Experiment 4 – Suppository

INTRODUCTION 

Suppositories are a dosage form which administered by rectum. They are used for local or systemic action. They also are designed in suitable size and shape to be easily for administration. The good properties of suppositories are shown by the releasing of drug to site of action and give the desired effects after this dosage form is administered by the patient.

As usual characteristic in formulation, the drug which will incorporate in suppositories formulation must be compatible with suppositories’ base. The good or ideal suppositories’ base must have certain characteristics such as melt in body temperature or dissolve in body fluids, solidify quickly after melts, easily moulded and removed from mould, chemically stable, easy to handle, non-toxic, and non-irritant. The different composition of suppositories’ base in a formulation can influence the rate of releasing the drug from the suppositories. It also can influence the physical characteristic of those suppositories. In this experiment, we are instructed to do several suppositories formulation with different composition of suppositories’ base. The result will be discussed.

OBJECTIVE

To study the effects of cooperating different composition of bases on the physical characteristics of the suppository formed and its release rate of drugs.

APPARATUS AND MATERIALS

Weighing balance

Weighing boat

Spatula

50ml and 100ml beaker

Hotplate

5ml Measuring cylinder

Suppository mould

Water bath

10 cm Dialysis beg

Thread

Glass rod

5ml Pipette

Pipette bulb

Plastic Cuvet

UV Spectrometer

PROCEDURE

1. Paracetamol stock solution was prepared by dissolving 10g paracetamol in 5 mL distilled water.
2. Paracetamol suppositories were prepared based on the following formula:

Suppository	Group	Ingredient		Paracetamol Stock Solution (g)	Total (g)
		PEG 1000	PEG 6000
1	1,5	9	0	1	10
2	2,6	6	3	1	10
3	3,7	3	6	1	10
4	4,8	0	9	1	10

3. A suppository mould was used to shape the suppository.
4. After the suppository harden, one of the suppository was put in a beaker containing 10 ml of distilled water maintained at 37^o The time taken for the suppository to completely melt was taken.
5. Another suppository was inserted in a dialysis bag. Both ends of the dialysis bag were tied tightly. The bag was put in a beaker containing 50mL distilled water that was maintained at 37^o
6. At every 5 minutes interval, a sample aliquot was pipetted and aracetamol release from the suppository was determined using UV-visible spectrometer. The water inside the beaker was stirred before taking the sample.

QUESTIONS

1) Compare the physical appearance of the suppositories and give explanation.

The table above shows the comparison between the physical appearances of suppositories in different formulations depends on the effects of different amount of the Polyethylene Glycol (PEG) in the suppository base mixtures. All the four suppositories produced are bullet-shaped following the shape of mould and they are in white tone.

The two bases used in this experiment are PEG 1000 and PEG 6000. Since PEG 6000 consists of more hydroxyl (-OH) bonds in its chemical structure, it shows greater hydrophilicity compared to PEG 1000. Hence, the suppositories become less greasy from formulation I to IV. From the results, it is found that the hardness of the suppositories increases with increasing amount of PEG 6000. This is due to the physicochemical characters of PEG 6000 forms more strong and stable hydrogen bonds in the suppository compared to PEG 1000. This explains why suppositories with more PEG 1000 and less PEG 6000 are softer.

2. Plot a graph of the time needed to melt the suppository vs. the amount of PEG 6000 in the                   formulation. Compare and explain the results.

The time needed for the suppository to melt is 74.00 minutes for our group.

Based on the graph above, the average time needed for the suppository to completely melt decreases initially until PEG 6000 3ml and increases with the increase of the amount of PEG 6000 starting from 6mg. The time needed for the suppository with 0g of PEG 6000 to melt is supposed to be the shortest. However, errors may have occured in the procedure of this experiment. The increase of the time needed is due to the presence of hydroxyl(OH) group in the suppository. The higher the amount of PEG 6000 used, the more the OH group that will be present in the suppository. Therefore, there will be more hydroxyl group forming hydrogen bond with paracetamol. With the increase in the formation of hydrogen bond, the suppository formed will be harder and time needed to completely melt the suppository will increase in water. This is because time needed for the dissociation of the hydrogen bond is more difficult with the increase in the formation of the hydrogen bond. From the graph, it can be seen that the average time needed for the suppository with 9g PEG 6000 increases abruptly after the time needed for suppository with 6g PEG 6000

The inaccuracy of the result might due to the inappropriate temperature where higher temperature will increase the melting of the suppository, producing the shorter time needed as measured in for 3g PEG 6000.  Besides, the accidental addition of water from the water bath into the beaker containing suppository will also increase the dissolution of suppository as more water to dissociate the hydrogen bond. A Noyes-Whitney equation of dissolution can explain the theory behind where the additional water will reduce the value of C, concentration of solid in bulk solution.

 is the rate of dissolution.

A is the surface area of the solid.

C is the concentration of the solid in the bulk dissolution medium.

C_s is the concentration of the solid in the diffusion layer surrounding the solid.

D is the diffusion coefficient.

L is the diffusion layer thickness.

With the decrease in the value of C, the value for (C_s – C) will be larger. Hence, the rate of dissolution will be faster, time taken for the suppository to melt will be shorter. Another error that might cause the inaccuracy may be due to the stirring of water when the suppository is melting by using glass rod. Hence, these errors has to avoided in order to improve the accuracy of the experimental result.

3) Plot UV absorption against time graph. Give comments.

The suppository was immersed in a water bath that is maintained at 37^oC to imitate the normal body temperature. The dialysis bag imitates the biological membrane at the rectum.

As time passes, the pattern of average UV absorption increases gradually which tells the amount of paracetamol released from the dialysis bag. The drug released will be absorbed by the UV ray thus giving a reading proportional to the drug content. At the first 5 minutes, the paracetamol release is more rapid as compared to the later stage of the experiment. This may be due to the concentration difference between the inside and outside of dialysis bag, thus allowing a more rapid diffusion of paracetamol out from the dialysis bag.

However, there are two constant reading at minutes 10 to 15, and 20 to 25. This can be due to errors while conducting the sampling. For example, the water bath is not properly stirred before taking the sample eliquot, thus giving lower value if the sample was taken far from the dialysis bag and vice versa. The cuvette may also not cleaned properly, thus giving impuritites that can interfere with the UV absorption.

To have a suppository that melts at body temperature is important to ensure maximum drug diffusion through the rectum lining. However, the formulation must be able to melt only within the body temperature’s range to prevent insufficient drug delivery to the rectum due to excessive melting, leakage or wastage upon application of the dosage form. The suppository also must melt gradually because if it takes too long to melt, it will give unpleasant feeling to the patient and if the suppository melts too quickly, it may cause the liquid to leak out from the rectum.

4) Plot a graph of UV absorption against time for formulation suppository that have different composition. Differentiate and discuss the result.

Theoretically, the graph obtained should obey the sigmoid curve. But, all the graph obtained by us does not obey the sigmoid curve. The results obtained shown on the graph are increased and decrease at several times. The result shows the unusual sigmoid curve. According to the experimental result obtained, Formulation I suppository has the highest peak, followed by Formulation IV, II and III. Beer-Lambert Laws states that the higher the absorbtion of UV, the higher the amount of drug in the solution. This means that Formulation I has the highest drug release rate compared to the other formulations. While the Formulation III has the lowest drug release rate.

Formulation I and IV suppository have higher peak than Formulation II and III. This means Formulation I and IV have higher drug release rate compared to Formulation II and III. Theoretically, the ideal formulation for suppository to have the highest drug release rate is 40% of PEG 1000 and 60% of PEG 6000. So, the ideal formulation used in this experiment should be the formulation III which contain 33.33% of PEG 1000 and 60% of PEG 6000. The higher the percentage the PEG 6000 used, the higher the drug release rate. This is because PEG 6000 will enhance the drug release. But, too high percentage of PEG 6000 will decrease the drug release rate instead of enhance it. Based on the results obtained from experiment, the suppository that shows highest release rate is from Formulation I. This may be due to some experimental errors occur.

Theoretically, formulation I has the lowest drug release rate because the absence of PEG 6000 in the formulation. Although there is PEG 1000 in the formulation, it only has little effect on drug release. PEG 6000 has much more influence on drug release compared to PEG 1000. But the result that we get is contraindicated with the theory.

Although formulation IV has the highest percentage of PEG 6000, it does not show the highest drug release rate. This is because too many PEG 6000 will cause the formation hydrogen bond between PEG and paracetamol. It requires the longer time to reach the highest value of drug release, as the suppository is the hardest. As a result, the drug release rate decreases.  The inaccurate result that we get from the experiment is due to some error happen when conducting the experiment. Example of errors that happen are uneven stirring of the solution, unstable temperature, mistake in the suppository formation and impurities can all lead to the experimental errors.

5) What is the role of each ingredient used in the preparation of this suppository? How PEG 1000 and PEG 6000 affect the physical characteristic of the formulation and drug release of suppository ?

PEG  act as carrier bases, solubilisers and absorption improvers for the drugs. PEG such as PEG 1000 and PEG 6000 that is used in this experiment increases the effective dispersion and delivery of drugs through the rectal route by diffusing out from the PEG as PEG degrades.

Active ingredient in this formulation is Paracetamol. Basically it is used as therapeutic agent in this suppository. It is an anti-pyretic agent, and it is also known as a pain-killer. It is commonly used for the relief of headache and minor pain.

When water diffused in, PEG 1000 will miscible with water evenly where the drug can dissolves in water. Therefore, constant rate of release of drug can be achieved while PEG 6000 has the characteristic of higher viscosity make it less available to miscible with water. This causes the drug to be difficult to diffuse out into the water. As increased of PEG molecular weight resulted in decreased solubility in water and solvent.

CONCLUSION

Different portion of combination of PEG 1000 and PEG 6000 affects the physical characteristics of the suppository and the rate of release of the active ingredient. The lower the molecular weight of the PEG, the higher the hydrophilicity, as it has higher hydroxyl value as compared to higher molecular weight PEG.

REFERENCES

1. http://pharmlabs.unc.edu/labs/suppository/bases.htm

2. http://www.pharmacopeia.cn/v29240/usp29nf24s0_c1151s68.html

Experiment 3 – Emulsion

INTRODUCTION

An emulsion is a type of dispersion in which two normally immiscible substances are stabilised by another substance, called an emulsifier. For example, olive oil and water will not dissolve in one another, as their intermolecular forces differ (like dissolves like). While we can agitate to form a suspension, it is temporary and the oil and water will eventually separate into distinct layers. There are two types of emulsion which are oil/water emulsion and water/oil emulsion. In emulsion preparation, a series of trials are performed to select a proper surfactant system.  Each surfactant has a specific HLB (hydrophile-lipophile balance) number.  If the required HLB for an oil is known, select surfactants or surfactant blends with HLB numbers close to the required HLB number.  A number of different surfactants with the required HLB number should be examined.  Not all surfactants having the same HLB value will be acceptable. When the HLB value for the ingredient(s) to be emulsified is unknown, the following procedure can be used.  Two surfactants with different HLB values are selected — usually one is oil soluble, the other is water soluble.  Blends of these surfactants in different proportions are prepared.  If an emulsion appears to be stable, then this system should be optimized and tested for long-term storage stability.

OBJECTIVE

1. To determine the effect of HLB surfactant on the emulsion stability.
2. To study the effect on physical and stability of the emulsion when the different amount of emulsifying agent have been used.

APPARATUS AND MATERIALS

8 tests tube

50ml measuringcylinder

2 sets pasture pipette and dropper

Vortex mixture

Weighing boats

1 set mortar & pestle

Light Microscope

Microscope slides

A set of 5ml pipette and bulb

A 50mL beaker

A 15ml centrifugation tube

Coulter counter apparatus

Centrifugation apparatus

Viscometer

Water bath 45^oC

Refrigerator 4°

Mineral oil

Arachis oil

Olive oil

Distilled water

Span 20

Tween 80

Sudan III solution

PROCEDURE

1. Every test tube is labelled and a straight line of 1cm is drawn from the bottom of the test tube.
2. 4ml of oil (mineral oil) and 4ml of distilled water are mixed in a test tube.
3. Span 20 (6 drops) and Tween 80 (9 drops) are dropped into the mixture of mineral oil and distilled water. The test tube is closed and mixed with Vortex Mixer for 45 seconds. Time taken is recorded for the interface to reach the 1cm line. The HLB value is determined for every sample.
4. A few drops of Sudan III are added to 1g of emulsion. The separation of colours in the sample are described and compared. An amount of sample is transferred onto a slide and observed under microscope. The shape and size of globule produced are hed, described and compared.
5. A formulation of Mineral Oil Emulsion is formulate using the wet gum method as below :

Mineral oil	25ml
Acacia gum	6.25g
Syrup	5ml
Vanillin	2g
Alcohol	3ml
Distilled water,qs	50ml

6. 40g of the emulsion produced is placed into a 50ml beaker and homogenized for 2 minutes using a homogenizer.
7. 2g of the homogenized and non-homogenized emulsion are weighed and labelled. A few drops of Sudan III are added. The texture, consistency, oiliness, and separation of colours of sample are observed and compare under microscope.
8. The viscosity of homogenized emulsion (15g in a 50ml beaker) are determined using a viscometer calibrated with Type LV-4 “Spindle”. The sample is exposed to water bath, 45°C for 30 minutes and refrigerator,  4°C for 30 minutes. The viscosities are determined after completing the cycle and left to be cooled to room temperature for 10-15 minutes.
9. 5g of homogenized emulsion is placed into a centrifuge tube (4500rpm, 10 minutes, 25°C). The ratio of height separation is measured.

QUESTIONS

1) What is the HLB value that will produce good and stable emulsion? Discuss.

Physically stable emulsions are the best achieved by the presence of a condensed layer of emulgent at the oil/water interface and that the complex interfacial films formed by a blend of an oil-soluble emulsifying agent with a water-soluble one produce the most satisfactory emulsions. Hydrophile-lipophile balance (HLB) method is used to calculate the relative quantities of the emulgents necessary to produce the most physically stable emulsion for oil/water combination.

Each surfactant is allocated an HLB number representing the relative proportions of the lipophilic and hydrophilic parts of the molecules.  High numbers up to a theoretical maximum of 20 indicate a surfactant exhibiting mainly hydrophilic or polar properties whereas low numbers represent lipophilic or non-polar characteristics.

In this experiment, a series of trial emulsions used to assess for stability. It based on fact that the degree of creaming or separation is at minimum at the optimum HLB value. Our result shows that test tube 1, 2 and 4 does not show any separation until reach 1cm. All stop at 0.5cm, 0.7cm, and 0.3cm. Therefore, it might be not suitable HLB value since the emulsion also already become caking and condensed on the top.

A stable emulsion may be defined as a system in which the globules retains their initial character and remains uniformly distributed throughout the continuous phase.  Therefore, the most suitable HLB value is 11.3, 13.2 and 14.1.

2) Compare the physical appearance of the Mineral Oil emulsions formed and give comments. What is Sudan III Test? Compare the colour dispersion produced explain. For Sudan III Test in procedure 4: Based on the experiment, our group had conducted the experiment for 25ml of mineral oil. Note that before the homogenization, the emulsion is unstable because the oily phase is immiscible with aqueous phase. Under the microscope, the globules appear in different sizes and larger. They are all unevenly dispersed. But after the homogenization process, we can denote that the emulsion become more stable as it become less oily and more viscosity. When it is observed under microscope, the globules are in smaller size and more evenly distributed throughout the emulsion.

For Sudan III test in procedure 7: (25 mL of Mineral oil) 3) a. Plot a graph of sample emulsion viscosity before and after the temperature cycle against different amount of mineral oil. All type of emulsion is exposed to the same temperature. The emulsions all have the same volume, but different proportion of oil, water and emulsifying agent proportions. From the graph, it shows that the viscosity of emulsion before the temperature cycle increases with the increasing amount of mineral oil like palm oil, arachis oil and olive oil, except for mineral oil, which shows a decrease in viscosity with increasing amount of mineral oil. The increasing proportion of mineral oil also indicates the increasing amount of acacia as emulsifying agent from the ratio of 4:2:1. The ratio indicates 4 parts of oil, 2 parts of water and 1 part of acacia. The increase amount of acacia will reduce the surface tension and increases the viscosity of the emulsion, so that the emulsion will not easily separate into two layers. For amount of mineral oil of 25 and 30 mL, the graph shows the decreasing in the viscosity of the emulsion. The viscosity of emulsion after the temperature cycle increases with increasing amount of mineral oil for both olive and mineral oil. Whereas the viscosity of emulsion after the temperature cycle decreases with increasing amount of mineral oil for arachis oil, palm oil has the same value of viscosity regardless the amount of mineral oil. This may be caused by loss of some proportion of volume of emulsion during the heating process.  Some proportion of water loss in heating and cause the emulsifying agent becomes less soluble in the continuous phase, thus it cannot increase the viscosity. During the heating process too, the molecules vibrate more, but interact less. That’s why the viscosity of emulsion reduced. These inaccuracy of results also may be due to some errors that occur during the experiment.  Example of error is may be due to the insufficient or excess emulsion proportions and also error in the manufacturing process of emulsion. This is because increasing amount of mineral oil used indicates the increasing amount of emulsifying agent, thus increases the viscosity of the emulsion.  Due to temperature cycle, increasing amount of mineral oil used will cause some loss of water. Then, it will form water-in-oil emulsion. The emulsifying agent becomes less soluble in the continuous phase, so it reduces the viscosity of emulsion after the temperature cycle.

4) Plot a graph of separated phase ratio formed from the centrifugation process versus the amount of mineral oil used. Explain.

Sudan III is lysochrome (fat soluble dye) it is a staining agent used to determine the presence and location of oily solution by staining it red. Sudan III will stain non polar substances including oils, fats, waxes and greases due to its higher affinity towards these compounds as compared to aqueous phase. It is usually being added to o/w emulsion, in background will be colorless with colored spots if dispersed phase. Note that this test may fail if ionic emulsifiers are used in the preparation. Sudan III testing shows the shape and physical characteristic of oily phase in an emulsion by staining the oil phase and observed under light microscope. We can also determine the type of the emulsion whether water in oil (w/o) or oil in water emulsion (o/w) by using this test. Red environment with globules indicate a w/o emulsion whereas red globules in a clear field indicate an o/w emulsion. Phase separation ratio is used to indicate stability of an emulsion. A high ratio of phase separation represents an unstable emulsion where two separated phases can be observed. Due to unstable emulsion, uniformity of drug in the emulsion will be greatly altered and the accuracy of dose being administered into patient might affected. Based on the graph plotted, separated phase ratio increases from 20mL palm oil emulsion to 25mL arachis oil emulsion, decreases from 25mL arachis oil emulsion to 30mL olive oil emulsion. According to theory, the separation phase ratio should increase with the increasing of the mineral oil contained in the formulation. This is because the added amount of oil in emulsion has exceeded the oil amount at which a stable emulsion can be formed.  Phase separation will occur at a faster rate. However, the results we obtained from the experiment do not follow this theory. This may be due to several errors that occurred during experiment. This may due to some experimental errors such as measurement mistakes and systemic error. The experimental errors include inaccuracy in measuring the amount of oil before forming the emulsion, insufficient homogenisation that has been carried out on emulsion and the height of separated phase is not being measured accurately.Thus, an appropriate amount of oil should be used in the emulsion to prevent phase separation as the amount of oil affects the uniformity and stability of the dose of drug administered. The drug particles should be dispersed in the emulsion uniformly in an ideal emulsion.

5) What are the function of each ingredients that used in the preparation of an emulsion? How the used of these ingredients effect the pyhysically and stability of formulation of an emulsion?

Palm oil, arachis oil, olive oil and mineral oil act as oil phase (internal phase) in the oil in water emulsion (o/w emulsion). Acacia, Span 20 and Tween 80 are emulsifying agents used to emulsify two immiscible liquid which are liquid and oil into a miscible form called emulsion. There have two region which hydrophobic tail and hydrophilic head that will in contact with the aqueous phase. This will lower the surface tension of water molecule and provide an evenly mixing between oil and water molecule. Vanillin acts as flavouring agent which can increase the taste of emulsion to increase the compliance of patient as an emulsion always contains a taste not preferred by most of the patients. Alcohol is used as preservative to prevent growth of microorganism since distilled water is used as continuos phase in oil in water emulsion. Syrup is used to mask the unpleasant taste of mineral oil and it is also can  increase the viscosity of the emulsion  but its amount need to be controlled to prevent to viscous. As the level of the internal phase volume increased and consequently the amount of water decreased, emulsion viscosity increased. The consistency of emulsions can be increased by increasing the viscousity of the continuos phase , and also increasing proportion of emulsifying agents into it. Emulsions prepared by homogenization at the beginning of the process of emulsification were stable with small internal phase particle diameter. Uses of alcohol ratio there is a phase volume/emulsifier HLB ratio, which results in optimum stability.

CONCLUSION

Emulsifying agents plays a role to the formation of a stable emulsion. By using combination of Tween 20 and Span 80, a more stable emulsion is formed as compared to when using either one  with the total absence of emulsifying agent. Different combination of hydrophilic and lipophilic emulsifier will give different HLB values which will determine the extent of the emulsion’s stability.

REFERENCES

1. http://deltauniv.edu.eg/UploadFiles/UploadFiles/3%281%29%20physical%20pharmacy_Hydrophilic-Lipophilic%20Balance.pdf

2. https://books.google.com.my/books?id=wYfODY0PPfYC&pg=PA135&lpg=PA135&dq=sudan+iii+test+in+emulsion&source=bl&ots=kNL6wSwi7y&sig=TqMDsuZChLNleFRpnY7IT6sVX0s&hl=en&sa=X&ei=rOprVeuEF4eXuASq5oKwCQ&sqi=2&redir_esc=y#v=onepage&q=sudan%20iii%20test%20in%20emulsion&f=false

Experiment 2 – Suspension

INTRODUCTION

A suspension formulation is a type of disperse system where the insoluble solid particles are dispersed homogenously in the liquid phase. A good suspension must be in a homogenous form after shaking, easily poured from the container, has a uniform solid particle size and possess better feel and taste. Sediment that is formed upon storage has to be easily redispersed after shaking, forming homogenous suspension.

In general, suspension comprises of an active ingredient (solid phase) in a liquid carrier, a wetting agent, flavouring and colouring agents and preservatives. The function of a wetting agent such as tragacanth is to reduce the interfacial tension between the solid particles and the liquid. Suspension can be classified into a coarse suspension where the diameter of the particles is more than 1 µm or a colloidal suspension where the particle size is less than 1 µm. In pharmaceutical practice, suspension is used to improve the stability of the active ingredient, taste, and bioavailability.

OBJECTIVES

To study the effect of the composition of tragacanth on the physical appearance, the rate of sedimentation and viscosity of the suspensions of different compositions.

APPARATUS & MATERIALS

1. Weighing balance
2. Viscometer
3. Centrifuge
4. 100ml Beaker
5. Weighing boat
6. Pestle and mortar
7. 50ml and 200ml Measuring cylinder
8. Glass rod
9. 150ml Plastic bottle
10. 1ml Pipette and bulb
11. 15ml Centrifudge tube
12. Ruler
13. Chalk
14. Tragacanth
15. Concentrated peppermint water
16. Syrup BP
17. Double-strength chloroform water
18. Distilled water

PROCEDURE

1. A suspension of Paediatric Chalk Mixture (150ml) is prepared using the formulation below:

Chalk                                                                                       3g Tragacanth                                                                    Refer to table 1 Concentrated Peppermint Water                                           0.6mL Syrup BP                                                                               15mL Double Strength Chloroform Water                                      75mL Distilled Water, q.s.                                                              150mL

Table 1

Paediatric Chalk Mixture	Group	Tragacanth (g)
I	1, 5, 9	0.0
II	2, 6, 10	0.1
III	3, 7, 11	0.3
IV	4, 8, 12	0.5

2. 5 ml of the suspension that had been prepared was transferred into a weighing boat and labelled. The texture,clarity and colour was compared and explained.
3. Then, 50 ml of the suspension that had been prepared was transferred into 50 ml measuring cylinder and the height of sedimented solid in the cylinder was measured every 0,5,10,15,20,25,30,35,40,45,50,55, and 60 minutes.
   
4. 95 ml of the remaining suspension was poured inton100ml beaker and the viscosity of the suspension was determined by using Viscometer.
   
5. 10 ml of the suspension was poured into centrifuge tube and the height of solid phase after centrifuged was measured (1000 rpm, 5 minute, 25 0C).

QUESTION

1. Compare the physical characteristics of the suspensions and give your comment.

Group	Amount of Tragacanth (g)	Texture	Clarity, appearance	Colour of suspension
1	0.0	Less viscous, rough texture and disperse easily	Less cloudy, has 2 layer	Less milky
2	0.1	Viscous	Moderately cloudy	Milky
3	0.3	Viscous	Opaque, cloudy	Milky
4	0.5	More viscous	Opaque, cloudy	Milky

Tracaganth acts as a suspending agent to ensure the ingredients are even and proportional in the mixture to support the suspensoid. As the amount of tragacanth added to the formulation is increased, the suspension becomes more viscous. The more viscous a suspension is, the longer time it requires for the suspended particles to sediment and this allow the product to be poured out easily from the vehicle without particles clinging at the bottom. As the amount of tragacanth that added to the formulation increases, the more opaque and cloudy the product will be. The opacity and the cloudiness indicates that the disperse particles is totally dispersed in the medium. If no tragacanth is added to the formulation, the product will appear to be separated in two phase in which the supposed to be disperse phase seems to sediment at the bottom of the container. . But if sedimentation occurs, it will be easily redisperse and stay disperse in the medium for a longer time compared to those formulation without tragacanth.

 

2. Plot graph of sediment against time. Give comments

Time (minutes)	0	5	10	15	20	25	30	35	40	45	50	55	60
Sediment height (mm)	0	1	4	8	10	15	18	21	22	22	22	22	22

 

The graph above shows the relationship between the heights of sedimentation (mm) against time (min). Based on the graph, as the time taken increases, the height of sedimentation increases as well. The suspension was white in colour. Some light are used to observe the sedimentation clearly to avoid from making error. For the last 20 minutes, the height of sedimentation of suspension has become no change which is it maintain at 22 mm. This is because all of the Tragacanth in the suspension has been sedimentated.

 

3. Plot average sediment height against time for suspensions with different Tragacanth content. Give comments

Time(min)		Average Sediment Height (mm)
		0	5	10	15	20	25	30	35	40	45	50	55	60
Tragacanth Content (g)	0.0	95	18	12	12	12	12	12	12	12	12	12	12	12
	0.1	12	11.8	11.6	11.5	11.5	11.5	11.4	11.3	11.3	11.3	11.2	11.1	10.9
	0.3	0	9	11	11	12	13	13	14	14	14	14	14	14
	0.5	0	1	4	8	10	15	18	20	21	22	22	22	22

 

From the result, a comparison can be made based on the time needed for the first sediment to settle. The higher the amount of tragacanth mixed, the longer the chalk can suspend, thus needs longer time to sediment. For 0.0g and 0.1 tragacanth, the absence of sufficient suspending agents causes the suspension to be very unstable, thus enables sedimentation to occur rapidly as soon as the mixing stopped. At highest tragacanth 0.5g concentration, no sedimentation can be seen on the first few minutes, because the suspending agents allows the chalk particles to suspend longer.

Next, we can see an obvious difference in the sedimentation pattern in low and high concentration of tragacanth. At 0.0g, the sediment height become shorter as time passes before comes to a plateau and this pattern is inversely true for higher concentration (0.5g). This is due to the presence of tragacanth which makes the sediment is less packed, as compared to suspension with lower tragacanth concentration.

4. Elaborate the viscometer analysis mechanism. Plot a graph on viscosity of suspension against Tragacanth content (g). Give comments

Viscometer is an instrument for the determination of viscosity. The most common types of viscometer are capillary, rotary, falling-sphere and ultrasonic.

In this experiment we use rotary viscometer. Rotational viscometers use the idea that the torque required to turn an object in a fluid is a function of the viscosity of that fluid. They measure the torque required to rotate a disk or bob in a fluid at a known speed.

‘Cup and bob’ viscometers work by defining the exact volume of a sample which is to be sheared within a test cell. The torque required to achieve a certain rotational speed is measured and plotted. There are two classical geometries in “cup and bob” viscometers, known as either the “Couette” or “Searle” systems. It is distinguished by whether the cup or bob rotates. The rotating cup is preferred in some cases because it reduces the onset of Taylor vortices, but is more difficult to measure accurately in instrument.

‘Cone and Plate’ viscometers use a cone of very shallow angle in bare contact with a flat plate. With this system the shear rate beneath the plate is constant to a modest degree of precision and deconvolution of a flow curve.

The advantages of this rotational viscometer are auto-zero function to ensure precision torque measurement, select all functions from user-friendly keypad, time stop feature to measure viscosity at precise user specified time interval, time to torque feature to measure the time interval for sample to reach user defined torque value, senses and displays continuously viscosity (cP or mPa·s), temperature (°C or °F), % torque, speed (rpm) and spindle used.

Tragacanth (g)	0.0	0.1	0.3	0.5
Viscosity (cP) ( x ± SD)	2.72 ± 0.98	2.45±0.48	2.92± 0.88	2.98 ± 0.46

 

6. Plot a graph of height of sedimentation formed after centrifuge against the composition of tragacanth(g). Give your explanations.

Tragacanth (g)	Before centrifuge (mm)	After  centrifuge(mm)	Height ratio
0.0	240	60	4.00
0.1	50	140	0.35
0.3	79	14	5.64
0.5	20	180	0.11

For this experiment, centrifuges Is used to investigate the effect of different amount of tragacanth on the height separation produced from the suspension after undergoes centrifugation at 1000 rotation per minute for 5 minutes within 25ºC. Based on centrifugation theory, centrifuge helps to separate suspended material from the mediums they are mixed with.  Particles of higher density or larger size typically travel at a faster rate and at some point will be separated from particles less dense or smaller. This can be explained by Stokes equation. Sedimentation rate will decrease as the medium viscosity increases, whereas increases as the gravitational force. Theoretically, theory, the result would be decreased in the ratio separation as the amount of tragacanth used increase. This is because tragacanth acts as a suspending agent, in which it imparts viscosity to the solution, besides forming film around particles and decrease interparticle attraction. It also acts as thickening agent which is important in the increase of viscosity of the solution that helps in preventing sedimentation of the suspended particles.  Therefore, there might be some errors during preparation that lead to increasing in height ratio which conducted by group 3

7. What is the function of each material that is used in the suspension formulation? How is the use of different amount of Tragacanth affect the physical characteristic and stability of a suspension formulation?

Chalk is an adsorbent and antacid and is an ingredient of mixture and powders used in the treatment of diarrhea. Tragacanth forms viscous solutions or gels with water, depending on the concentration. It is used as a suspending agent. Concentrated peppermint water is a carminative. It has mildly antiseptic properties. Double strength chloroform water acts as carminative, flavouring agent and preservative. Syrup BP is a sweetening and flavoring agent. Varied amount of Tragacanth will affect the physical characteristic of the suspension either it will be rough or smooth. The stability of suspension which use Tragacanth is good and but on prolonged storage is hard to redisperse. Suspensions that do not use Tragacanth do not have good stability and the solid particles inside will sediment rapidly but easy to redisperse.

CONCLUSION

We can evaluate the effect when using varied amount of Tragacanth and this will influence the physical characteristics and stability of the suspension.

 REFERENCES

1. http://en.wikipedia.org/wiki/Viscometer#Rotational_viscometers
2. http://www.brookfieldengineering.com/products/viscometers/laboratory-dv-i.asp

 

Experiment 1- Ointment

INTRODUCTION

Ointment is a semi-solid containing active ingredient and other excipient. The active ingredient may be one or more. It is intended to be used topically onto our skin and provide an emollient effect. It is advisable to consult a patient to use the ointment preparation after bath and during the skin still wet. This is because the ointment will form a thin layer coating the skin surface preventing water loss from skin. A good ointment preparation must have a good texture, easily to applied and not too greasy.

OBJECTIVE

1. To investigate physical characteristics by using different composition of ointments.
2. To find out rate of drug being released by using different composition of ointments.

APPARATUS & MATERIALS

1. Weighing balance
2. Heater
3. Spectrometer
4. 100ml Beaker
5. Weighing boat
6. Pestle and mortar
7. Glass slap
8. Glass rod
9. Spatula
10. 10cm Dialysis beg
11. 5ml Pipette and bulb
12. Plastic cuvette
13. Thread
14. Emulsifying wax
15. White soft paraffin
16. Liquid paraffin
17. 5g Acetylsalicylic acid
18. Distilled water

PROCEDURES

1. Emulsifying Ointment (50 g) was prepared following this formula:

Emulsifying ointment	Group	Ingredients			Total (g)
		Emulsifying wax	White soft paraffin	Liquid paraffin
I	1, 5	21	25	4	50
II	2, 6	17	25	8	50
III	3, 7	13	25	12	50
IV	4, 8	9	25	16	50

2. Some of the ointment formed (5 g) was taken and placed into a weighing boat and was labeled. The texture, clarity and color of the ointment formed was described and compared.
3. The acetylsalicylic acid powder was incorporated (1.5 g) into 30 g ointment which has been prepared using the levigation technique. The acetylsalicylic acid was triturated using the mortar and pestle.
   
4. Fill the acetylsalicylate acid ointment into the dialysis bag and both of the end of the beg were tied tightly, as shown in the figure below:
5. The bag was inserted into a beaker (100 ml) containing distilled water (50 ml) which has been heated to 37⁰C.
6. In the interval of 5 minutes, an aliquot sample (2-5 ml) was pipette and the acetylsalicylic acid released from the ointment base was determined using the UV-visible spectrometer. The distilled water was ensured to be stirred before the sample was taken.

RESULTS

Time (minutes)	UV Absorption
UV Absorption at 300 nm	0	5	10	15	20	25	30
	0.128	0.137	0.162	0.173	0.255	0.476	0.522

 

DISCUSSION

1. Compare the physical characteristic for the ointments and explain.

Experiment	Material (g)			Texture of ointment			Clarity
	Emulsifying Wax	White soft paraffin	Liquid paraffin	Spreadibility	Greasiness	Hardness
I.	21	25	4	Difficult	Least greasy	Hardest	Turbid
II.	17	25	8	Little difficult	Little greasy	Less harder	Turbid
III.	13	25	12	Less difficult	More greasy	Little soft	Turbid
IV.	9	25	16	Easy	Most greasy	Softest	Turbid

 

2. Plot a graph of UV absorption against time and give explanation.

 

The UV spectrometer measures the releasing of acetylsalicylic acid from the ointment in the dialysis bag to the distilled water. The release of the drugs from the ointment involved the diffusion mechanism. The water is set to 37 ⁰C to mimic the temperature of human body as ointment will be applied to human skin. Meanwhile, the dialysis bag represents the skin membrane.

The amount of drug absorbed into the blood circulation is represented by the concentrations of the drug in the distilled water. The UV absorption represents the concentration of the drug that crosses the membrane and reaches into  the distilled water. In theory, the UV absorption is proportional to the time of the release of acetylsalicyclic acid across the membrane to distilled water. The gradient of the graph shows the rate of drug release across the membrane of dialysis bag.

The graph shows that as the time increases, the concentration of the acetylsalicyclic acid in the distilled water is increasing. Along the time until a particular point, the gradient of the graph decreases thus the releasing rate is reduced. Along the progression of the experiment, the concentration of salicyclic acid in the dialysis bag and the distilled water become isotonic because the salicyclic acid keeps moving into the distilled water and equilibrium occurs.

This will result in the reduced gradient of the graph when time passes by. If the experiment is continued beyond 60 minutes, it may result in a straight line graph as the acetylsalicyclic acid diffuses into and out of the membrane of dialysis bag in an equilibrium state. At the late stage, the releasing rate increases. This may be contributed by the experimental error such as leakage of the drug whereby the thread is not tight tightly enough or  the distilled water is stirred too vigorously.

 

3. Plot a graph of UV Absorption against time for ointment formulations of different composition. Compare and discuss the results obtained.

Time (min)	UV Adsorption
	0	5	10	15	20	25	30
UV absorption at 300nm	0.128	0.137	0.162	0.173	0.255	0.476	0.522

 

Time (min)		Average UV absorption at 300nm (x ± SD)
Emulsifying ointment		0	5	10	15	20	25	30
	I	0.049	0.060	0.370	0.280	0.050	0.315	0.031
		0.075	0.093	0.257	0.301	0.334	0.428	0.443
	AVG	0.062	0.0765	0.3135	0.2905	0.1920	0.743	0.237
	SD	0.0183	0.0233	0.0798	0.01485	0.2008	0.5314	0.2913
	II	0.202	0.205	0.194	0.239	0.288	0.318	0.337
		0.360	0.390	0.413	0.415	0.498	0.525	0.544
	AVG	0.281	0.2975	0.3035	0.327	0.393	0.4215	0.4405
	SD	0.1117	0.1308	0.1549	0.1245	0.1485	0.1464	0.1464
	III	0.070	0.088	0.076	0.166	0.153	0.087	0.163
		0.004	0.024	0.034	0.081	0.078	0.089	0.127
	AVG	0.037	0.056	0.055	0.124	0.116	0.088	0.145
	SD	0.0467	0.0453	0.297	0.0601	0.0530	0.0014	0.0255
	IV	0.128	0.137	0.162	0.173	0.255	0.476	0.522
		0.114	0.335	0.329	0.350	0.192	0.281	0.655
	AVG	0.121	0.236	0.246	0.262	0.224	0.379	0.589
	SD	0.009	0.140	0.118	0.125	0.045	0.138	0.094

 

The ideal ratio of amount Emulsifying Wax, White Soft Paraffin and Liquid Paraffin in the preparation ointment is 30 : 50 : 20. Theoretically, an ointment that contains the highest amount of emulsifying wax and lowest amount of liquid paraffin will take longer time for the acetylsalicylic acid to penetrate from the dialysis membrane. This results in the lowest reading of UV absorption by the acetylsalicylic acid. This can be explained by the theory stated that when there is a low amount of emulsifying wax,  the acetylsalicylic acid cannot disperse well in the ointment. As we increase the amount liquid paraffin in the formulation, greater amount of acetylsalicylic acid is able to penetrate the dialysis membrane at a faster rate. This is due to the role of liquid paraffin in the formulation that improves hydration, thus increase the effectiveness of the absorption of the ointment at the percutaneous membrane.

In the experiment conducted, four formulations were prepared. The formulations possessed different amount of emulsifying wax and liquid paraffin. Results obtained have shown that Formulation 1 which contained the least amount of liquid paraffin resulted in the lowest UV absorption value. Formulation 3 showed higher UV absorption than Formulation 1. This is followed by Formulation 4 and Formulation 2 with the greatest amount of UV absorption. As the amount of emulsifying wax reduces, with an increase in the amount of liquid paraffin, the UV absorption of acetylsalicylic acid should increase. From the results obtained, there is a slight inaccuracy because supposedly Formulation 2 shows lower UV absorption than Formulation 3 and Formulation 4. This inaccuracy might be due to several possible errors done during the experiment.

The possible errors may be due to the inaccurate measurement of amount of ingredients used in the formulation prepared. This will affect the permeation rate of Acetylsalicylic acid and the UV absorption. Secondly, the absorption of UV may get affected by the inaccurate or different amount of ointment filled in the dialysis bag. Besides, the presence of contaminants on the dialysis bag could also contribute to the inconsistent results. Moreover, the spillage of the drug from the dialysis could be one of the errors that could alter the results.

4. what is the function of each ingredients used in the preparation of the ointment? How does the usage of different compositions of the emulsifying wax and liquid paraffin affects the physical properties of the ointment and the rate of drug release from it?

Emulsifying wax Is a type of anionic emulsifying agent which act as needed to mix two phases, (oil-soluble and water-soluble)phases and forms a layer known as interface. It helps in decreasing the interfacial surface tension to enable the drug particles to be distributed evenly in the ointment which prevents  sedimentation from occurring.

White soft paraffin and liquid paraffin are hydrocarbon base. White soft paraffin also known as white petroleum  jelly. It act as moisturizer and emollient by providing a layer of oil on the surface of the skin to prevent water  evaporating  from the skin surface. It increases the greasiness of the ointment and is able to penetrate through the hydrophobic layer of the skin more readily. It also serve as stabilizer. Liquid paraffin is used to reduce the viscosity of the base so that the emollient will easily expelled from its container. Combination of white soft paraffin and liquid paraffin is to achieve viscosity.

Acetylsalicylic  acid is the active ingredient in this ointment preparation and play a major role as the active ingredient. It also known as aspirin that give therapeutic effect by given antipyretic, anti-inflammatory and analgesic effect by enzyme inhibitor. The grainy of texture of dissolved aspirin also serves to exfoliate the skin.

The composition of each bases in ointment very important to determine characteristic of ointment produce. Emulsifying wax will give hardness to the ointment due to its high melting point. Then, liquid paraffin can give softness to the ointment if large amount added. Lower portion of emulsifying wax and large portion of liquid paraffin will increase spreadibility and greasiness and enhance the release of drug and help in penetrate the skin lipid bilayer readily.

CONCLUSION

The proportion of emulsifying wax and liquid paraffin should be in balance in order to produce an ointment with desired hardness. This can ensure that the active ingredients can be released at an optimum rate and increase the bioavailability.