Furosemide (FUR) is a high-loop diuretic widely used for the treatment of conditions leading to excessive accumulation of
water in the body (edema), which are normally associated with cardiovascular disorders such as heart failure, infarction and
hypertension. It has been reported that FUR has a bioavailability problem and initially shows an adverse temporary peak diuretic
effect.1,2 To eliminate this, various efforts have been made to develop prolonged-release forms of FUR; however, it has been reported
that the bioavailability of such preparations decreases to 40–60% compared with conventional tablet forms.1
Although it has not been demonstrated for humans, animal studies of bioavailability have shown that there may be regions in
the stomach and/or the upper part of the small intestine where FUR is specifically absorbed; the short stay of controlled
release preparations in this specific region of absorption leads to bioavailability problems.3 Accordingly, developing a floating dosage form with a controlled release pattern aimed at prolonging the residence of FUR
at the site of maximum absorption (upper gastro intestinal tract (GIT)) has been proposed as an option for improving bioavailability
and, at the same time, reducing the side effect of peak diuresis associated with conventional formulations.4 However, formulating a gastro-retentive dosage form of FUR poses a great challenge because of FUR's insufficient aqueous
solubility in acidic mediums (pH 1.2), which may be considered the rate-limiting step in the absorption process.5,6
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The present work was conducted to develop a novel floating dosage form of FUR to enhance its solubility by formulating a self
microemulsifying drug delivery system (SMEDDS) of FUR, followed by its adsorption onto a mixture of high functionality excipient
(SMCC), matrix forming polymers (HPMC K4M and HPMC E50 LV) and a gas-generating agent (NaHCO3) to achieve a buoyant matrix with a controlled release profile.
Materials and methods
FUR was obtained from Aarti drugs (India). The following materials were donated by Gattefosse (India) and were used as received:
Labrafac CM10 (C8‑C10 polyglycolized glycerides), Masine 35-1 (Glyceryl momolinoleate), Lauroglycol FCC (Propylene glycol
laurate), Labrafil 1944 CS (Apricot kernel oil PEG 6 esters) and Labrafac PG (Propylene glycol caprylate/caprate). Cremophor
RH 40 (Polyoxyl 40 Hydrogenated castor oil), Cremophor EL (polyethoxylated castor oil) and Solutol HS 15 (Polyoxyethylene
esters of 12-hydroxystearic acid) were obtained from BASF (India). Gelucire 44/14 (PEG‑32 glyceryl laurate), 50/13 (PEG‑32
glyceryl palmistearate) and Triacetin (glyceryl triacetate) were received from Colorcon Asia (India). Span 20 (sorbitan monolaurate),
Tween 80 (polyoxyethylene sorbitan mono-oleate), PEG 400 and sodium bicarbonate were bought from Merck Co (India). Silicised
microscrystalline cellulose ProSolv SMCC was received from DMV International (India). Grades of hydroxypropyl methylcellulose
(HPMC K4M and HPMC E50 LV) were received as gift samples from Colorcon Asia. Deionised water was prepared using a Milli‑Q
purification system from Millipore (France).
Acetonitrile and methanol used in the study were of HPLC grade. All other chemicals were reagent grade. Empty hard gelatin
capsule shells were donated by ACG capsules (India).
The solubility of FUR in various components (oils, surfactants and cosurfactants) was determined and 500 mg of each of the
selected vehicles were added to each cap vial containing an excess of FUR (1 g). After sealing, the mixture was heated at
40 °C in a water bath to facilitate solubilisation. Mixing of the systems was conducted using a vortex mixer and the formed
suspensions were then shaken with a shaker at 25 °C for 48 h. After reaching equilibrium, each vial was centrifuged at 3000
rpm for 5 min and excess insoluble FUR was discarded by filtration using a membrane filter (0.45 µm, 13 mm; Whatman, USA).
The concentration of FUR was then quantified using HPLC.
Pseudo-ternary phase diagram
The pseudo‑ternary phase diagrams of oil, surfactant/cosurfactant (S/CoS) and water were developed using the water titration
method; the mixtures of oil and S/CoS at certain weight ratios were diluted with water in a dropwise manner. For each phase,
diagrams at a specific ratio of S/CoS, 1:1 and 3:1 (w/w), a transparent and homogenous mixture of oil and S/CoS was formed
under the mixing by vortexing for 5 min. Each mixture was then titrated with water and visually observed for phase clarity
and flowability. The concentrations of water at which turbidity‑to‑transparency and transparency‑to‑turbidity transitions
occurred were derived from the weight measurements, and these values were then used to determine the boundaries of the microemulsion
domain, corresponding to the chosen value of oils, as well as the S/CoS mixing ratio. The effect of adding triacetin to oil
phase‑on‑phase behaviour of the mixture was also studied. To determine the effect of drug addition on microemulsion boundary,
phase diagrams were also constructed in drug presence, wherein the oil component was enriched with the drug. Phase diagrams
were then constructed using Tri plot v1‑4 software.7
Preparation of SMEDDS
SMEDDS were prepared using Tween 80 and PEG 400 as the S/CoS combination, and Labrafac Hydro WL (with 40% triacetin) as the
oil component (Table 1). In all formulations, the level of FUR was kept constant (10% (w/w) of the total formulation weight). Briefly, accurately
weighed FUR was placed in a glass vial followed by the addition of oil and S/CoS, and the components were then mixed by gentle
stirring and vortex mixing, and heated at 40 °C using a magnetic stirrer until the FUR was perfectly dissolved. The formulations
were then subjected to three to four freeze thaw cycles, which included freezing at –4 °C for 24 h, followed by thawing at
40 °C for 24 h. The formulations were then observed for phase separation. Only formulations that were stable to phase separation
were selected for further studies.
Table 1: SMEDDS formulations.
Emulsion droplet size analysis
SMEDDS formulation (100 µl) was diluted to 250 mL in a beaker and gently mixed using a glass rod. The resultant emulsion was
then subjected to particle size analysis, using a Malvern Meta‑sizer equipped with 2000 Hydro MU (Malvern, India] with a particle
size measurement range of 0.02–2000 µm. Particle size was calculated from the volume size distribution and all studies were
repeated in triplicate, with good agreement being found between measurements.
Assessment of self emulsification
The evaluation of self‑emulsifying properties was conducted using visual assessment, as previously reported.8 In brief, visual assessment was performed by drop‑wise addition of the preconcentrate (SMEDDS) into 250 mL distilled water.
This was done in a glass beaker at room temperature and the contents were gently stirred magnetically at approximately 100
rpm. Compositions were then categorised on speed of emulsification, clarity and apparent stability of the resultant emulsion.
In vitro dissolution studies
The quantitative in vitro release test was performed in 900 mL of buffer pH 1.2, using USP XXIII Type 1 apparatus, at 37 +/- 0.5 °C. The basket shafts
were rotated at 50 rpm. The SMEDDS formulation was filled in hard gelatin capsules (1 size) and used for drug release studies
to compared with plain FUR. During the release studies, 5 mL sample of medium was taken out and subjected to drug analysis
using HPLC. The removed volume was replaced each time with fresh buffer pH 1.2. For determining the in vitro dissolution of plain FUR, the medium was changed to 900 mL of buffer pH 1.2 with Tween 80 (equivalent to the amount used
in formulation). Similarly, dissolution studies were also conducted in other mediums (buffer pH 4.5 and 7.2) to observe the
effect of pH on drug release.
HPLC analysis of FUR
FUR concentration in the samples was determined by HPLC analysis. The HPLC analysis system consisted of Jasco PU 980 Intelligent
pump (Jasco Pumps, India) and Jasco MD‑2015 plus multi‑wavelength detector. The chromatographic column was a C‑18 Lichrosphere
10RP-18e (5 µm) 4.6 mm x 250 mm (Merck, India). The chromatographic conditions were:
- mobile phase: methanol: 0.01 M KH2PO4 (37:63)
- flow rate: 1 mL/min; loop size: 100 µL
- detection at 274 nm and retention time 9.0 +/- 9.5 min.
The validation parameters were:
- linearity (range = 5–30 µg/mL, coefficient of correlation = 0.9999)
- limit of detection = 126 ng/mL
- limit of quantification = 420 ng/mL.