EXPERIMENT 4: EVALUATION OF THE EFFECTS OF DIFFERENT FORMULATIONS ON SUPPOSITORIES CHARACTERISTICS

Objectives:

i) To determine the effect of different composition of base on the physical characteristics of suppository.

ii) To determine the effects of different formulations on the drug release from suppository.

Date of experiment: 20/5/2015

Introduction:

Suppositories are solid dosage forms intended for insertion into body orifices (rectum, vagina, urethra where they melt, soften, or dissolve and exert a local or systemic effect. Rectal suppositories intended for localized action are most frequently used to relieve constipation or pain, irritation, itching, and inflammation associated with hemorrhoids.

An ideal suppository base should have the following properties:

i)     Melts at body temperature or dissolves in body fluids.
ii)    Non-toxic and non-irritant.
iii)   Compatible with any medicament.
iv)   Releases any medicament readily.
v)    Easily moulded and removed from the mould.
vi)   Stable to heating above the melting point.
vii)  Easy to handle. 

viii) Stable on storage.

Drugs should be dispersed in a suitable suppository base. The base must be capable of melting, softening, or dissolving to release its drug components for absorption. Different base composition would affect the rate and limit of the drug released from the suppository.

The rate at which a drug is released from a suppository and absorbed by the rectal mucous membrane is directly related to its solubility in the vehicle. When the drugs are highly soluble in the vehicle, the tendency to leave the vehicle will be small and so the release rate into the rectal fluid will be low.

The goal of this experiment is mainly to investigate the effect of different base composition on the physical characteristics of suppository formed and the effect of drug release from the suppository.

Apparatus:

Weighing balance, weighing boat, spatula, 50 ml beaker, 100 ml beaker, hotplate, 5ml measuring cylinder, suppository mould, water bath (37^oC) , dialysis bag (10cm), thread, glass rod, 5 ml pipette and pipette bulb, Kuvets plastics, spectrophotometer UV/Vis

Materials:

Polyethylene glycol (PEG) 1000
Polyethylene glycol (PEG) 6000 
Paracetamol
Distilled water

Procedure:

1. A saturated paracetamol stock solution is prepared by adding 10g Paracetamol in 5 ml of  distilled water.

2. The 10g Paracetamol suppository  is prepared by using the following formula:

Suppository	Group	Ingredient (g)		Paracetamol stock solution (g)	Total (g)
		PEG 1000	PEG 6000
I	1, 5	9	0	1	10
II	2, 6	6	3	1	10
III	3, 7	3	6	1	10
IV	4,8	0	9	1	10

3. The suppository is shaped  by using  suppository-mould . The shape, texture and colour of the suppository formed is observed and compared.

4. The suppository is inserted into a beaker which contain 10ml distilled water (37^oC) and the time that is needed to melt the suppository is recorded.

5. A suppository is inserted into a dialysis bag and both ends are tightly tied with thread. The bag is inserted into a 100ml beaker which contains 50ml distilled water that has been heated to 37^oC.

6. At each 5 minutes interval, an aliquot sample(3-4ml) is pipetted and the release of Paracetamol from the suppository is determined by using UV-visible spectrometer. The distilled water is stirred by using a glass rod before the sample is taken.

Results:

Time (min)	UV-visible absorption at 520 nm
	0	5	10	15	20	25	30
Group 2	0.0200	0.0250	0.0310	0.0340	0.0340	0.0360	0.0360

 Discussion:

1. Compare the physical appearance of the suppository formed.

Physical characteristic	Suppositories
	I 1, 5	II 2, 6	III 3, 7	IV 4, 8
Shape	Bullet shaped, solid	Bullet shaped, solid	Bullet shaped, solid	Bullet shaped, solid
Hardness	+	++	+++	++++
Greasiness	++++	+++	++	+
Colour	White	White	White	White

The entire suppositories produced are of bullet shaped, but the time taken for them to be solidified varies. This is due to the different composition of PEG1000 and PEG6000 in the suppositories. Formulation with higher content of PEG1000 will took longer to be solidified.

PEG 1000 is greasier and less hydrophilic than PEG6000 which possess more hydroxyl group. Thus the greasiness of the suppositories increased with the increase of the amount of PEG1000.

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Besides, the hardness characteristic of the suppository is increasing from formulation I to formulation IV with the increasing amount of PEG 6000.

                                                                                                            

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

Content PEG 6000 (g)	0		3		6		9
Time (min)	Group 1	65	Group 2	10	Group 3	60	Group 4	74
	Group 5	58	Group 6	65	Group 7	43	Group 8	61
Average time ± SD		61.5 ± 4.95		37.5  ± 38.89		51.5 ±12.02		67.5 ±9.19

Based on the graph shown, we can observe that the time taken for the suppository to melt is not directly proportional to the PEG 6000 content in gram. The function of the PEG 6000 content is as a suppository base. As the general knowledge, increasing the mass of the PEG 6000 will make the suppository more solid. For that, the time taken for the suppository to melt will be longer as the mass of the PEG 6000 increase. As shown on the graph above, the time taken for the suppositories to melt which contain 9g of PEG 6000 is 67.5 minutes which the longest time among the result. While the shortest time taken for the suppository to melt is the suppository that contain 3 grams and 6 grams of PEG 6000.

Theoretically, the time taken for the lowest amount of PEG 6000 should be lowest and vice versa but the results we obtained is totally inappropriate. The deviation of the result from the story is majorly effected by the errors occur while conduct the experiment. Defect of suppository made reduction in mass and will reduce the time for suppository to dissolve. Error made during measured, and transferred of the ingredient during making of suppository also may alter the result. There also possibility that suppository does not solid enough when we removed it from the mould. The unsolidified suppository made it easier to be dissolved in water bath. The heating process using water bath also may produce this result. Some of the group might stir the beaker containing suppository which make it faster to dissolve.    

3. Plot the graph of UV absorption against time (Step 6). Explain the plotted graph. 

Initially, there is small amount of paracetamol detected at 0 time. This is an error that occurs due to inappropriate filling of paracetamol into the dialysis bag.

In this experiment, paracetamol needs to pass through the dialysis bag before it can reach distilled water. Dialysis bag represents human biological membrane while the distilled water represents human blood plasma. The melting process,drug release and drug absorption processes occur in water bath at 37ºC, which represents human body temperature.

At the first 10 minutes, amount of paracetamol released into water in beaker shoots up. This is because there is a great difference in concentration gradient between water in beaker and suppository in dialysis bag. Therefore, paracetamol diffuses out quickly.

This graph deviates from the theory (sigmoid curve). Firstly, uneven heating of water bath which will lead to inconstant drug release rate from the suppository. Besides that, the suppository we made may not be homogenously formed. This may cause the brittleness of PEG suppository or trapped air space in the suppository thus reducing the size and altering the drug release rate of drug from suppository dosage form. In addition to that, distilled water in which the dialysis bag is exposed to may not be stirred evenly before it is taken to be tested on spectrophotometer UV/Vis.

4. Plot the graph of UV absorption versus time for the suppository formulation which has different composition. Compare and discuss the result.

	Group/ Time (min)	UV absorption at 520 nm (X ± SD)
		0	5	10	15	20	25	30
S U P P O S I T O R Y	1	0.0130	0.1970	0.2870	0.2080	0.2110	0.2260	0.2970
	5	0.0013	0.0045	0.0074	0.0068	0.0082	0.0112	0.0110
	Average (X ± SD)	0.0072      ± 0.0083	0.1008 ± 0.1361	0.1472 ± 0.1977	0.1074 ± 0.1423	0.1096 ± 0.1434	0.1106 ± 0.1519	0.1540 ± 0.2022
	2	0.0200	0.0250	0.0310	0.0340	0.0340	0.0360	0.0360
	6	0.0000	0.0012	0.0140	0.0110	0.0120	0.0110	0.0160
	Average (X ± SD)	0.0100 ± 0.0141	0.0131 ± 0.0168	0.0225 ± 0.0120	0.0225 ± 0.0163	0.0230 ± 0.0156	0.0235 ± 0.0177	0.0260 ± 0.0141
	3	0.0020	0.0030	0.0040	0.0070	0.0210	0.0130	0.0190
	7	0.0200	0.0330	0.0300	0.0100	0.0230	0.0130	0.0130
	Average (X ± SD)	0.1010 ± 0.1400	0.0180 ± 0.0212	0.0170 ± 0.0184	0.0085 ± 0.0021	0.0220 ± 0.0014	0.0130 ± 0	0.0160 ± 0.0042
	4	0.0000	0.0020	0.0030	0.0030	0.0040	0.0040	0.0050
	8	0.0100	0.0180	0.0780	0.0330	0.0460	0.0440	0.0460
	Average (X ± SD)	0.0050 ± 0.0071	0.0100 ± 0.0113	0.0405 ± 0.0530	0.0180 ± 0.0212	0.0250 ± 0.0297	0.0240 ± 0.0283	0.0255 ± 0.0290

From the graph above, it seems that there are fluctuations on the UV absorption readings from the UV-visible spectrometer on all four types of suppository samples analysed in this experiment.

            There are 4 parameters of an in vitro suppository drug release: Temperature, contact area, release medium, and membranes.

           

            The first parameter is temperature. In the whole set of experiment, we are using 37^oC as the body temperature for the tests of drug release from suppositories. Although for most practical purposes they can be set at 37^oC but that is not always the reality. As the body temperature may have slight variations at different periods of time, for example, during sleep time, when our metabolism rate is minimal, the temperature may slightly decrease. At this point, measurements of drug release at 37^oC may become an overestimation due to differences in melting range of the suppository. Also, comparing bases at 37^oC may cause such erroneous conclusions, as what we can see above. We should be aware that the temperature at which testing is performed is crucial, especially when aging has occurred.

            We shall also consider the release medium that is used in this test. We could consider the release medium in this test as the distilled water, which traverses through the dialysis bag, once it is immersed into the beaker of distilled water. But we need to realize that there is no ideal solution yet due to the problem of choosing a suitable volume and composition of the release medium that suits the condition in the rectal area.

            Another problem is due to the membrane factor. The ‘membrane’ that we used in this experiment is the dialysis bag. This may come with an enormous drawback as the release measured in the outer compartment is not equal to the actual release that is taking place in the inner compartment. We do need to consider that, the membrane may form a resistance to passing drug molecules, and that the actual release may be underestimated, although there are stands saying that membrane signifies the rectal mucosa.

            Taking back a look on the experiment results, theoretically all four suppositories should obtain a graph of increasing values, only to be varied is the degree of the increase and the values. Suppository I which contain the least PEG 6000 base should have meet the conditions for an optimum drug release, yet much to our disappointment, the results are not in line with our theory.

            The results produced by Suppository II could meet with our theory, i.e. the suppository which contains PEG 6000 and PEG1000 as the base could release the drug in a short period of time, and amount of drug release is increasing and keep constant after 15-20 minutes

5. What is the function of every substance used in this suppository preparation? How can the different contents of PEG 1000 and PEG 6000 affect the physical characteristics of the formulation of a suppository and the rate of release of drug from it?

Paracetamol used in the preparation of suppository is the active ingredient. Polyethylene glycol used acts as water soluble suppository base.  It enables the suppository to dissolve in rectal solution and release the medication easily. 
Combining PEG of different molecular weights in different ratio provides some control over the consistency, the drug release rate as well as the solidity/hardness of the suppository.
Increasing in the amount of PEG6000 over PEG1000 make the suppository becomes harder, but decrease the drug release rate.
PEG1000 has higher hydrophilicity compared to PEG6000. The drug release rate depends on the hydrophilicity whereby higher hydrophilicity give better drug release rate.

Conclusion:

The different amount of combination of PEG 1000 and PEG 6000 in the suppository preparation affects the physical and chemical characteristics of suppositories such as the rate of drug release, greasiness texture, and shape of the suppository.

References:

https://books.google.com.my/books?id=NFGSSSbaWjwC&pg=PA884&lpg=PA884&dq=function+PEG+6000+suppositories&source=bl&ots=V8_RNbgszr&sig=WdvXCgX0U414qj7Ja_nx1pOBNcE&hl=en&sa=X&ei=z11fVdaWFM7juQS6ooDACA&ved=0CDEQ6AEwAw#v=onepage&q=function%20PEG%206000%20suppositories&f=false

http://www.drugs.com/inactive/polyethylene-glycol-6000-274.html