INTRODUCTION:
Suspension dosage form is a preferred and widely accepted dosage forms for insoluble or poorly soluble drugs for various therapeutic applications and has long been used for insoluble and poorly soluble drugs for making oral, topical and parenteral products. Physically stable dispersions of a drug in a suitable suspension vehicle are important to create a drug product with: consistent and reproducible dosage, predictable shelf life batch to batch consistency. Addition of a stabilizer or suspending agent which reduces the rate of settling and permits easy redispersion of any settled particulate matter both by protective colloidal action and by increasing the consistency of the suspending medium1-3. Suspending agents are (i) inorganic materials, (ii) synthetic compounds, or (iii) polysaccharides. Natural gums like Acacia, Tragacanth, Khaya, Karaya and Eulophia campestris belong to the latter group4. Gums have been wildly used as tablet binders, emulgents and thickeners in cosmetics and suspensions as film-forming agents and transitional colloids. There are reports about the successful use of Ocimum gratissimum, Butea monospermama, Albizia zygia gum and Leucaena leucocephala seed gum as suspending agent5-8.
Eulophia campestris Wall. (family Orchidaceae) is a genus of terrestrial orchids, an herbaceous plant. It is found throught India and indigenous to persia and Afganistan. Tubers of E. campestris yield a large quantity of mucilage to water and, on boiling even with 40 parts of water, form a thick jelly which is highly nutritious9-10. Aim of our present study to evaluate the suspending property of mucilage of the Eulophia campestris and forulate a deflocculated suspension. Gum of the Eulophia campestris has been reported to have binder potential for tablet formulations11.
MATERIALS AND METHODS:
The fresh tubers of E. camphesteris plant were collected from Mandav region of ujjain and authenticated in Botanay Department, Vikram University, Ujjain. Metronidazole (Cipla ltd., Vikhroli, Mumbai), gum tragacanth, acacia, sodium CMC (Loba Chemie, Mumbai) of pharmaceutical grade was used. All the other solvents, reagents used were of analytical grade.
Extraction of the Mucilage12-14:
Mucilage are viscid, somewhat tenacious and generally adhesive liquids, prepared with water as a solvent. The mucilage was isolated from freshly dried and coarsely powdered tubers of E. campestris. The tubers were steeped in water 24 h, boiled for 1 h and kept aside for 2 h to release mucilage into water. The material was squeezed in a muslin bag to remove the marc from the filtrate. The filtrate was again filtered under vaccum and added to equal volume of acetone to precipitate the mucilage. The mucilage was separated, dried in an oven at temperature less than 50°C, powdered and passed through sieve number 80. The powdered mucilage was stored in desicator until further use (yield: 30% w/w).
Phytochemical Examination:
Preliminary tests were performed to confirm the nature of mucilage obtained. The chemical tests that were conducted are: Ruthenium red test, Molisch test, test for reducing sugars and Ninhydrin test15.
PREPARATION OF METRONIDAZOLE SUSPENSION:
Metronidazole solution (4%w/v) in distilled water and tragacanth powder (1.0 g) was triturated with glycerin (10.0 ml) separately. The mixture was transferred to 100 ml amber coloured bottle, volume was made up with distilled water and then shaken vigorously for 2 min (thus making 1.0%w/v of the gum in the preparation). Suspension contained 0.1% w/v benzoic acid as a preservative. Similarly suspension of 2, 3, 4% w/v were produced with powder tragacanth. The procedure was repeated for acacia, sodium carboxy methyl cellulose and Eulophia campestris mucilage. All the suspension were deflocculated. To determine the degree of flocculation we have also prepared flocculated suspension using SLS 0.1% w/w as flocculating agent.
EVALUATION OF SUSPENSION:
Rate of separation:
The rate of separation of the suspensions were determined by keeping 50 ml portion of each suspensions in stoppered measuring cylinder and stored undisturbed at room temperature. The separation of clear liquid was noted at intervals of 5 d up to 30 d. The sedimentation volume, F (%), was then calculated using the following equation16
F = 100Vu/Vo
Where Vu is the ultimate volume of the sediment and Vo is the original volume of the suspension.
Degree of Flocculation:
The degree of flocculation was determined17 following the equation β= F/Fα, where F is the ultimate sedimentation volume in the flocculated suspension and Fα is the ultimate sedimentation volume in the deflocculated suspension.
Redispersion:
Fixed volume of each suspension (50 ml) was kept in calibrated tubes which were stored at room temperature for various time intervals (5d, 10…45 d). At regular interval of 5 d, one tube was removed and shaken vigorously to redistribute the sediment and the presence of deposit if any was recorded.
Rheology:
The time required for each suspension sample to flow through a 10 ml pipette was determined and the apparent viscosity (ηα in mls–1) was calculated using the equation :
Volume of pipette (ml)
Flow Rate = ηα =
Flow time (sec.)
The viscosity (in poise) of the samples was determined at 25 C using the Brookfield viscometer, model DV-E (Brookfield Laboratories, Massachusetts) at 30 revolutions/min (Spindle #4). All determinations were made in at least triplicate and the results obtained are expressed as the mean values.
Particle Size Analysis:
After shaking, 10 ml of each sample was separately transferred into 200 ml cylinder. Distilled water (150 ml) was then added, mixed, and 10 ml aliquot was removed at a distance of 10 cm below the surface of the mixture and at 1, 5, 10, 15, 20, 25 and 30 min. This was transferred into an evaporating dish and evaporated to dryness in an oven at 105 C and the residue weighed. The particle diameter (d in cm) was then calculated using the Stoke's equation18:
18ηh
d =
(ρs – ρo)
where, h is the distance of fall of the particle (cm), t is the time (s), η is the viscosity of the dispersion medium (poise), ρs – ρo is the density gradient between the dispersed particles and the liquid (g cm–3) and g is the gravitational constant (cm s–2).
Determination of the pH of the suspensions:
The pH of each of the prepared suspension was measured using pH meter (Digital Telethermameter, Model no.-111) at weekly intervals for 4w. for ease of redispersibility, 10 ml of each suspension was poured into four calibrated tubes, which were stored at room temperature for 1,2,3 and 4 w. at the end of each storage period, each tube was shaken at constant moderate rate of 30 shake/min. the time (s) taken to redisperse the sedimented suspension was recorded.
The method essentially consisted of holding the sample tube straight in upright position between two fingers with thumb at the bottom and the middle finger at the top followed by the almost uniform rotation through 1800 and brought back through the same path. The pair of successive upward and downward movement each of approximately equal force, constituted one complete shake. The number of shakes required for complete elimination of sediment from the bottom of the tube was recorded. At this juncture the sample was observed for homogeneity of the suspension and the total time(s) recorded to redisperse the sedimented suspension. This was based on the empirical understanding that not more than that force should be required and the same that is routinely applied by the consumer in the event of “shake well before use” maximum care was taken to exert approximately the same amount of force every time and the same time interval.
RESULT AND DISCUSSION:
It is quite understand that the better is the suspending medium the lesser the rate of sedimentation. Suspensions are routinely evaluated for their rate of separation which indicates its suspending property. To evaluate the suspending properties of the gum, suspensions were prepared with fixed concentration of metronidazole but with varying concentration of test mucilage (1.0 to 4.0%w/v) as well as the traditional suspending agents such as tragacanth, acacia and sodium CMC. The sedimentation volume profile of the suspension prepared with 1%, 2%, 3% and 4% of suspending agent shown in fig. 1, 2, 3 and 4 respectively. Here Eulophia campestris mucilage shows its superiority over tragacanth and acacia. Eulophia campestris mucilage showed a comparable result to that of sodium CMC.
Fig. 1: Sedimentation profile of suspension made with 1% concentration of Sodium CMC, E. campestris, tragacanth and acacia
Fig. 2: Sedimentation profile of suspension made with 2% concentration of Sodium CMC, E. campestris, tragacanth and acacia
Fig. 3: Sedimentation profile of suspension made with 3% concentration of Sodium CMC, E. campestris, tragacanth and acacia
Fig. 4: Sedimentation profile of suspension made with 4% concentration of Sodium CMC, E. campestris, tragacanth and acacia
able 1: Physical characterization of mucilage of E. camphestris
|
Parameters |
Observation |
|
Solubility |
Swells in cold water considerably but quickly dissolves in warm water forming viscous colloidal solution. Insoluble in ethanol, methanol, chloroform and ethyl acetate. |
|
Swelling index |
12.6% |
|
pH (1% w/v solution) |
7.4 |
|
Loss on drying |
8.6% |
|
Microbial load a. Bacteria (no. of CFU/g mucilage) b. Fungi (no. of CFU/g mucilage) |
110
105 |
The dispersed particles sediment at a faster rate in suspensions containing 1% and 2% w/v of suspending agent and the initial sedimentation during first 20 days are much faster than afterwards. The suspensions prepared with 3% suspending agent shown a constant decline in sedimentation volume up to 20 days whereas, the decline was minimized after 25 days. However the suspensions prepared with 4% suspending agent the change in sedimentation volume was minimum throughout the 45 days of study. According Martin etal.16 the sedimentation volume provides only a qualitative account of flocculation. The degree of flocculation (β) is more useful parameter, which is the ratio of ultimate sedimentation volume in the flocculated and deflocculated system. A comparison of β values (Table 3) of suspensions prepared with Eulophia campestris mucilage, gum tragacanth and sodium CMC shows a slight higher values at the 3 and 4% w/v level for Eulophia campestris mucilage and tragacanth. These observations show that Eulophia campestris mucilage is a better suspending agent than Sodium CMC. Since the suspension produces sediment on storage it must be readily dispersible so as to ensure the uniformity of the dose. If sediment remains even after shaking vigorously for specified time, the system is described as caked. The suspension with 1 and 2%w/v tragacanth have shown to be caked after 25 and 35 days respectively indicating its effectiveness as suspending agent at this concentration. However the suspensions with sodium CMC and Eulophia campestris mucilage were found to be redispersible irrespective of their concentration.