Formulation of a Diuretic Oral Suspension of Basella alba

L. (Basellaceae)
Noreen Mae E. Chavez, Bianca Mae G. Limjoco, Melchi Esrom L. Opelario, Edmin Christian R. Tunod, Ma. Beatrice M.
Vega

Faculty of Pharmacy, University of Santo Tomas, Metro Manila, Philippines

ABSTRACT
The ethanolic leaf extract of Basella alba L. has been shown to possess diuretic activity owing to the presence of
saponins. The study focused on the formulation of a diuretic oral suspension, as potential alternative to oral solid diuretic
medications, using the ethanolic leaf extract of Basella alba L. as active ingredient. The ethanolic leaf extract of Basella
alba L. is an aromatic, non-greasy, dark green, viscous mass. The test for saponins produced a positive result in the froth
test, but a negative result in the hemolysis test. Differential scanning calorimetry (DSC) analysis showed potential inter-
actions with sodium chloride, carboxymethylcellulose, sodium benzoate, citric acid, and saccharin. An oral suspension
containing 50 mg/mL of Basella alba leaf extract was formulated, and viscosity, pH, and redispersibility were assessed.
Using the Lipschitz method, diuretic activity was evaluated using four groups of Sprague Dawley rats: Group 1 was
treated with base suspension, Group 2 received 10 mg/kg Furosemide, and Groups 3 and 4 were treated with 250 mg/kg
and 500 mg/kg of the oral suspension formulation, respectively. The urine volume and electrolyte content of the urine
were used as indicators of diuretic activity. The formulated oral suspension with the dose of 500 mg/kg showed a compa-
rable diuretic effect to furosemide having a diuretic index of 1, while the formulated oral suspension with a dose of 250
mg/kg produced a mild diuretic effect. There was a significant increase in urine Na+ and K+ output after administration of
both test concentrations of Basella alba oral suspension (p<0.0001). The study findings suggest that the formulated oral
suspension containing Basella alba leaf extract exhibits dose-dependent diuretic activity and maybe a suitable alternative
to oral solid diuretic medications.

Keywords: Basella alba Linn., diuretic, furosemide, oral suspension, saponins

1. Introduction sitivity, ototoxicity and electrolyte imbalance due to di-

Diuretics refer to drugs which enhances the rate
of urine flow. Water and electrolytes such as sodium,
potassium, chloride and bicarbonate are also regulated in
the presence of diuretics. These are the body's response to
maintain a state of homeostasis. There are different types
of diuretics according to their mechanism of action.
Furosemide, the prototype drug from a type of diuretic
called Loop diuretics inhibits Sodium Potassium Chloride
transporter in the thick ascending loop of Henle. It is also
known for increasing renal blood flow and reducing the
pulmonary congestion and left ventricular filling pres-
sures in the heart (Katzung & Trevor, 2017), thus making
it the drug of choice for heart failure. Adverse effects as-
sociated with taking this drug are hypovolemia, hypersen-
uresis (Se & Sang, 2015).
Basella alba L., from the Basellaceae family, is
locally known in the Philippines as “Alugbati”. It is a fast
growing perennial climbers that is commonly known as
Ceylon spinach or Malabar Spinach (Kumar, et al., 2013).
Phytochemical Studies shows that the leaves of
Basella alba L. have been found to contain vitamin K
(Adegoke and Ojo, 2017). It also contains basella
saponins A, B, C, and D, which are responsible for the
diuretic activity. An alternative theory is that the potassi-
um content of the plant also contribute to the diuretic ac-
tivity (Hostettmann and Marston, 2015). Sridevi, K.. ,
Ravishankar, K. & Kiranmayi, G.V.N. (2014) stated that
the ethanolic leaf extract of Basella alba L. can increase
urine output as well as in the sodium, potassium, chloride
and bicarbonate excretion.
 Suspensions are a class of dispersed system in
which a finely divided solid is dispersed uniformly in a
liquid dispersion medium (Nutan, M. & Reddy, I., 2010).
In formulating a suspension, excipients are employed to a
certain extent to allow the production of a stable and un-
compromised preparation. The essential components of a
pharmaceutical suspension that are commonly used are
the active ingredient, wetting agent/surfactant, flocculat-
ing agent, suspending agent, buffers, preservatives, sol-
vent, and flavorant.
Physicochemical tests according to the ASEAN
Guideline on Stability of Drug Products Oral Solutions
and Suspensions (2013) should be evaluated for appear-
ance (including formation of precipitate, clarity for solu-
tions), color, odour, assay, degradation products, pH, vis-
cosity, preservative content and microbial limits. Addi-
tionally for suspensions, redispersibility, rheological
properties, mean size and distribution of particles should
be considered. After storage, sample of suspensions
should be prepared for assay according to the recom-
mended labeling (e.g. shake well before using).
Lipschitz Method is a standard method and a very
useful tool for screening of potential diuretics. The
method is based on water and sodium excretion in test
animals (Nayak, et al.,2012). Analysis of the urine in-
cluded estimation of sodium and potassium ions (Sridevi,
K., et al. 2014) and Ion selective electrodes or ISEs is
used due to its ability to provide a wide range of ap-
plications in determining ions in water and other medi-
ums where such determinations are not subject to inter-
ferences (Department of Chemistry and Biochemistry,
New Mexico State University, n.d.).
This study centers on the development of an oral
suspension to aid in the administration of the diuretic
formulation to geriatrics. Since there are no commercially
available oral diuretic suspensions, the study aimed to
determine if the ethanolic leaf extract which possesses
diuretic activity will elicit the same activity if it is formu-
lated into an oral suspension.
2. Materials and Methods
2.1 Study Design
The leaves of Basella alba Linn. plant were pur-
chased from Dizon Farms. Analytical grade ethanol used
for Soxhlet extraction was purchased from Belman Labo-
ratories. All excipients used for the formulation of the
suspension such as carboxymethylcellulose, citric acid,
sodium benzoate, saccharin, and sodium chloride were
purchased from Alysons' Chemical Enterprises. The ref-
erence drug whis is Furosemide (Lasix) was purchased
from Mercury Drugstore.
Sprague-Dawley rats weighing approximately
100-200g were utilized as test animals and were obtained
from Joselito Plegaria Biological and Zoological Supply.
The rats were acclimatized in metal cages in a room tem-
perature of 25 ± 2°C in the animal housing facility of Re-
search Center for the Natural Sciences and Applied Sci-
ences (RCNAS), University of Santo Tomas for 7 days.
The animals were kept at constant 12 hours light/dark
cycle and were given free access to food and water for the
entire duration except 17-24 hours prior to the experi-
ment.. This study protocol was approved by the Institu-
tional Animal Care & Use Committee of University of
Santo Tomas (UST-IACUC).
2.1.1 Extraction Procedure
Fifty grams of leaves were cleared from dirt by
washing the leaves with water. After washing, the leaves
were left to dry in a shade inside a room for a span of two
weeks. After thorough drying, the dried leaves were
coarsely powdered using a mechanical grinder. The pul-
verized leaves were subjected to Soxhlet method of ex-
traction. 200 mL of ethanol was added to the round bot-
tom flask, attached to the Soxhlet extractor and condenser
on an isomantle. The ground leaves were loaded into the
thimble. Using the isomantle, the solvent was heated and
was made to evaporate, moving through the apparatus to
the condenser. The process ran for a total of 16 hours
(Redfern, et al., 2014). Once the process has finished, a
rotary evaporator at 50°C under reduced pressure was
used to evaporate the ethanol solvent, leaving a small
yield of extracted plant material (about 2 to 3 mL) in the
glass bottom flask. The extract was stored until the time
of the formulation.
2.1.2 Test for Saponins
2.1.2.1 Froth Test
One gram of the extract was dissolved in 10 mL
of distilled water in a test tube and vigorously shaken for
1-2 minutes.
Formation of honeycomb froth of 1 cm in height
that lasted for a minimum of 30 minutes indicated the
presence of saponins (Gunjal, et al., 2015).
2.1.2.2 Hemolysis Test
A blood agar plate was used with four equidistant
wells with depths of 4 mm cut using a sterile well cutter.
120 µl each of the ethanolic extract, ethanol, aqueous
Gugo solution (positive control) , and sterile water (nega-
tive control) were dispensed into the wells and kept cov-
ered for 24 hours at room temperature (Almutairi & Ali,
2015).
 A presence of a zone of clearance or “hemolytic
halo” is an indication of the presence of saponins.
2.1.3 Differential Scanning Calorimetry
The materials tested for compatibility with the
active ingredient are found in Table 1. Binary mixtures
were obtained by manual mixing of 1:1 ratios of the
ethanolic leaf extract and excipients using a mortar and
pestle. Thermal behavior was assessed by weighing 4–6
mg of the sample, enough to uniformly cover the alu-
minum crucible. A heating rate of 20°C min−1 was used
during this study. The thermograms for each excipient
and ethanolic extract was compared to the thermogram of
the binary mixture. If there are no insignificant changes in
the peaks of the thermogram of the extract and the binary
mixture, the excipient and the ethanolic leaf extract are
considered compatible. Glycerin was also tested for ther-
mal stability using the oven by subjecting it to 50 ℃ for a
duration of 4 weeks.
Table 1. Excipients used for compatibility test
Components of an Oral
Excipients
Suspension
Flocculating agent Sodium chloride
Viscosity/ thickening
Carboxymethycellulose
agent/ suspending agent
Buffer Citric acid
Preservative Sodium benzoate
Sweetener Saccharin
2.1.4 Formulation of the Oral Suspension
To prepare a 50 mL oral suspension, 1.4 g of car-
boxymethylcellulose was first triturated in a mortar and
pestle. Followed by the addition of the ethanolic leaf ex-
tract of Basella alba L. mixed with 10 mL glycerin. 10
mL volume of water was added prior to the addition of
the following: 0.01 g of citric acid, 0.5 g of saccharin, 0.1
g of sodium chloride and 0.1g of sodium benzoate. Final-
ly, water was added in portions to bring up the final vol-
ume of 50 mL with continuous trituration to ensure a uni-
form product.


2.1.5 Physicochemical Tests for Oral Suspensions
2.1.5.1 Organoleptic Evaluation
Organoleptic properties of formulated suspension
were described based on color, odor, and consistency.
Samples were placed at 300 C and 500 C temperature con-
ditions for one week and reassessed using the same para-
meters.
2.1.5.2 pH Determination
The pH of the formulated suspension was mea-
sured using a calibrated pH meter. Three independent
measurements were made. The pH of samples stored at
room temperature and oven conditions for one week were
compared to the pH of the suspension at time 0.
2.1.5.3 Viscosity Determination
Viscosity of the different samples were measured
using a Brookefield viscometer, spindle 4 at 250 rpm.
2.1.5.4 Redispersibility Evaluation
Cylinder was used to determine the redispersibili-
ty of the suspension based on the method reported by
Mittal and co-workers (2014). The suspension was al-
lowed to settle in a measuring cylinder. The mouth of the
cylinder was closed and inverted to 180º and the number
of inversions necessary to restore a homogeneous suspen-
sion was determined. If the homogeneity of the suspen-
sion was attained in one inversion, then the suspension is
considered 100% easily redispersible. Every additional
inversion decreases the percentage of ease of redis-
persibility by 5%.
2.1.6 Test for Diuretic Activity
2.1.6.1 Lipschitz Method
The test was performed following the Lipschitz
method with minor modifications. 12 male Sprague Daw-
ley rats each housed individually in a metabolic cage.
Food was withdrawn 17 hours before the experiment but
water was given ad libitum. On the day of the experiment,
gentle pressure was applied to the urinary bladders of the
rats to have a uniform content of urine. Followed by that
was the administration of 15 mL saline solution as a
priming dose to all of the test animals regardless of body
weight (Ganegamage, Abeytunga, and Ratnasooriya,
2014). An oral gavage was used to introduce the base
suspension, furosemide (Lasix) dispersed in the base sus-
pension, and the formulated suspension with the dosage
strength of 50 mg/mL. The first group which is the nega-
tive control group was treated with the base suspension
and the second group which will serve as the positive
control group, was treated with 10 mg/kg furosemide
(Lasix) dispersed in the base suspension. After the admin-
istration of the two suspensions a washout period was
done for 1 week. After the washout period, the first and
second group acted as the third and fourth groups, respec-
tively. The third group and fourth group received varying
250 mg/kg and 500 mg/kg doses of the formulated oral
suspension respectively.
For the collection of the urine sample, the tip of the tails
of the rats were tugged and the urine volume for analysis
of Na+ and K+ was recorded 5 hours after drug adminis-
tration (Sridevi, K., et al. 2014). From the performed pro-
cedure, the diuretic index of the urine output was calcu-
lated. Diuretic index is the ratio between the volume of
excretion of the test groups and the standard group,
(Asif, Jabeen, Atif, Majid, & Qamar-Uz-Zaman, 2015)
checked using Anderson-Darling test. Duncan Multiple
Range test was used to check what levels of the treatment
are significantly different from each other with respect to
the dependent variable. All tests were performed at 0.05
level of significance.
3. Results
3.1 Ethanolic Leaf Extract of Basella alba L.
The ethanolic leaf extract of Basella alba L. is a
non-greasy, dark green mass, with a viscous consistency
and an aromatic odour.
3.2 Test for Saponins
The ethanolic leaf extract produced a honeycomb
froth with the height of 1 cm which lasted for a few min-
utes. However, it did not produce a zone of hemolysis or
“hemolytic halo”.

Diuretic Index (I.D.) = VT / VS 3.3 Differential Scanning Calorimetry Analysis

Where The thermogram of the ethanolic extract of Basel-

la alba L. showed that a thermal process occurred be-
VT - Total volume of urine excreted by the test groups tween 106.81°C to 135.80 °C, producing and endothermic
peak at 116.50 °C, and an enthalpy of 243.9739 J/g. This
VS- Total volume of urine excreted by the standard is probably related to the loss of volatile constituents of
the sample, such as ethanol. Moreover, thermographs
2.1.6.2. Urinary Electrolyte Excretion presenting the various excipients utilized and its binary
mixtures caused the disappearance of the peak of the ex-
Urine samples were submitted to Majime Animal
tract but retained the peaks of the excipient.
Diagnostics Laboratory in Antipolo for analysis of sodi-
Aside from the aforementioned solid excipients,
um and potassium ions. The diagnostic laboratory utilized
glycerin and the binary mixture of the extract with glyc-
Hitachi Ion Selective Electrodes for both sodium and
erin stored at 50℃ for one week were found to be physi-
potassium content determination. Urine samples were
cally stable.
incubated for 24 hours. The urine was allowed to equili-
brate at room temperature, then mixed properly to ensure
3.4 Formulation of the Oral Suspension
homogeneity. A volume of 250 µl from a urine sample
was aspirated and added to 250 µl of the urine diluent
Three prototypes were made until the final com-
which is ISE diluent Gen 2. Inversion of the mixed urine
position of the formulation was achieved. The final com-
sample and diluent was repeated for 5-10 times. The urine
position is stated in table 2. The first and second proto-
samples were then loaded into the instrument. All calcula-
type produced a gummy consistency so the suspending
tions were performed by the system using a machine-
agent was reduced. In addition to that, the first two proto-
stored calibration curve.
types exceed the suggested amount of saccharin in an oral
suspension and have pH values which were below the
2.2 Statistical Analysis
specified range of 4-6, therefore saccharin and citric acid
Two separate Analysis of Covariance (ANACO- were reduced to render a more appropriate formulation.
VA) tests were performed. One using sodium as the de-
pendent variable and one using potassium as the depen-
dent variable. The treatments were the categorical vari-
able and urine amount was the continuous variable that
may affect sodium or potassium. The assumptions of
ANACOVA are normality of error terms and constancy of
variance. Since there is a duality nature between the two
assumptions, only the normality of error terms was
 Table 2. Quantitative Compositions of the Prototype stand for 1 week at a 40º C oven has the largest differ-
Formulations ence, which may be caused by heat which can influence
the viscosity of a suspension. The result is that liquids
Prototype
Prototype show a reduction in viscosity with increasing tempera-
Excipient Prototype1 3
2 tures.
(FINAL)
CMC 5.0 g 3.5 g 2.5 g 3.5.4 Redispersibility

Glycerin Upon standing of the suspension at baseline, sus-

10 mL 5.0 mL 10 mL
pension allowed to stand for 1 week at room temperature,
Extract and suspension allowed to stand for 1 week at 40ºC oven
2.5 g 2.5 g 2.5 g
in a measuring cylinder, the suspensions all required one
Citric acid inversion to render them homogenous. The results indi-
0.1 g 0.05 g 0.01 g cate that the three formulated suspensions are all 100%
Saccharin easily redispersible regardless of the environment and
5.0 g 2.0 g 0.5 g temperature condition.
NaCl 1.0 g 0.5 g 0.1 g 3.6 Test for Diuretic Activity
Sodium
3.6.1 Lipschitz Method
0.1 g 0.1 g 0.1 g
benzoate
The diuretic activity of the sample formulations
Water 50 mL qs 50 mL qs were determined as the diuretic index of the different
50 mL qs ad formulations. The result for the base suspension showed a
ad ad
diuretic index of .60 which is not included in any range
specified thus attributed to no diuretic activity. The 250
mg/kg suspension resulted to a diuretic index of 0.82
which falls in the range for mild diuretic effect, lastly the
3.5 Physicochemical Tests for Oral Suspensions
500 mg/kg suspension produced the greatest diuretic in-
3.5.1 Organoleptic Evaluation
dex among the three, the diuretic index is 1.00 which is in
the range of moderate diuretic effect.
The suspension at baseline, the suspension al-
lowed to stand for 1 week at room temperature, and the
Table 3. Diuretic Indices of the Formulated Suspen-
suspension allowed to stand for 1 week at 40º C oven re-
sions
tained the same color, odour, and taste. However, the sus-
pension allowed to stand for 1 week at 40º C oven dif- Treatment Diuretic Index
fered in consistency because it produced a less viscous
property when compared to the first two suspensions. Base suspension 0.60

Formulated oral
3.5.2 pH 0.82
suspension (250 mg/kg)
The pH of the suspension at baseline, suspension Formulated oral
allowed to stand for 1 week at room temperature and the 1.00
suspension (500 mg/kg)
suspension allowed to stand for 1 week at 40ºC oven were
4.102, 4.586, and 4.574 respectively. Therefore, the pH of
each suspension conforms with the specifications indicat-
ed for suitability for use. 3.6.2. Urinary Electrolyte Excretion

There is enough evidence that the ANACOVA

3.5.3 Viscosity


The suspension at baseline and the suspension
allowed to stand for 1 week at room temperature didn’t
largely differ, this may be attributed to the constant tem-
perature environment in which they were formulated and
located. On the other hand, the suspension allowed to
will give reliable results at 0.05 level of significance.
However, to confirm if there is an overall statistically sig-
nificant difference among the treatment levels, a post hoc
test was conducted.
For the Duncan test of Na+ ion, the formulated
oral suspension has a significantly higher mean than the
base suspension and a significantly lower mean than the the excipients. On the other hand, glycerin is compatible
standard (Furosemide) suspension. with the excipient based from its physical attributes. The
results of the formulations showed that increased amounts
Table 6. Duncan Grouping for Sodium Ion of the suspending agent will render a suspension with a
gummy consistency. Saccharin, in amounts higher than
Duncan
Mean Treatment the specified range, will render a suspension too sweet-
Grouping
tasting and will be harmful to patients. Lastly, citric acid
A 16.500 Furosemide suspension will extremely lower pH if added in excess. The three
formulated oral suspensions showed uniformity on their
Formulated oral suspension organoleptic properties. However, a Slight decrease in
B 12.667
(500 mg/kg) viscosity for sample stored at 400 was observed and may
be considered a natural behavior of systems at elevated
B temperature All pH determinations were within the ac-
ceptable range of 4-6. The use of citric acid as a buffer
Formulated oral suspension
B 11.000 efficiently resisted pH changes and stabilized the suspen-
(250 mg/kg)
sions for a given period and temperature condition.
C 7.833 Base suspension Therefore, the pH of each suspension conforms with the
specifications indicated for suitability for use (Patil, P.D.,
Means with the same letter are not significantly different
Desai, S.R., & Disouza, J., I. 2016). The formulated sus-
pensions subjected in differing environments are all 100%
For the Duncan test of K+ ion, the formulated oral suspen-
sion has a mean not significantly higher standard easily redispersible. The formulated oral suspension with
(Furosemide) suspension and significantly higher mean the dose of 500 mg/kg showed a comparable diuretic ef-
than the base suspension. fect to furosemide. As reported by Sridevi, Ravishankar,
and Kiranmayi (2014), the 500 mg/kg dose of formulated
Table 7. Duncan Grouping for Potassium Ion suspension showed an enhanced diuretic effect compared
to the dose of 250 mg/kg. Both doses of the formulated
Duncan
Mean Treatment oral suspensions gave significantly higher Na+ and K+
Grouping
output than the base suspension (p<0.05). One important
A 21.000 Furosemide suspension factor which determines the variability of the response to
diuretics is the level of endogenous urine electrolyte se-
A cretion, since high levels of these in the urine manifest
diuresis. (Campus, n.d.).

Formulated oral suspension

B A 19.167
(500 mg/kg)
B The study shows that the formulated oral suspen-
Formulated oral suspension
B 18.000
(250 mg/kg)
C 11.667 Base Suspension
Means with the same letter are not significantly different
4. Discussion
The ethanolic leaf extract produced a positive
result in the froth test but a negative result in the hemoly-
sis test. This may be due to the low concentration and
incomplete diffusion of the ethanolic extract on the blood
agar plate. Thermograms of the ethanolic extract, in com-
parison with the corresponding excipients and its corre-
sponding binary mixture are found to be incompatible on
the basis of disappearance of the characteristic peak
found in the ethanolic extract alone. This may be an indi-
cation of an interaction between the active ingredient and
5. Conclusion
sion of Basella alba L. is effective in inducing diuresis
and may be used as an alternative medication to solid di-
uretic dosage forms.
6. Recommendations
For future establishment and investigation, the
researchers recommend the following:
1. Confirm excipient compatibility using an alternative
method like Fourier-transform infrared spectroscopy
(FTIR).
2. Conduct stability testing of formulated suspension.
3. Develop stability indicating assay for formulated sus-
pension.
4. Investigate mechanism of action of the diuretic activity
of Basella alba L. extract.
Conflicts of Interest
No conflict of interest to declare.
Acknowledgements
The authors would like to express their deepest
gratitude to the Research Center of Natural and Applied
Sciences (RCNAS) of the University of Santo Tomas
(UST), the UST Herbarium and Asst. Prof. Gina C. Cas-
tro from the Faculty of Pharmacy of the University of
Santo Tomas for providing invaluable guidance and
knowledge throughout the research.
References
Ali, H., Ahmed, A., El-Haj, B. & Saad, R., (2016) Extem-
poraneous Furosemide Suspensions for Pediatrics Use
Prepared from Commercially Available Tablets. In-
ternational Journal of Pharmacy & Pharmaceutical Re-
search. Vol 5, Issue 2.
Almutairi, M., Ali, M., (2015) Direct Detection Of
Saponins In Crude Extracts Of Soapnuts By FTIR. School
of Civil Engineering & Surveying, University of
Portsmouth, Portland Building, Portland Street,
Portsmouth, United Kingdom. Retrieved from: https://
www.ncbi.nlm.nih.gov/pubmed/25537113
Amanpreet K., Shishu G., & Om. (2016) Thermal Analy-
sis and Quantitative Characterization of Compatibility
between Diflunisal and Lipid Excipients as Raw Materials
for Development of Solid Lipid Nanoparticles. Ther-
mochimica Acta. Vol 643, p2 3-32. Retrieved from
https://www.sciencedirect.com/science/article/pii/
S0040603116302490?via%3Dihub
Anandarajagopal K. , Sudhahar D. , Ajaykumar T.V. ,
Muthukumaran G. (2011). Evaluation of CNS Depressant
Activity of Aerial Parts of Basella alba Linn. IJPI J.
Pharmacol. Toxicol. 1:5.
Asif, M., Jabeen, Q., Atif, M., Majid, A., & Qamar-Uz-
Zaman, M. (2015). Diuretic Activity of Achyranthes as-
pera Linn Crude Aqueous Extract in Albino Rats. Tropical
Journal of Pharmaceutical Research,13(12), 2039. doi:
10.4314/tjpr.v13i12.14
Aziz, M.M., Saqib, N.U., Akhtar, N., Asif, H.M.,
Jamshaid, M., Sultana, S., & Bashir, K. (2013). Phyto-
chemical Screening and Evaluation of the Diuretic Activi-
ty of Aqueous Methanol Extract from Aerial Parts of
Mentha viridis Linn (Labiatae) in Albino Rats. Retrieved
from https://www.ajol.info/index.php/tjpr/article/view/
107891
Bajaj, S., Singla, D., & Sakhuja, N. (2012). Stability
Testing of Pharmaceutical Products. Journal of
Applied Pharmaceutical Science. 02 (03): 129-138.
Bamidele O, Akinnuga AM, Olorunfemi JO, Odetola OA,
Oparaji CK, Ezeigbo N (2010). Effects of Aqueous Ex-
tract of Basella alba leaves on Haematological and Bio-
chemical Parameters in Albino rats. African Journal of
Biotechnology. 9(41):6952-6955.
Bernal1, N. et al. (2014). Development, Physical-Chemi-
cal Stability, and Release Studies of Four Alcohol-Free
Spironolactone Suspensions for Use in Pediatrics. De-
partment of Pharmacy and Pharmaceutical Technology,
Faculty of Pharmacy, University of Barcelona. Avda. Joan
XXIII s/n, 08028 Barcelona, Spain. dx.doi.org/10.14227/
DT210114P19
Bhaduri L., Padmanabha R., Pulla Sireesha S. , Ramya
Krishna. P. , & Nagarjuna Y. (2011) Comparative study
of Acacia nilotica and Acacia sinuata for diuretic activity.
Division of Pharmacology, Centre for Pharmaceutical
Research (CPR), Raghavendra Institute of Pharmaceutical
Education and Research, Chiyyedu, Anantapur, Andhra
Pradesh, India.2 (6):17-22
Bresolina,T.M., Camargo, S.S., Donald,R.M., Filhoa,
V.C., Quintãoa,N.L., Silvaa, C.M.,Silvaa. R.M. (2015)
Development of an oral suspension containing dry extract
of Aleurites moluccanus with anti-inflammatory activi-
ty.Revista Brasileira de Farmacognosia Vol. 26 (2016)
68–76
Coggins, Mark D. (2013) Evaluating Potential Diuretic
Overuse. Retrieved from: http://www.todaysgeriatricmed-
icine.com/archive/110113p5.shtml
Dash, D. (2018) Effect of Particle Size on Stability of
Suspension. Retrieved from:
https://www.pharmatutor.org/articles/effect-of-particle-
size-on-stability-of-suspension
Desai, S., Desai, D. G. & Kaur, H (2009) Saponins and
their Biological Activities. Pharma Times. Vol. 41 No.
3.Retrieved from:https://www.researchgate.net/publica-
tion/288126191_Saponins_and_their_biological activities
Deshmukh, S. & Gaikwad, D. (2014) A review of the tax-
onomy, ethnobotany, phytochemistry and pharmacology
of Basella alba (Basellaceae). Journal of Applied Pharma-
ceutical Science Vol. 4 (01), pp. 153-165. DOI:10.7324/
JAPS.2014.40125
Devgan, M., Maddi, R. & Sarkar, B.K. (2014) Develop-
ment and Evaluation of Oral Herbal Formulations of Lan-
tana Camara Extract. Journal of Pharmaceutical Biology,
Vol. 4(1): 23-25.
Development and Stability Evaluation of Liquid Crystal-
Based Formulations Containing Glycolic Plant Extracts
and Nano-Actives. (2018). Cosmetics,5(2), 25. doi:
10.3390/cosmetics5020025
Diuresis [Def. 1]. (n.d) National Cancer Institute Dictio-
nary of Cancer Terms . Retrieved from: https://www.can-
cer.gov/publications/dictionaries/cancer-terms/def/diure-
sis
D. Kiss, et al. (2006) Application of DSC and NIRS to
Study the Compatibility of Metronidazole with Different
Pharmaceutical Excipients. Journal of Thermal Analysis
and Calorimetry, Vol. 84 (2006) 2, 447–45. Retrieved
from: https://link.springer.com/article/10.1007/
s10943-005-7257-9
Extracts [Def. 2]. (n.d.) Miller-Keane Encyclopedia and
Dictionary of Medicine, Nursing, and Allied Health, Sev-
enth Edition. (2003). Retrieved March 31 2018 from
https://medical-dictionary.thefreedictionary.com/extracts
Flocculation [Def. 3]. (n.d.) Retrieved from: https://
www.britannica.com/science/flocculation
Furosemide [Def. 4]. (n.d.) EMedicineHealth retrieved
from: https://www.emedicinehealth.com/drug-
furosemide/article_em.htm
Ganegamage, S., Abeytunga, Y., & Ratnasooriya, W.
(2014). Antidiuretic Activity of the Methanol Extract of
Aporusa lindleyana Wight (Euphorbiaceae) Baillon in
Rats. Tropical Journal of Pharmaceutical Research,13(7),
1099. doi:10.4314/tjpr.v13i7.13
Geriatrics [Def. 5]. (n.d.) MedicineNet Retrieved from:
https://www.medicinenet.com/script/main/art.asp?arti-
clekey=18385
Govindasamy, G., Krishnamoorthy, K., Rajappan, M.
(2013) selection of excipients for nanoparticles formula-
tions of nateglinide through drug-excipients compatibility
study. International Journal of Pharmacy and Pharmaceu-
tical Sciences. Retrieved from https://innovareacademic-
s.in/journal/ijpps/vol5suppl2/6832.pdf
Ingleson, K. (2017). Ascites: Causes, symptoms, and
treatment. Retrieved from https://www.medicalnewsto-
day.com/articles/318775.php
Ion Selective Electrodes (ISE) are membrane electrodes.
(n.d.). Retrieved from New Mexico State University web-
site: https://web.nmsu.edu/~kburke/Instrumentation/IS_-
Electrod.html?fbclid=IwAR1jaLjb7fgw2PourceeepUb-
y1LCTbzvEWs270Mxqt7o0An9NJ5-H-9_rM
Isolate [Def. 6]. (n.d) Collins dictionary. Retrieved from:
(https://www.collinsdictionary.com/dictionary/english/
isolate)
Kaur, K. (2013). Basics of an Ion-Selective Electrode.
Retrieved from https://www.azosensors.com/article.aspx?
ArticleID=244&fbclid=IwAR3sacrsrPcRjYEwnUhQ4w-
daUl4NLovW0KAsP-bAtWYT3AXFBGt77aIIKo0
Kodre KV, et al. (2014). Differential Scanning Calorime-
try: A Review. Research and Reviews: Journal of Phar-
maceutical Analysis, Vol. 3, Issue 3.
Logrippo, S. et al. (2017) Oral drug therapy in elderly
with dysphagia: between a rock and a hard place! 64:9-20
Retrieved from: https://www.ncbi.nlm.nih.gov/pmc/arti-
cles/PMC5293185/
Madjid, S., Naser, D.M., Djavad, F. (2003) Prevention of
crystal growth in acetaminophen suspension by the use of
polyvinyl pyrrolidone and bovine serum albumin. De-
partment of Pharmaceutics, Faculty of Pharmacy, Mazan-
daran University of Medical Sciences, Sari, Iran, Vol.
11(3)
Marciano, M. (2018) Saponins. The Naturopathic Herbal-
ist. Retrieved from: https://thenaturopathicherbalist.com/
plant-constituents/saponins/
Murthy, P. N., Devi, M. V., Sahoo, S. K., Mahapatra, A.
K., Khandai, M. (2015) Evaluation of sedimentation sta-
bility in paracetamol suspensions with Plantago ovata
mucilage as suspending agent using near-infrared trans-
mission measurements. Der Pharmacia Lettre 7 (7):85-96.
Retrieved from : http://www.scholarsresearchlibrary.com/
articles/evaluation-of-sedimentation-stability-in-parac-
etamol-suspensions-withplantago-ovata-mucilage-as-sus-
pending-agent-using-n.pdf
Pal, Dilipkumar & Nayak, Amit & Kalia, Samir. (2010).
Studies on Basella alba L. leaves mucilage: Evaluation of
suspending properties. International Journal Drug Discov
Tech. 1. 15-20.
Pani, N.R., Nath, K.L., Acharya, S. (2011) Compatibility
studies of nateglinide with excipients in immediate re-
lease tablets. Acta Pharm. 61 (2011) 237–247 DOI:
10.2478/v10007-011-0016-4. Retrieved from https://
www.researchgate.net/publication/51230327_Compatibil-
ity_studies_of_nateglinide_with_excipients_in_immedi-
ate_release_tablets
Patil P. D , Desai S. R, Disouza, J. I. (2016) Stability
analysis and quantitative evaluation of metronidazole
suspension. Indian J. Pharm. Biol. Res., 2016; 4(3):5-14.
Retrieved from http://ijpbr.in/pdf/2-Stability-analysis-
and-quantitative-evaluation-of-metronidazole-suspen-
sion.pdf?fbclid=IwAR0McpUNc56P1L3kfama_8-
SOpxmzTva-L3851jhK3F5Ksbdi5Cc6RAq44lQ
Peterson, M. (2013). Toxicologic Decontamination. Small
Animal Toxicology 3rd edition. Retrieved from:
https://www.sciencedirect.com/topics/pharmacology-toxi-
cology-and-pharmaceutical-science/forced-diuresis
Pimple, A., Shahi, S., Khan, S., Shaikh, S., Chate, R. &
Dube, A. (2017) Oral Suspension: A Review. Journal of
Medical and Pharmaceutical Innovation. 4(20). Retrieved
from: http://www.jmedpharm.com/index.php?
journal=JMPI&page=article&op=view&path%5B%5D=2
49&path%5B%5D=310Plant, L. (2003). Clinical Use of
Diuretics. CME Renal Medicine, Vol 3 No 6 (2003). Re-
trieved from: https://pdfs.semanticscholar.org/1f86/0cd-
d6479f7315b1b326e2f1d9ed41cd989f0.pdf
Pirkle, J. (2016, March). Roche Ion-Selective Electrode.
Retrieved from https://wwwn.cdc.gov/nchs/data/nhanes/
2013-2014/labmethods/URLT_H_R_MET_Elec-
trolytes.pdf?fbclid=IwAR1tNiOtLD8lXe_9NIM7WZZh-
IVzBTMP2vx4RprNxx_OXtc4NiVUk0k5Yxw8
Proel, E. & Nelson, W. (2018). The Flame Photometer in
Determination of Sodium and Potassium. American Jour-
nal of Clinical Pathology, Vol 20, Issue 9, (1950) Pages
806–813. Retrieved from: https://doi.org/10.1093/ajcp/
20.9.806
Qavi, A., Kamal, R., & Schrier, R. (2015). Clinical Use of
Diuretics in Heart Failure, Cirrhosis, and Nephrotic Syn-
drome. International Journal of Nephrology, Vol 2015
(2015). Retrieved from: https://www.hindawi.com/jour-
nals/ijn/2015/975934/
Rathee S, Ahuja D, Rathee P, Thanki M, Rathee D (2010).
Cytotoxic and Antibacterial Activity of Basella alba
Whole Plant: A Relatively Unexplored Plant. Pharmacol-
ogyonline 3:651-658.
Redfern, J., Kinninmonth, M., Burdass, D., & Verran, J.
(2014). Using Soxhlet Ethanol Extraction to Produce and
Test Plant Material (Essential Oils) for Their Antimicro-
bial Properties. Journal of Microbiology & Biology Edu-
cation, 15(1), 45–46. Retrieved from: http://doi.org/
10.1128/jmbe.v15i1.656
Roshan Adhikari, Naveen Kumar HN, Shruthi SD.
(2012). A Review on Medicinal Importance of Basella
alba L. International Journal of Pharmaceutical Sciences
and Drug Research. 4(2): 110-114
Sridevi, K. , Ravishankar, K. & Kiranmayi, G.V.N.
(2014). Evaluation of Diuretic and Antiurolithiatic Activi-
ties of Ethanolic Leaf Extract of Basella alba. In-
ternational Journal of Pharmacy. 4(1): 145-149
Shukla, S., Patel, R., and Kukkari, R. (2009) Study of
Phytochemical and Diuretic Potential of Methanol and
Aqueous Extracts of Aerial Parts of Phyla Nodiflora
Linn.. Retrieved from: http://ijppsjournal.com/
Vol%201%20Issue%201/114.pdf
Subhas, K., Kapoor, M. & Akshay R. P. (2015) Efficient
pharmacognostical, phytochemical and pharmacological
survey on Basella alba L. An ethno therapeutic plant. In-
ternational Journal of Pharmacognosy and Phytochem-
istry Vol. 2 (1) pp. 059-063. Retrieved from: http://inter-
nationalscholarsjournals.org/download.php?
id=936526414921807456.pdf&type=application/
pdf&file=Efficient%20pharmacognostical,%20phyto-
chemical%20and%20pharmacological%20survey%20on
%20Basella%20alba%20L.%20An%20ethno%20thera-
peutic%20plant.pdf.
The International Pharmacopoeia (2017). (7th Ed.). De-
termination of pH. Retrieved from: http://apps.who.int/
phint/pdf/b/7.1.13.1.13-Determination-of-pH.pdf
Tongco, J., Añis, A & Tamayo, P. (2015). Nutritional
Analysis, Phytochemical Screening, and Total Phenolic
Content of Basella alba Leaves from the Philippines. In-
ternational Journal of Pharmacognosy and Phytochemical
Research. 7(5); 1031-1033
Uddin, M. S., Mamun, A. A., Akter, N., Sarwar, M. S.,
Rashid, M., Amran, M. S. (2016) Pharmacopoeial Stan-
dards and Specifications for Pharmaceutical Oral Liquid
Preparations. Archives of Current Research International.
Retrieved from: https://www.researchgate.net/publication/
292130182_Pharmacopoeial_Standards_and_Specifica-
tions_for_Pharmaceutical_Oral_Liquid_Preparations
Ueoka, A. R., Moraes, C. A. P. (2018) Development and
Stability Evaluation of Liquid Crystal-Based Formula-
tions Containing Glycolic Plant Extracts and Nano-Ac-
tives. Cosmetics 5(2), 25 Retrieved from: http://
www.mdpi.com/2079-9284/5/2/25
U. S. Food and Drug Administration. (2014) Guide to
inspections oral solutions and suspensions.Retrieved
from:https://www.fda.gov/ICECI/Inspections/Inspection-
Guides/ucm074935.htm

Vijender K, Bhat ZA, Dinesh K, Puja B, Sheela S (2011).

In-vitro antiinflammatory activity of leaf extracts of
Basella alba linn. Var. Alba. International Journal of drug
development and research. April-June, 3(2):176-179

Viscosity[Def.8].(n.d.)ChemistryLibreTexts.Retrieved-
from:https://chem.libretexts.org/Core/Physical_and_The-
oretical_Chemistry/Physical_Properties_of_Matter/
States_of_Matter/Properties_of_Liquids/Viscosity

Woldemedhin, B., Nedi, T., Shibeshi, W., & Sisay, M.

(2017). Evaluation of the Diuretic Activity of the Aque-
ous and 80% Methanol Extracts of the root of Euclea
divinorum Hiern (Ebenaceae) in Sprague Dawley rats.
Journal of Ethnopharmacology,202, 114–121. doi:
10.1016/j.jep.2017.01.015

BOOKS:
Abramson, S. (2010). Extracorporeal Treatment of Poi-
sonings. Chronic Kidney Disease, Dialysis, and Trans-
plantation (3rd Ed.). Philadelphia, PA: Saunders, an im-
print of Elsevier Inc.

Bardal, S. K., & Martin, D.S. (2011). Toxicology. Applied

Pharmacology. Missouri, MO: Saunders, an imprint of
Elsevier Inc.

Hostettmann, K. & Marston, A. (2005, September 29).

Saponins. New York, NY: Cambridge University Press
Kulshreshtha, A. K., Singh, O. N., & Wall, G. M. (2010).
Pharmaceutical suspensions from formulation develop-
ment to manufacturing. New York: Springer.

Katzung, B. G., Kruidering-Hall, M., Trevor, A.J.

(2017).Pharmacology Examination and Board Review.
(11th Ed.).Chapter 15 Diuretics and Other Drugs that Act
on Kidney.New York: McGraw Hill Education.pp.
132-142

Peterson, M. E.(2013).Toxicologic Decontamination.

Small Animal Toxicology (3rd Ed.). Missouri, MO:
Saunders, an imprint of Elsevier Inc.

Remington, J. P., & Beringer, P. (2006). Remington: The

Science and Practice of Pharmacy. Philadelphia: Lippin-
cott Williams & Wilkins. Retrieved 2018-03-29.

Vogel, W. H. (2015). Drug discovery and evaluation:

Pharmacological Assays (H. G. Vogel, Ed.). New York:
Springer. p. 461