mg/
ml) as positive control, Group 2 received distilled water (0.2ml) as
negative control, Group 3 was administered with 150 mg/kg hydrogel
containing chitosan with no extract (2C/0E, 3.85%), Group 4 received
150 mg/kg of hydrogels with 2:2 mixture of chitosan and extract (2C/
2E), Group 5 received 150 mg/kg of batch containing 1:3 chitosan and
extract mixture (1C/3E), while Group 6 was administered with 150 mg/
kg hydrogels containing extract with no chitosan (0C/4E, 7.7%).
An hour after the administration of test substance, ulcer was induced
by oral administration of 1ml of absolute ethanol to all the animals. The
animals were sacrificed an hour later by cervical dislocation, the stom­
ach removed and opened along the greater curve. The stomach was
rinsed in distilled water, pinned to a flat white surface and observed with
a hand lens ( × 10). Mean ulcer score for each group was calculated and
expressed as the ulcer index (UI) which considers number of animals.
The Ulcer protection (%) values of the treated groups were calculated
using Equation 3;
Ulcer Protection ​ (%) = Ulc − Ult/Ulc × 100 3 ature applied (Montembault et al., 2004).
Where Ulc is the ulcer index of the control group and Ult is the ulcer
index of the treated group.
Method of determining Ulcer score was adapted from Adami et al.
(1964) and is presented in Table 1.
Furthermore, visual examination was used to assess the colour of the
gastric mucosa.
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Sustainable Chemistry and Pharmacy 21 (2021) 100414
Table 1
Criteria for ulcer scores.
Ulcer Score Description
0 No ulcer
1 Haemorrhagic and slight ulcer, length < 2mm
2 One haemorrhagic and slight ulcer, length <5mm
3 More than one grade 2 ulcers
4 One ulcer of length <5mm and diameter <2mm
5 One to three ulcers of Grade 4
6 Four to six ulcers of Grade 4
7 More than six ulcers of Grade 4
8 Complete lesion of the mucosa with haemorrhage
3. Results and discussion
3.1. Physical properties and percentage yield of extract
The extract was greenish black in colour, it had a characteristic
odour and was sticky to touch. The Viscosity was found to be 313 mPa/s.
The percentage yield was calculated to be 32.68%
3.2. Gas Chromatography-mass spectroscopy
The GC-MS data and spectra (Fig. 1) obtained from this study and the
correlating library search revealed the presence of bioactive chemical
groups such as Quinoline derivatives, cyclohexanedicarboxylic acid,
pyridines and pyrimidines, anthracene, naphthalene carboxylic acids,
naphthoquinones, phenanthryl derivatives, propanedioic acids etc. The
retention times and relative amounts of constituents are presented in
Table 2. The presence of diols suggest that the extract can partially
interact with boric acid to establish a unique reversible bonding that
may be vital in biomimetic or biological activity.
3.3. Characterization of hydrogel
3.3.1. pH and viscosity
The pH and viscosity of the different formulations are presented in
Table 3. Most of the formulations have pH values within the range of
5.0–6.2 while the formulation prepared with extract and boric acid
showed a low pH value of 1.5. The observed pH range show that the
preparations will be easily tolerated in the gastrointestinal tract when
administered.
The viscosity values were in the range of 252–490 mPa/s. The
viscous nature of these preparations would reduce the sedimentation
rate of the particulate plant matters present. The gelling of the initial
liquid precursors increased the viscosity of the product. Assessment of
viscosity was more difficult for the extract-boric acid batch because of
high particulate matter mobility. The study also showed clearly that
increasing concentration of chitosan resulted in increased viscosity
whereas increase in the plant extract reduced the viscosity in extract-
chitosan systems. Chitosan is a viscosity-enhancing agent in acidic
environment which is because of its high molecular weight and linear
unbranched molecular structure. It has been reported and confirmed
that viscosity of chitosan solutions increases with increasing concen­
trations of chitosan (Rowe et al., 2019). Earlier investigations have also
shown that gelation of chitosan is influenced by some factors such as
apparent charge density, degree of acetylation of chitosan and temper­
The jelly and viscous nature of these preparations would also facil­
itate adsorption on internal bio-surfaces such as walls of gastro-
intestinal tract and this would influence efficacy of the investigative
anti-ulcer remedy.
3.3.2. Sedimentation volume
The result of sedimentation test is presented in Table 4. The formu­
lations prepared with only chitosan and boric acid (cross-linker) showed
C.O. Agubata et al. Sustainable Chemistry and Pharmacy 21 (2021) 100414

Fig. 1. Gas Chromatograph of methanol extract of Mangifera indica leaves.

Table 2 Table 3
Data from GCMS. pH and Viscosity of hydrogel formulations.
Peak Retention Time (Min) % relative amount of Total Concentration Viscosity (mPa/s) pH

1 33.11 4.21 4g extract/boric acid ———a 1.50

2 50.67 1.83 4g chitosan/boric acid 490.0 6.20
3 53.76 1.23 3g chitosan+ 1g extract/boric acid 489.5 6.10
4 54.98 1.69 1g chitosan+ 3g extract/boric acid 252.0 5.00
5 59.35 0.83 2g chitosan+ 2g extract/boric acid 425.0 5.70
6 63.97 2.39 2g chitosan/boric acid 421.0 6.20
7 67.16 6.16 a
8 68.84 2.01 Equipment could not measure with accuracy due to dynamic nature of sus­
9 69.09 1.81 pended particles.
10 71.94 27.25
11 72.95 4.03
viscosity is inversely related to rate of sedimentation. Therefore high
12 74.63 18.43
13 76.71 4.30
viscosity slows down the sedimentation of particles thereby improving
14 77.27 18.16 sedimentation volume. The sedimentation volume is defined as the ratio
15 79.04 5.67 of the equilibrium volume of suspension zone occupied by sediments Vu,
to the entire volume of the suspension, Vo. The value ranges between
0 and 1 and increases as the volume of the suspension that is occupied by
the highest value of sedimentation volume (F) as 0.8, although the
the sediment increases. The value of F provides a qualitative knowledge
amount of particles present was small compared to extract-based ones.
about the physical stability of the suspension. The greater the value of F,
The lowest F value of 0.1 was seen in the formulations containing only
the more stable the suspension and the better its suspendability.
plant extract and boric acid (without chitosan). The sedimentation
volume of the preparations increased with increasing concentration of
3.3.3. Redispersibility number
chitosan whereas the values decreased with increasing concentrations of
The highest value of redispersibility number of 13 was seen in 1:1
plant extract. This influence of viscosity on sedimentation volume can
chitosan/extract formulation while the lowest redispersibility value of 2
also be explained using the stoke law/equation which shows that

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C.O. Agubata et al. Sustainable Chemistry and Pharmacy 21 (2021) 100414

Table 4 was seen in the extract formulation (without chitosan). The redis­
Sedimentation volume of hydrogel formulations. persibility values for all the formulations are expressed in Table 4.
Content Sedimentation Sedimentation Redispersibility Although the extract-based formulation without chitosan showed low
Volume (F) Volume (%) number redispersibility number, this is basically because of its relatively low
4g extract/boric 0.100 10.0 2 resistance to flow whereas the high chitosan–containing formulations
acid with high viscosity showed high redispersibility numbers. A clearer ef­
4g chitosan/boric 0.800 80.0 7 fect of the extract was observed when comparing formulations prepared
acid using 2g chitosan (without extract) and the 2g chitosan/2g extract
3g chitosan + 1g 0.600 60.0 12
extract/boric acid
formulation where the former showed lower values with the later
1g chitosan+ 3g 0.350 35.0 7 showing around 100% more redispersibility number.
extract/boric acid It is desirable that suspensions are easily redispersible (that is low
2g chitosan + 2g 0.425 42.5 13 redispersibility number) so as to ensure uniformity of administered drug
extract/boric acid
doses after shaking (Bhurat et al., 2012). This implies that the number of
2g chitosan/boric 0.350 35.0 6
acid inversions or cycles required to redisperse the suspension after settling
should be low. The endpoint of this test was taken when the inside of the

Fig. 2. FT-IR spectra of extract (a), chitosan (b), hydrogel with extract (c) and hydrogel without extract (d).

5
C.O. Agubata et al.
base of the cylinder was clear of sediment. However, the higher viscosity
of some preparations delayed the redispersion although no caking was
observed in these preparations and they were slower in forming
sediments.
3.3.4. FTIR spectra
Fourier Transform infrared spectroscopy is useful in providing in­
formation about the presence or absence of specific functional groups
and provides molecular fingerprints for identification.
The FTIR spectrum of the extract (Fig. 2a) showed broad band at
3246.5 cm-1 which corresponds to –OH bonds present in the extract.
Also C–H bond stretching was observed at 2833.4 cm-1. The spectrum of
chitosan (Fig. 2b) showed peaks at 3382 cm− 1, 3291.2 cm− 1, 2870
cm− 1and 1640 cm− 1 representing peaks or bands due to –NH2, –OH,
C–H and C– – O respectively. The FTIR spectra of the hydrogels with and
without extract (Fig. 2c and d) contain bands corresponding to chemical
groups found in the constituents. These spectra showed only the broad
band of –OH at 3308 cm− 1/3285 cm− 1 due to chitosan, –OH bearing
bioactives from extract and boric acid. The spectra also showed peaks at
1638 cm− 1 signifying the presence of –C–– O bond. It was also clear that
the peaks of amines were missing and this may imply that there was
cyclization during complexation and maybe some of the primary amines
of chitosan have been transformed to the secondary forms which do not
show any peak in FTIR spectra. This could involve a condensation re­
action. These activities could be an early indication of a form of
reversible dynamic covalent interactions.
3.3.5. Drug release studies
A UV/VIS spectrophotometer was used to scan a dilute aliquot of the
Fig. 3. Standard calibration curve for Mangifera indica leaf extract.
6
Sustainable Chemistry and Pharmacy 21 (2021) 100414
plant extract at different wavelengths to obtain a λmax of 220 nm at
which point maximum light absorption was observed which corresponds
to the most significantly abundant and sensitive chromophore in the
extract. The standard calibration curve of the extract marker is pre­
sented in Fig. 3 and used to establish the release profile from the
formulations.
The release profile of the extract component was then monitored as
increasing absorbances at 220 nm and is presented in Fig. 4. At the onset
of release, batches containing high amounts of extract (1C/3E, 0C/4E)
showed higher levels of release and diffusion. However as the study
progressed, preparation with the lowest extract content (3C/1E) started
showing improved delivery and eventually showed release higher than
the other batches. The accumulation of extract materials at the surface of
dialysis membrane may have ultimately reduced the rate and extent of
passage of phytoconstituents across the membrane for preparations with
higher concentration of extract, thereby favouring the eventual higher
passage of plant material in the receiver medium for a low dose prepa­
ration. After 30 min, about 31% and 29% of extract marker was released
and permeated across the dialysis membrane from 1C/3E and 0C/4E
respectively, whereas 22% was detected across the membrane from 3C/
1E and 2C/2E hydrogels. Subsequently, 3C/1E batches released 100% of
the phyto-marker across the membrane after 3h while 1C/3E, 2C/2E and
0C/4E hydrogels only achieved 79%, 71% and 62% release respectively,
after 3h of investigation. The release rate of the phytoconstituent marker
is facilitated by the use of receiver medium with pH around 1 (0.1N
HCl).
3.3.6. In-vivo animal studies
The ethanol-induced ulcer model was used to test for the gastro-
C.O. Agubata et al. Sustainable Chemistry and Pharmacy 21 (2021) 100414

Fig. 5. Gastro-protection profile of hydrogels based on chitosan and Mangifera

indica extract.

compared to negative control after it was observed that ulcer scores/

indices were not significantly different. This may be that omeprazole
mechanism of action of inhibition of proton pump and acid secretion has
more value for treatment of ulcer rather than preventing ulceration by a
chemical agent.
In this ethanol-induced ulcer model, the ethanol may have pene­
trated the gastric mucosa because it can solubilize the protective mucous
Fig. 4. Bioactive release and diffusion profile of Mangifera indica. and expose the mucosa to the proteolytic and hydrolytic actions of hy­
drochloric acid and pepsin (Oates and Hakkinen, 1988) which would
protective effect of the different liquid hydrogel formulations. Distilled have resulted in damaged membrane (Sener et al., 2004). The ethanol
water (DW) was used as the negative control while Omeprazole (as could have also stimulated gastric acid secretion and increase oxidative
dispersed granules from 20 mg omeprazole capsule) served as the pos­ stress which all have deleterious effects on the gastric mucosa.
itive control. The comparisons of ulcer index and percentage protection Chitosan-based hydrogels may have reduced the ulcerative effect of the
were made against the negative control. ethanol by forming a protective mucous-like layer which is not
Effect of extract and chitosan-based hydrogels on absolute ethanol- completely dissolved in ethanol. The basic nature of chitosan also allows
induced ulcer is presented in Table 5, Figs. 5 and 6. Any test sample it to bind to bile acid thereby reducing the interaction of the later with
having an ulcer index greater than that of the negative control is said to the injured gastro-surface. Moreover, Chitosan is mucoadhesive and
have no ability to prevent ethanol-induced ulcer. Hence, the positive induces mucin aggregation through electrostatic forces, hydrogen
control (drug dispersion) showed no gastro-protective effect, probably bonding and other mechanisms (Sogias et al., 2008).
because it provided ‘no preventive barrier effect’ although the drug is The borate-chitosan complex may have also played a crucial role
known to heal ulcer. The 1C/3E formulation showed 52.6% ulcer pro­ using its self-healing property to prevent exposure of the gastric mucosa
tection, 2C/2E and 2C/0E formulations showed 49.5% ulcer protection, to the whole bulk of the ethanol. As the ethanol causes damage during
whereas 17.9% ulcer protection was observed with 0C/4E formulation. passage through the hydrogel layer, the hydrogel may simultaneously
The anti-ulcer effect provided by the different formulations seem to be repair thereby preventing further disruption and protecting the surface
due to the viscosity-dependent barrier nature of the hydrogels as the of the mucosa. However, this hydrogel behavior which is based on
least viscous of the formulations (4E/0C) showed the lowest percentage reversible covalent interaction and hydrogen bonding would face in­
gastro-protection in the study. The formulations containing chitosan terferences from physiological conditions and other substances present.
showed a significant ulcer-preventive effect (p < 0.05) compared to the One of the basic reasons why the formulations based on just extract
negative control. This may imply that chitosan has the ability to provide and boric acid only showed a low percentage ulcer protection is that
a ‘screen coat’ on the gastric wall and/or the hydrogels formed with these formulations were much more acidic with pH of 1.2 whereas those
boric acid have self-reparative effect that also catalyzes natural healing containing chitosan were observed to have a pH values of 5.0–6.2.
process. It had earlier been reported that chitosan can bind with red Moreover, these extract-based formulations (without chitosan) lack the
blood cells thereby promoting rapid clotting (Kozen et al., 2008). The ‘slimy’ cover observed in those with chitosan. However, preparations
interaction of the fluid chitosan-based hydrogels with biological mac­ containing the extract showed less adverse mucosal colour changes.
romolecules may have reduced the immediate effect of ulcerative agent
(ethanol). 4. Conclusion
Omeprazole (OPZ) showed similar percent gastro-protection
Hydrogels based on boric acid, chitosan and/or methanol extract of
Mangifera indica leaves were developed and show effective gastro-
Table 5 protection against chemical agents resulting in minimal adverse ef­
Effect of hydrogels containing chitosan and extract on ethanol-induced ulcer. fects on gastric mucosa.
Treatment Dose (mg/kg) Ulcer index % Protection
Credit author statement
Distilled water (DW) 5 ml/kg 0.95 ± 0.33 –
Omeprazole (OPZ) 20 0.98 ± 0.29 − 3.2
0E/2C 0.4 0.48 ± 0.11 49.5 Agubata CO: Conceptualization, methodology, supervision,
2E/2C 0.4 + 0.4 0.48 ± 0.18 49.5 Writing-Review and editing.
3E/1C 0.6 + 0.2 0.45 ± 0.25 52.6 Ezeanya NI: Investigation, Writing-original draft preparation
4E/0C 0.8 0.78 ± 0.43 17.9
Nzekwe IT: Validation, Writing- review and editing.

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C.O. Agubata et al. Sustainable Chemistry and Pharmacy 21 (2021) 100414

Fig. 6. Gross image of stomach harvested from animals in group 1 (A), group 2 (B), group 3 (C), group 4 (D) and group 5 (E).

Uzondu SW: Formal analysis
Mbah CC: Resources
Declaration of competing interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influence
the work reported in this paper.
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