Analysis of Pharmaceutical Ingredients, Excipients and Dosage Forms (Practical Workbook)

University of Szeged
ANALYSIS OF PHARMACEUTICAL INGREDIENTS,

EXCIPIENTS AND DOSAGE FORMS
(PRACTICAL WORKBOOK)
Edited by:
Dr. Zoltán Aigner
Dr. Piroska Szabó-Révész
Authors:
Dr. Mária Budai-Sz"cs
Dr. Erzsébet Csányi
Dr. Katalin Kristó
Dr. Péter Láng
Szeged, 2015
This work is supported by the European Union, co-financed by the European Social Fund,
within the framework of "Coordinated, practice-oriented, student-friendly modernization of
biomedical education in three Hungarian universities (Pécs, Debrecen, Szeged), with focus on
the strengthening of international competitiveness" TÁMOP-4.1.1.C-13/1/KONV-2014-0001
project.
The curriculum can not be sold in any form!
Analysis of pharmaceutical ingredients, excipients and dosage forms 2
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Contents
Introduction.................................................................................................................4
I. Preformulation ............................................................................................................6
1. Determination of the dissolution rate of the active substance ....................................7

2. Determination of the spectral transmission of glass containers................................13
3. Investigation of the gel formation of Carbomera......................................................19
4. Investigation of the consistency of suppository bases – Determination of the
resistance to rupture and flow point of suppositories ...............................................22
II. Investigation of dosage forms ...................................................................................28
5. Determination of the drop-weight of dosage forms used dropwise ..........................29

6. Determination of the viscosity of dextran infusion and the average
molecular weight of dextran .....................................................................................38
7. Investigation of emulsions ........................................................................................43
8. Investigation of suspensions .....................................................................................49
9. Investigation of ointments I. – Investigation of drop point, solidifying point, oil
number and water absorbing capacity.......................................................................56
10. Investigation of ointments II. – Investigation of the washability and rheological
characteristics of ointments.......................................................................................61
11. Investigation of ointments III. – Characterisation of consistency by viscosity,
spreadability and adhesion investigations.................................................................66
12. Investigation of the water uptake of tablets ..............................................................74
13. Investigation of tablets ..............................................................................................80
14. Investigation of the disintegration of rectal and vaginal suppositories.....................85
III. Investigation of stability ...........................................................................................88
15. Determination of the decomposition (caramelization) of glucose solution ..............89

16. Investigation of the viscosity changes of hydrophilic sols I. ...................................94
17. Investigation of the viscosity changes of hydrophilic sols II....................................99
18. Influence of humidity on the geometric parameters of tablets ...............................103
19. Investigation of the stability of tablets containing acetylsalicylic acid I.
– Investigation of decomposition kinetics, calculation of shelf life using
(real time) long-term stability test...........................................................................108
20. Investigation of the stability of tablets containing acetylsalicylic acid II.
– Investigation of decomposition kinetics, calculation of shelf life
using stress test .......................................................................................................113
IV. Biopharmaceutical investigations ...........................................................................117
21. Investigation of the drug release of emulsions by static diffusion method.............118

22. Investigation of drug release by agar plate diffusion method.................................124
23. Investigation of drug release from suppositories by dynamic diffusion method....131
24. Investigation of the mucoadhesivity of hydrogels ..................................................136
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Appendix.................................................................................................................142
Helios Alpha UV-Vis spectrophotometer ...............................................................143

Anton Paar RHEOLAB MC 1 rheometer ...............................................................145
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Introduction
Medicine is a special commodity, which means that besides the active pharmaceutical
ingredients (API) (equivalent terms: active substance, active ingredient, drug substance,
medicinal substance) and the excipients, the product also involves the sum of technological
processes and additional information which leads to effective therapy.
For the formulation of a medicine with the desired effect and stability, comprehensive
physical, physical-chemical and biopharmaceutical knowledge is needed, which must be
applied already during the API research and development, and later through the process of
development, licensing and production as well. It is very important for the professionals
participating in development, production and control to adopt the approach which leads to the
solution of the above problem.
As the first step of development, preformulation studies are made. During the
preformulation studies the physical-chemical properties of the API and excipients are
investigated, they are qualified from the aspect of pharmaceutical technology and the
compatibility test of the ingredients is performed. The investigation of polymorphism, the
selection of API salts, early method developments and the analysis of the effects of
technological procedures are carried out in this stage of development. Examples for
preformulation, such as determination of solubility and dissolution rate as physical-chemical
tasks; studying of the effect of salts and pH on hydrophilic sols and gels as compatibility tests,
and investigation of the consistency of suppositories are all included in this book.
After – or possibly parallel to – the preformulation studies, stability tests are started in the
formulation development. With regard to stability, we can speak of chemical, physical-
chemical, microbiological, therapeutic and toxicological stability. The book discusses the
degradation kinetics of the API in the dosage form and its interpretation; and then in view of
kinetics, the calculation of shelf life is presented by using long term and stress tests.
Parameters indicating stability need to be determined in order to establish the physical-
chemical stability of dosage forms, because international guidelines (ICH, EMA) regulating
stability tests do not specify special parameters for the description of systems, they must be
ascertained during the development. The book provides some examples for the
characterization of dosage form stability, such as phase separation rate (emulsions), half-time
(suspensions), oil number (ointments), rheological parameters (yield point, viscosity,
thixotropy), disintegration time (tablets, suppositories).
_______________________________________________________________________________
The investigation of dosage forms can be conducted on the basis of the methods of the
Pharmacopoeia (disintegration, dissolution, viscosity measurements), but in many cases there
are no specific investigations for the dosage form, and then we can opt for investigations not
listed in the Pharmacopoeia. Such investigations are also presented in this book, for example,
redispersibility of suspensions, determination of the spreadability, adhesion and washability
of ointments.
In the book, great emphasis is laid on biopharmaceutical investigations. Dissolution and
mucoadhesivity tests belong to this type of investigations. In case of dissolution and diffusion
tests, pharmacopoeial and non-pharmacopoeial investigations of emulsions, ointments and
suppositories are presented. As regards mucosal drug delivery, the adhesion of the form on
the mucosa is of key importance, because in the absence of this drug absorption is
questionable. The book presents in vitro rheological methods for the investigation of
mucoadhesion, which can indicate the in vivo adhesivity of the system.
The investigation of containers is discussed in the Pharmacopeia in great detail. This type
of measurements is represented by the light permeability measurements of glass containers.
The book summarizes the main pharmacopoeial and non-pharmacopoeial investigations.
Because of the complexity and manual needs of the tasks, the students perform them in pairs.
The execution of the tasks, the preparation of the report and the (statistical) evaluation of the
results help the students to adopt the proper approach and to acquire manual skills.
Szeged, 1st June, 2015
Dr. Mária Budai-Sz"cs Dr. Piroska Szabó-Révész Dr. Zoltán Aigner

_______________________________________________________________________________
I. Preformulation
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1. Determination of the dissolution rate of the active substance
Introduction
The development of the effect of an active substance depends on several factors. One of
the most important of these is dissolution rate, which determines what quantity of the
pharmaceutical product is dissolved during its residence time in the gastrointestinal tract. The
bioavailability of the active substance may be impaired greatly by its possibly slow
dissolution rate. During the practice, we are going to examine the time course of the
dissolution of active substances, which can provide important data for formulation.
The rate of dissolution is the amount of the dissolved material (dw) that diffuses on surface
(A), perpendicularly to the surface, through layer thickness (d(x)), at constant temperature, as
a result of concentration change (d(c)) in time dt:
dw d(c)
=-D A (1)
dt d(x)
where D is the diffusion constant (Fick’s law).
The dimension of the dissolution rate constant is: mg · cm-2 ·hour-1.
According to the Noyes-Whitney equation, the dissolution rate constant can be determined
with the following equation in the case of first order kinetics:
2.303 [Cs ]
k= log (2)
t [Cs ] [C ]
where t is the time, C is the current concentration of the solute, Cs is the saturation
concentration of the solute (e.g. at room temperature 0.50 g of theobromine dissolves in 1000
mL of distilled water).
The rate of dissolution depends on the following factors:
Test factors:
– mixing intensity, mixer type, geometric factors;
– flow conditions;
– concentration gradient;
– material quality of the solvent;
– temperature of the solvent, etc.
Physical-chemical factors:
– amorphous or crystalline state of the active substance;
_______________________________________________________________________________
– polymorphism;
– particle size;
– complex formation, eutectic formation, solid solution;
– chemical structure;
– presence of surfactants.
In practice, the rate of dissolution is usually tested by measuring the concentration increase
in the solution surrounding the solid material, or more rarely by determining the mass
decrease of the solid material not yet dissolved. There are several methods in literature for
determining the rate of dissolution, but it is important to distinguish between the dissolution
rate of the pure active substance and the rate of release of the active substance from the
dosage form. The specific properties of the active substance are determined in the first case,
while in the second case the quantity of the active substance released from the dosage form is
measured.
Task
1. During the practice, you are going to examine the dissolution rate of a specific quantity
of theobromine with the rotating paddle apparatus described in the Hungarian
Pharmacopoeia. Determine the active substance concentration of the samples taken at
the given times spectrophotometrically. Include the obtained results in a table.
Apparatus
Several methods and apparatuses have been developed for testing the rate of dissolution,
and the two most common ones, the rotating basket and the rotating paddle procedures are
official both in the American and the Hungarian Pharmacopoeias (Figure 1.1) (Ph. Hg. VIII.
2.9.3.).
_______________________________________________________________________________
Investigation method
Measure theobromine samples for three parallel examinations. 0.2500 g of theobromine

measured with analytical precision is used during the examination. A powder form is tested, it
is either spread on the top of the dissolution medium or filled in a capsule.
Perform three parallel examinations in the paddle dissolution apparatus with the
observation of the given parameters. Start your work by heating up the dissolution apparatus,
measuring and warming the dissolution medium. Measure 900 mL of dissolution liquid
(distilled water) into the vessel of the apparatus and set the temperature to 37 ± 0.5 °C with
the thermostat. The rotational speed of the paddle should be 100 rotations/minute, and
samples should be taken 5, 10, 20, 30, 60 and 90 minutes after the start of the examination.
Take 10 mL of sample with a pipette and filter it on a paper filter. Sampling should be made
at half height of the vessel, from the middle between the shaft of the paddle and the wall of
the dissolution vessel. Do not supplement the volume of the samples taken.
Figure 1.1: Basket and paddle dissolution apparatus
Prepare appropriate dilutions from the filtered samples in volumetric flasks in order to
obtain a measurable extinction (0.3 – 0.8 A) and measure their light absorption with a
spectrophotometer. Perform the measurements at a wavelength of R = 273 nm, 0.1 A = 1.6529
µg/mL.
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2. Calculate the dissolution rate constant of theobromine (k).
3. Plot the average of c* against sampling time.
4. Write a short evaluation of the results of the test.

_______________________________________________________________________________
Report
1. Determination of the dissolution rate of the active substance
Name: ..................................................... Group: ....................... Date: ............................
1. During the practice, you are going to examine the dissolution rate of a specific quantity
of theobromine with the rotating paddle apparatus described in the Hungarian
Pharmacopoeia. Determine the active substance concentration of the samples taken at
the given times spectrophotometrically. Include the obtained results in a table. (c* is the
amount of dissolved theobromine expressed as the percentage of the measured amount.)
2. Calculate the dissolution rate constant of theobromine (k).
Measurement of theobromine
Test 1 Test 2 Test 3
_________ with material (g)
_________ empty (g)
Measurement (g)
Results of the dissolution test of theobromine

Measurement Time (minute) 5 10 20 30 60 90
A
dilution
1. c (µg/mL)
dissolved theobromine (mg)
c*
A
dilution
2. c (µg/mL)
c*
A
dilution
3. c (µg/mL)
c*
c* average
k
_______________________________________________________________________________
3. Plot the average of c* against sampling time.
4. Write a short evaluation of the results of the test.

_______________________________________________________________________________
2. Determination of the spectral transmission of glass containers
Introduction
Glass containers are most commonly used for storing and dispensing liquid dosage forms.
A great number of APIs, excipients and dosage forms are light-sensitive, that is why they
must be dispensed and stored in glass containers protecting against light, which are tested
according to the Pharmacopeia.
According to Ph. Hg. VIII., glass containers for pharmaceutical use are glass articles
intended to come into direct contact with pharmaceutical preparations. Coloured and
colourless glass can be distinguished. Colourless glass is highly transparent in the visible
spectrum. Coloured glass is obtained by the addition of small amounts of metal oxide(s),
chosen according to the desired spectral absorbance.
The Hungarian Pharmacopeia (Ph. Hg. VIII., 3.2.1.) makes the following
recommendations about glass containers: Preparations for parenteral administration are
normally presented in colourless glass, but coloured glass may be used for substances known
to be light-sensitive. Colourless or coloured glass is used for the other pharmaceutical
preparations. It is recommended that all glass containers for liquid preparations and for
powders for parenteral administration permit the visual inspection of the contents.
Concerning glass containers, the Hungarian Pharmacopeia prescribes physical and
chemical investigations.
Physical investigations:
– determination of filling volume
– spectral transmission for coloured glass containers
Chemical investigations:
– investigation of hydrolytic resistance
A. hydrolytic resistance of the inner surface of glass containers (surface test)
B. hydrolytic resistance of glass grains (glass grains test)
C. to determine whether the containers have been surface-treated (etching test)
D. surface hydrolytic resistance - determination by flame atomic absorption
spectrometry (faas)
– investigation for arsenic compounds
_______________________________________________________________________________
Task
Break the glass container or cut it. Select sections representative of the wall thickness and
trim them as suitable for mounting in a spectrophotometer. Make sure the length of the
specimen is greater than that of the slit. Before placing in the holder, wash, dry and wipe the
specimen with lens tissue. Take care to avoid leaving fingerprints or other marks.
Apparatus: UV-VIS spectrophotometer, in transmittance mode
Place the specimen in the spectrophotometer in such a way that the light beam is
perpendicular to the surface of the section (Figure 2.1). Measure the transmission
(transmittance, T) of the specimen with reference to air in the spectral region of 290-450 nm,
at intervals of 20 nm.
Figure 2.1: Placing of the specimen into the cuvette holder of the spectrophotometer
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Limits (Ph. Hg. VIII.; 3.2.1.)
The observed spectral transmission for coloured glass containers for preparations that are
not for parenteral use does not exceed 10 per cent at any wavelength in the range of 290 nm to
450 nm, irrespective of the type and the capacity of the glass container. The observed spectral
transmission in coloured glass containers for parenteral preparations does not exceed the
limits given in the Table below.
Limits of spectral transmission for coloured glass containers for parenteral preparations
(Ph. Hg. VIII, Table 3.2.1.-5.)
Maximum percentage of spectral

transmission at any wavelength between
290 nm and 450 nm
Filling volume (mL) Flame-sealed Containers with
containers closures
Up to 1 50 25
Above 1 and up to 2 45 20
Above 20 25 10
1. Measure each specimen in 4 different positions, in this way you will get 4 values for
each sample. Calculate the average of 4 measurements, and plot the data against the
wavelength. Evaluate the results according to the pharmacopoeial limits.
2. Correlate the different glass samples in such a way that their transmission refers to a
wall thickness of 1 mm, by using the following equation:
lg T%' lg T%''
lg T% = l + lg T%'' (1)
l1
where T% is the light permeability of the minimum thickness of the glass container in %
transmittance, T’% is the maximum % light permeability measured in the investigated
wavelength range, T”% is the theoretical light permeability (92%) of the glass container
of a wall thickness of 0 mm (lg T’’% = 1.9638), l1 is the average thickness of the glass
sample in mm, l = 1 mm.
Measure the thickness of the glass samples at 10 points of the specimen with a
micrometer screw. Calculate the average values of wall thickness.
_______________________________________________________________________________
Report
2. Determination of the spectral transmission of glass containers
Name: ..................................................... Group: ....................... Date: ............................
1. Measure each specimen in 4 different positions, in this way you will get 4 values for
each sample. Calculate the average of 4 measurements, and plot the data against the
wavelength. Evaluate the results according to the pharmacopoeial limits.
Glass sample (code, characteristic):

Sample 1: .................................................................
Sample 2: .................................................................
Sample 3: .................................................................
Sample 4: .................................................................
Sample 1 Sample 2
<
(nm) 1 2 3 4 average 1 2 3 4 average
290
310
330
350
370
390
410
430
450
< Sample 3 Sample 4
(nm) 1 2 3 4 average 1 2 3 4 average
290
310
330
350
370
390
410
430
450
_______________________________________________________________________________
Evaluation:
_______________________________________________________________________________
2. Measure the thickness of the glass samples at 10 points of the specimen with a
micrometer screw. Calculate the average values of wall thickness.
Measurement Thickness (mm)

Sample 1 Sample 2 Sample 3 Sample 4
1
2
3
4
5
6
7
8
9
10
average
Transmittances referring to a thickness of 1 mm:
Sample 1, T% = ..................................................
Sample 2, T% = ..................................................
Sample 3, T% = ..................................................
Sample 4, T% = ..................................................
Evaluation:
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3. Investigation of the gel formation of Carbomera
Introduction
Carbomers are high molecular mass polymers of acrylic acid cross-linked with polyalkenyl
ethers of sugars or polyalcohols (Ph. Hg. VIII. 04/2009:1299).
A wide variety of Carbomera product types are used in the field of pharmaceutical
industry. Carbomers are excellent viscosity increasing excipients, and they are used as
transparent aqueous or hydroalcoholic gel forming agents. Their favourable properties are
mostly due to the carboxylic groups of acrylic acid. Various degrees of neutralization are
needed for gelation in an aqueous phase. The aim of the present work is to investigate the
neutralization effect on gel formation.
The aim of the practice is the investigation of the gel formation of Carbomers depending
on pH and the determination of viscosity at different pH values.
Task
1. Prepare 50.0 g of 0.3 % Cabomera containing system. The polymer should be poured
onto the surface of the previously measured distilled water while stirring continuously.
Measure the pH of the aqueous suspension, then the viscosity at an increasing shear rate
(0.1 – 100 1/s) at 12 points.
2. Prepare again 50.0 g of 0.3 % Cabomera containing system. The polymer should be
poured onto the surface of the previously measured 40 g of distilled water while stirring
continuously. Start neutralization up to pH = 5 by adding 0.1 N NaOH in parts. After
reaching the desired pH, add the remaining distilled water up to 50.0 g and stir
vigorously to homogenize it. Measure the viscosity at an increasing shear rate (0.1 –
100 1/s) at 12 points.
3. Prepare again Carbomera containing systems according to point 2. Adjust the pH up to

pH = 6; 7; 8; 9 and 10 by adding 0.1 N of NaOH in parts. Measure the viscosity at an
increasing shear rate (0.1 – 100 1/s) at 12 points.
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Report
3. Investigation of the gel formation of Carbomera
Name: ..................................................... Group: ....................... Date: ............................
1. Prepare 50.0 g of 0.3 % Cabomera containing system. The polymer should be poured
onto the surface of the previously measured distilled water while stirring continuously.
Measure the pH of the aqueous suspension, then the viscosity at an increasing shear rate
(0.1 – 100 1/s) at 12 points.
2. Prepare again 50.0 g of 0.3 % Cabomera containing system. The polymer should be
poured onto the surface of the previously measured 40 g of distilled water while stirring
continuously. Start neutralization up to pH = 5 by adding 0.1 N of NaOH in parts. After
reaching the desired pH, add the remaining distilled water up to 50.0 g and stir
vigorously to homogenize it. Measure the viscosity at an increasing shear rate (0.1 –
100 1/s) at 12 points.
3. Prepare again Carbomera containing systems according to point 2. Adjust the pH up to

pH = 6; 7; 8; 9 and 10 by adding 0.1 N of NaOH in parts. Measure the viscosity at an
increasing shear rate (0.1 – 100 1/s) at 12 points. Include the obtained results in a table.
pH without 5 6 7 8 9 10
adjustment
measured
pH
D B C B C B C B C B C B C B C
(1/s) (Pa·s) (Pa·s) (Pa·s) (Pa·s) (Pa·s) (Pa·s) (Pa·s)
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
11.
12.
13.
_______________________________________________________________________________
4. Plot the viscosity values measured at the highest shear rate against pH.
Give the minimum pH value needed for gel formation. Use the curve-fitting method.
pH = .......................................................................
Evaluation:
_______________________________________________________________________________
4. Investigation of the consistency of suppository bases –

Determination of the resistance to rupture and flow point of
suppositories
Introduction
The investigation of the consistency and hardness of suppositories is important with

respect to applicability. This investigation gives useful information on how the suppository
can retain its shape during packaging, storage and shipping. It is also important concerning
administration, as suppositories with not enough hardness cannot be administered easily,
while the ones that are too hard can cause pain.
Task
1. Determination of resistance to rupture

Apparatus: Apparatus for the determination of the resistance to rupture of suppositories
This test determines, under defined conditions, the resistance to rupture of suppositories
measured by the mass needed to rupture them by crushing (Ph. Hg. VIII. 2.9.24.) (Figure 4.1)
Check that the apparatus is vertical. Heat the thermostatted chamber to 25 °C. The dosage
form to be tested has been maintained for at least 24 h at the required measuring temperature.
Place the suppository vertically between the jaws in the sample holder with the point
upwards. The top pressure block of the suspension loading rod is carefully positioned and the
test chamber is closed with its glass window; for each determination, position the suppository
in the same manner with respect to the direction of the force applied. Wait for 1 min and add
the first 200 g disc. Again wait for 1 min and add another disc. Repeat the operation until the
suppository collapses. The mass required to crush the suppository is calculated by the sum of
the masses weighing on the suppository when it collapses [including the initial mass of the
device (600 g)] assessed as follows:
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– if the suppository collapses within 20 s of placing the last disc, do not take this mass into
account,
– if the suppository collapses between 20 s and 40 s of placing the last disc, use only half of
this mass in the calculation, i.e. 100 g,
– if the suppository remains uncrushed for more than 40 s after the last disc is placed, use all
the mass in the calculation.
Figure 4.1: Apparatus for the determination of the resistance to rupture of suppositories
Carry out each measurement on five suppositories, making sure that no residue remains
before each determination (Ph. Eur.). Fill the attached table.
2. When the measurement is finished, prepare the different types of suppositories of the
composition given by your practice leader. Calculate 3.0 of base per suppository
together with excess. Use the moulding technique and place the finished suppositories
into boxes.
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Compositions of series:
Series A:
1. Adeps solidus compositus
2. Adeps solidus compositus + 30 % Talcum
3. Adeps solidus compositus + 10 % Paraffinum liquidum
4. Adeps solidus compositus + 15 % Paraffinum liquidum + 30 % Talcum
Series B:
1. Adeps solidus
2. Adeps solidus + 30 % Talcum
3. Adeps solidus + 10 % Oleylum oleinicum
4. Adeps solidus + 15 % Oleylum oleinicum + 30 % Talcum
3. Determination of consistency by penetrometry

Apparatus: penetrometer (Ph. Hg. VIII. 2.9.9.) (Figure 4.2)
Determination of the consistency of test samples prepared from suppository bases with and
without excipients by measuring the flow point of suppositories. We measure the penetration
depth of the penetrating object into the test sample prepared from the series to be investigated.
The flow point ( Fp (N/cm2)) is calculated from the weight of penetrating object (G (N)) and
the depth of penetration (cm):

4G
Fp = (1)
s2
Place the test sample on the base of the penetrometer. Verify that its surface is
perpendicular to the vertical axis of the penetrating object. Bring the temperature of the
penetrating object to 25 ± 0.5 °C and then adjust its position such that its tip just touches the
object and hold it free for 5 s. Set the zero reading, release the penetrating object and measure
the depth of penetration in mm scale. Carry out 5 parallel measurements with each sample,
include the results in the table and evaluate them.
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Figure 4.2: Penetrometer
4. When the measurements are finished, prepare the test samples of the composition given
by your practice leader in a small Petri dish. (Calculate with 30.0 g of base per test
sample.)
Composition of series:
Series A:
1. Adeps solidus compositus
2. Adeps solidus compositus + 30 % Talcum
3. Adeps solidus compositus + 10 % Paraffinum liquidum
4. Adeps solidus compositus + 15 % Paraffinum liquidum + 30 % Talcum
Series B:
1. Adeps solidus
2. Adeps solidus + 30 % Talcum
3. Adeps solidus + 10 % Oleylum oleinicum
4. Adeps solidus + 15 % Oleylum oleinicum + 30 % Talcum
_______________________________________________________________________________
Report
4. Investigation of the consistency of suppository bases – Determination of the resistance
to rupture and flow point of suppositories
Name: ..................................................... Group: ....................... Date: ............................
1. Determination of resistance to rupture
Letter symbol of series: ..........................................................
Mark Measurement Measurement Measurement Measurement Measurement Average

of 1 2 3 4 5
supp. Mass F Mass F Mass F Mass F Mass F Mass F
(g) (N) (g) (N) (g) (N) (g) (N) (g) (N) (g) (N)
......./
1
......./
2
......./
3
......./
4
2. Determination of the flow point of suppository
Mark of
....... / 1 ....... / 2 ....... / 3 ....... / 4
supp.
Pit Fp Pit Fp Pit Fp Pit Fp
Measurement 2 2 2
(cm) (N/cm ) (cm) (N/cm ) (cm) (N/cm ) (cm) (N/cm2)
1.
2.
3.
4.
5.
average
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3. Evaluation:
_______________________________________________________________________________
II. Investigation of dosage forms

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5. Determination of the drop weight of dosage forms used

dropwise
Introduction
Numerous dosage forms administered in drop form are known. Ph. Hg. VIII. lists the
following in this field:
– Liquid preparations for oral use: oral drops
– Ear preparations: ear drops
– Nasal preparations: nasal drops
– Oromucosal preparations: oromucosal drops
– Eye preparations: eye drops
Drops are solutions, emulsions or suspensions that are administered in small volumes such
as drops by the means of a suitable device.
Ph. Hg. VIII. prescribes dosage form investigations, such as dose and uniformity of dose,
only for oral drops in the monograph (Ph. Hg. VIII., p. 2985). In the case of other types of
drops, there is no prescribed investigation related to the drop form.
The weight of the drop can be modified by the excipients in the formulation. The weight of
drops (m) dropped out from the dropping tube depend on the external diameter of the
dropping tube (d) and the surface tension of the liquid (+) (Tate 1864):
d
m= x
(1)
g
where m is the drop weight in g, d is the diameter of the dropping tube in cm, , = 3.14, g =
981 cm·s-2, +x is the surface tension (mN/m).
Besides the above factors, the drop weight also depends on the property of the dropping
tube, the speed of dropping, the way the dropping tube is held and the force between the
surface of the dropping tube and the liquid dropping out (e.g. adhesion, wetting).
When checking the dose of drops, the drop number and the drop weight also have to be
taken into consideration. The drop number shows how many drops can be prepared from 1 g
of solution with a standard dropping tube (Figure 5.1).
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Both the drop weight and the drop number change with changing surface tension.
Figure 5.1: Standard dropping tube
The aim of the investigation is to examine the changes of the drop weight of the dosage
form used dropwise as a result of the changing of the surface tension with different excipients.
There is proportionality between the drop weight and the surface tension of the purified
water and the solution containing different excipients:
mv v
= (2)
mx x
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where mv is the drop weight of purified water (g), mx is the drop weight of the examined
solution (g), +v is the surface tension of purified water (Aqua purificata) (mN/m), +x is the
surface tension of the examined solution (mN/m).
There is inverse proportionality between the drop number prepared from 1 g of solution
and the surface tension. The surface tension of purified water is known (73 mN/m at 20 °C),
mv and mx can be determined experimentally, so +x can be calculated:
x = mx 73 (3)
mv
Tasks
1. Prepare two of the solution series of the following composition, chosen by your practice
leader.
Components A/1 A/2 A/3 A/4 A/5 B/1 B/2 B/3 B/4 B/5
Alcoholum 96% (g) 1.5 3.0 4.5 6.0 7.5 – – – – –
Mucilago methylcellulosi (g) - - - - - 0.75 1.5 2.25 3.0 3.75
Aqua purificata (ad g) 15 15 15 15 15 15 15 15 15 15
Components C/1 C/2 C/3 C/4 C/5 D/1 D/2 D/3 D/4 D/5
Polysorbatum 20 (g) 0.15 0.45 0.75 1.05 1.35 – – – – –
Tinctura aromatica (g) – – – – – 1 2 3 4 5
Aqua purificata (ad g) 15 15 15 15 15 15 15 15 15 15
2. Determine the drop number of the prepared solutions and the purified water with the
given dropping tube. Make 10 parallel measurements.
3. Fill in the tables. Calculate the values of +x. Evaluate the results statistically. (Calculate
standard deviation, relative standard deviation according to the equations in Table
2.9.40-2 of Ph. Hg. VIII.)
_______________________________________________________________________________
4. Plot the measured drop weight values against the concentration of the examined
excipients on a graph paper. How does the surface tension of the solutions change with
the concentration of the excipients?
5. If 15 g of excipient-free aqueous solution contains 0.30 g of codeine hydrochloride
dihydrate, how is the codeine hydrochloride dihydrate content in one drop of solution
changed by adding the excipients in different concentrations? Fill in the table. Evaluate
the data.
_______________________________________________________________________________
Report
5. Determination of the drop weight of dosage forms used dropwise
Name: ..................................................... Group: ....................... Date: ............................
1. Fill in the tables. Calculate the values of +x. Evaluate the results statistically. (Calculate
standard deviation, relative standard deviation.)
Name of the excipient used: .....................................................
Drop Drop Drop Drop Drop Drop

number of number of number of number of number of number of
Measurements
water solution solution solution solution solution
.......... .......... .......... .......... ..........
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
average
stand. dev.
rel. stand. dev.
drop weight (cg)
\ (mN/m)
_______________________________________________________________________________
Drop Drop Drop Drop Drop

number of number of number of number of number of
Measurements
solution solution solution solution solution
.......... .......... .......... .......... ..........
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
average
stand. dev.
rel. stand. dev.
drop weight (cg)
\ (mN/m)
_______________________________________________________________________________
2. Plot the measured drop weight values against the concentration of the examined
excipients on a graph paper. How does the surface tension of the solutions change with
the concentration of the excipients?

_______________________________________________________________________________
3. Fill in the table corresponding to the composition series chosen by your practice leader.
Evaluate the results.
Alcoholum 96% Amount of Mucilago methylcellulosi Amount of

codeine codeine
(m/m %) hydrochloride (m/m %) hydrochloride
dihydrate in 1 dihydrate in 1
drop drop
(mg) (mg)
10 5
20 10
30 15
40 20
50 25
_______________________________________________________________________________
Polysorbate 20 Amount of Tinctura aromatica Amount of

codeine codeine
(m/m %) hydrochloride (m/m %) hydrochloride
dihydrate in 1 dihydrate in 1
drop drop
(mg) (mg)
1 6.6
3 13.3
5 20.0
7 26.7
9 33.3
Evaluation:
_______________________________________________________________________________
6. Determination of the viscosity of dextran infusion and the

average molecular weight of dextran
Introduction
On the basis of their origin or synthesis, macromolecular compounds can be natural (e.g.
starch, gelatine, etc.), semisynthetic (e.g. dextran, cellulose derivatives, etc.) or synthetic (e.g.
polyvinyl-alcohol, polyethylene glycol, etc.). Spherical and linear macromolecules can be
distinguished according to the shape of the macromolecules. The original structure and
concentration of the dissolved macromolecular material have a profound effect on the
properties of the formed colloidal solution, among which flow behavior is very important.
Threadlike macromolecules are inordinately located in all directions of the space or they may
adhere to each other, which is determinative in terms of viscosity.
Dextran is produced by the fermentation of sucrose using the bacterial strain and substrains
of Leuconostoc mesenteroides. In the 8th Edition of the Hungarian Pharmacopoeia 4 dextrans
with different average molecular weight are official. For medical purposes, the fraction
having a molecular weight of 30,000 – 80,000 is the most advantageous, because it has the
least harmful side effect. One of the requirements for dextran used as a plasma substitute is
that its viscosity should be constant and approximately identical to the viscosity of the blood
plasma, the other is that its molecular weight should not change during storage and
sterilization.
The average molecular weight of dextran can be determined with simple physical
investigations by using the dextran solution.
Task
Determination of the density and dynamic viscosity of dextran solution. Calculation of the
average molecular weight of dextran from the results by using a formula.
_______________________________________________________________________________
1. Determination of density
Apparatus: pycnometer
For the determination of the density of the liquid, use a pycnometer with a volume of 10.00
mL. Measure the weight of the empty and dry pycnometer on an analytical balance, then fill it
up with the test liquid of ~20 °C until the meniscus of the liquid is under the circular mark.
Then put the pycnometer into a water bath of 20 ± 0.1 °C. After 10 minutes, fill up the
pycnometer in the water bath with a pipette so that the lowest point of the meniscus touches
the circular mark. If necessary, dry the neck of the pycnometer inside with a filter paper and
close it with its plug.
After taking out the pycnometer, dry it carefully, place into the balance box for 10 minutes,
then measure its weight. The difference between the weight of the empty pycnometer and that
of the liquid-filled one gives the weight of the liquid enough to fill the flask (mf).
After that, empty the pycnometer and wash it carefully with water of ~20 °C; then fill it up
with water in the way described above and calculate the weight of water (mv).
From the quotient (mf/mv) of the weight of liquid (mf) and water (mv), measured in the same
way, the density at 20 °C (^20 °C) can be calculated with the following formula:
mf
20° C = 0.997003 + 0.0012 (1)
mv
Give the calculated value of density rounded to 3 decimal places.
2. Determination of viscosity
Apparatus: Ostwald-Fenske capillary viscometer No 5 (Figure 6.1)
The Hungarian Pharmacopoeia (Ph. Hg. VIII., 2.2.9.) prescribes the following for the
determination of viscosity using a capillary viscometer:
– the determination of viscosity using a capillary viscometer is carried out at a
temperature of 20 ± 0.1 °C, unless otherwise prescribed;
– the time required for the level of the liquid to drop from one mark to the other is
measured with a stop-watch to the nearest one-fifth of a second;
– the result is valid only if two consecutive readings do not differ by more than 1 %;
– the average of not fewer than three readings gives the flow time of the liquid to be
examined.
_______________________________________________________________________________
Figure 6.1: Capillary viscometer
The dextran infusion is diluted with isotonic sodium chloride solution up to 0.6 %. Then,
by using the Ostwald-Fenske capillary viscometer No. 5, the viscosity of the diluted infusion
is measured at 20 ± 0.1 °C.
For the determination of viscosity, the flow time of the defined volume of the liquid to be
examined in the capillary of a given length and diameter is measured. A certified capillary
viscometer should be used for the measurements. The capillary viscometer selected according
to the viscosity of the sample to be examined or its solution of the required concentration
should be placed into a water bath at constant (± 0.1 °C) temperature. The size of the water
bath should be sufficient for the examined sample in the capillary viscometer to be at least 20
mm under the surface of the water bath and also at least 20 mm from the bottom during the
entire examination.
Water should be used for filling the thermostat. During the measurement, the middle of the
thermometer's mercury reservoir should be at the same height as the middle of the capillary.
The flow time is measured with a stop-watch.
The bubble-free liquid to be examined is filled into the viscometer that has been degreased,
washed and dried carefully. About three-quarters of the spherical reservoir on the thick pipe
_______________________________________________________________________________
should be filled with the liquid to be examined. The viscometer is placed into the bath in such
a way that the stems ending in the gap are upright. The measurement can be started only if the
sample to be examined has taken over the temperature of the bath (approximately 10
minutes). After this time, the liquid to be examined has to be absorbed through the rubber
tube over the circular mark between the two balls above the capillary, then the absorption has
to be finished. The stop-watch should be started if the surface of the liquid sinking reaches the
circular mark between the two balls and it should be stopped if it reaches the circular mark
above the capillary. The time measured in this way is the flow time (t). The flow time should
be measured at least 5 times, and for the calculation of the result only the flow times which do
not show any one-way differences and differ not more than ± 0.5 % from each other should be
considered. The dynamic viscosity of the examined sample can be calculated by using the
following formula:
20°C =k 20°C t (2)

where 220°C is the dynamic viscosity in mPa·s at 20.0 °C, k is the constant of the instrument at
20.0 °C (according to the certification, it is: 8.65 °C·10-4 mm2/s2), t is the mean value of the
flow time in seconds (s), 620°C is the density of the sample to be examined at 20.0 °C in g/mL
(determined in an earlier examination).
Based on the results obtained, the average molecular weight can be calculated by using the
following formula:
= k M wa (3)
where 2 is the dynamic viscosity of the solution, k is a constant (9.78·10-4), a is the index
(0.5), Mw is the average molecular weight.
_______________________________________________________________________________
Report
6. Determination of the viscosity of dextran infusion and the average molecular weight
of dextran
Name: ..................................................... Group: ....................... Date: ............................
1. Prepare 50 mL of 0.9 % sodium chloride solution. How much NaCl should be used for
the preparation? .................................... g
2. Determine the density of the dextran infusion by using a pycnometer according to the
Hungarian Pharmacopoeia, with three parallel measurements, and fill in the table.
Number of pycnometer
Weight of empty pycnometer (g)
Weight of pycnometer filled with water (g)
Weight of pycnometer filled with dextran infusion (g)
Weight of water (g)
Weight of dextran infusion (g)
Density (g/cm3)
Average of density (rounded to three decimal places)
3. Determine the dynamic viscosity of the diluted dextran infusion with the help of a
modified Ostwald-Fenske capillary viscometer according to the Hungarian
Pharmacopoeia, with 5 parallel measurements. Include the results in the table below:
Number of measurement Flow time (s) Average of flow time (s) Viscosity (mPa·s)
1.
2.
3.
4.
5.
4. Calculate the average molecular weight of the dextran infusion according to the formula
given. The average molecular weight of dextran: ............................................
Evaluation:
_______________________________________________________________________________
7. Investigation of emulsions
Introduction
Emulsions are thermodynamically unstable systems. The reason for this is the large
specific surface area of the particles. The emulsified state lasts until the layers separating the
liquid drops retain their structure. Thus the constancy of the degree of dispersion is one of the
most important aspects concerning the stability of emulsions, because depending on the drop
size distribution – depending on time – they may float. The drop size is increasing with the
confluence of the floated or flocculated drops (coalescence); this leads to the breaking of the
emulsion, then the dispersed part begins to form a continuous phase, therefore a
thermodynamically favourable condition results. From the point of view of pharmaceutical
production, an emulsion is regarded stable if its degree of dispersion remains unchanged or
changes only within the prescribed limits until use.
During the investigation of the stability of emulsions, the rheological properties, which are
determined by the following factors, are also of great significance: the dispersed phase, the
viscosity of the dispersion medium, the amount and the material quality of the emulsifier
used. If the volumetric concentration of the dispersed phase is small enough, the system
behaves as a Newtonian or ideal viscous fluid. The yield curve of ideal viscous fluids is
characterized by a baseline starting from the origin, and they give a straight line. The flow
becomes pseudoplastic if the volumetric concentration is increased, as a result of which the
resistance of the system to flow increases. The yield curves of such materials also start from
the origin, but they are not linear. Viscosity changes depending on shear stress. The reason for
this is the instantaneous and reversible disaggregation or deformity of the emulsified drops
caused by shear force. The aim of this practice is the investigation of the characteristic
properties of emulsions in case of changing the factors influencing stability.
Task
1. The determination of the yield properties and viscosity of emulsions
The Hungarian Pharmacopoeia (Ph. Hg. VIII., 2.2.10.) lists the following types of
instruments for the determination of viscosity by using a rotating viscometer: spindle
_______________________________________________________________________________
viscometers, cone-plate viscometers and concentric cylinder viscometers. During the practice,
an Anton Paar Rheolab MC 1 Rheometer is used, which is a concentric cylinder type
viscometer. The fluid to be examined has to be located in the gap between the inner and outer
cylinders. Viscosity can be measured by rotating the inner cylinder ("Searle" type viscometer)
or by rotating the outer cylinder ("Couette" type viscometer).
Apparatus: Anton Paar Rheolab MC 1 Rheometer
Sample holder "Z3" (Figure 7.1) is filled hermetically with the emulsion to be examined up
to the inner mark. The cylindrical probe is affixed with the help of the retaining ring. The
device is installed according to the instructions of the manual and the required program is
started.
Figure 7.1: Sample holder "Z3" of the Anton Paar Rheolab MC 1 Rheometer
2. Investigation of spontaneous separation (not an official investigation method in Ph. Hg.

VIII.)
Apparatus: 25.0-mL measuring cylinder with a ground stopper
_______________________________________________________________________________
25.0 mL of emulsion is filled into a measuring cylinder with a ground stopper. The sample
is shaken a few times. The volume of the water separated and collected in the bottom of the
measuring cylinder is determined after a certain time (one and a half hours).
Calculation of the speed of separation:
1 V + Vt
v= log 0 (1)
t V0 - Vt
where v is the speed of separation (mL/hour), t is the time of observation (hour), Vt is the
volume of water separated during t time (mL), V0 is the original volume of the emulsion (25
mL).
Compositions to be examined:
Materials I. II. III. IV. V.

Paraffinum liquidum 10.0 g 30.0 g 60.0 g 30.0 g 30.0 g
Aqua purificata 68.0 g 48.0 g 8.0 g 47.0 g 49.5 g
Polysorbatum 20 2.0 g 2.0 g 2.0 g 3.0 g 0.5 g
Mucilago methylcellulosi 20.0 g 20.0 g 20.0 g 20.0 g 20.0 g
Apparatus: Mixer (stage 1, 1 min of mixing time)

_______________________________________________________________________________
Report
7. Investigation of emulsions
Name: ..................................................... Group: ....................... Date: ............................
1. The effect of the weight ratio of the dispersed phase on the properties of the emulsion:
Viscosity
Composition Speed of separation
(D = 656 1/s)
II. (10 %)
I. (30 %)
III. (60 %)
Evaluation:
2. The effect of the concentration of emulsifiers on the properties of the emulsion:
Viscosity
Composition Speed of separation
(D = 656 1/s)
V. (0.5 %)
I. (2 %)
IV. (3 %)
Evaluation:
_______________________________________________________________________________
3. Rheological characterization of the compositions tested (determination of flow type):
I. II. III. IV. V.

D B I D B I D B I D B I D B I
2 2 2 2
(1/s) (N/m ) (1/s) (N/m ) (1/s) (N/m ) (1/s) (N/m ) (1/s) (N/m2)
_______________________________________________________________________________
4. Plot and evaluate the yield curves of the compositions.
Evaluation:
_______________________________________________________________________________
8. Investigation of suspensions
Introduction
The two most important requirements to be met by suspensions are: the homogeneous
distribution of the active ingredient at least for the period between the mixing of the
suspension and the pouring of given dose, and the easy redispersibility of the sediment
formed during storage. Sedimentation cannot be prevented, but it can be delayed with various
technological methods. One possibility for this is to reduce the size of the non-dissolving
particles, or to enhance the viscosity of the dispersion medium with various macromolecular
materials (e.g. mucilages). With respect to the sediment formed, suspensions can be
flocculated or deflocculated. Flocculation occurs if the macromolecules adsorbed on the solid
particles are cross-linked, or if the surface parts of the particles not covered with
macromolecules stick together due to the high adhesion. The sediment of flocculated
suspensions is loose and bulky. Its advantage is that the floccules (grain aggregates) form a
sediment which is easily redispersible, but sedimentation is fast in this case. In contrast, the
sediment of deflocculated suspensions consists of particles in a close packed arrangement (so-
called "cemented sediment"), therefore it is difficult to shake, but its sedimentation is slower.
The sedimentation of suspensions can be free or hindered (see Figures 8.1 and 8.2).
Sedimentation is free if the suspension is sufficiently washy, in which the particles do not
hinder each other during sedimentation and a sediment with a gradually increasing volume is
formed, where the supernatant is cloudy because of the floating particles. Free sedimentation
is characterized by a high half-life and a low sedimentation rate. However, hindered
sedimentation is more typical of pharmaceutical suspensions. In this case, the particles hinder
each other during sedimentation. The reason for this is the higher concentration of the
particles in the given volume and the interactions between them, and also the presence of the
viscosity increasing excipient. Hindered sedimentation is characterized by a clear supernatant
and a gradually decreasing phase boundary between the sedimenting particles. The aim of this
practice is to determine the half-life (t1/2) of flocculated and deflocculated suspensions, and to
investigate the resuspensibility of the sediment.
_______________________________________________________________________________
Figure 8.1: Hindered settling suspension
Figure 8.2: Free settling suspension
Task
1. Investigation of the sedimentation of suspensions (not an official investigation method

in Ph. Hg. VIII.)
Apparatus: 25.0-mL measuring cylinder with a ground stopper
_______________________________________________________________________________
Pour the homogenous suspension into a measuring cylinder with a ground stopper and
shake it well. Observe the separation of the dispersed part and the dispersion medium. Note
the height of the sediment column in every 5 minutes during 30 minutes, then in every 15
minutes during 75 minutes. If the suspension becomes foamy after being poured into the
cylinder, the total distance should be measured from the bottom of the foam layer.
Determination of half-life: plot the height of the sediment column against time. In case of
hindered sedimentation:
l0 - l E (1)
l1/2 = l 0 -
2
In case of free sedimentation:
lE
l 1/2 = (2)
2
where l1/2 is the height of „half-distance”, l0 is the initial height, lE is the equilibrium height.
Plot the sediment volume against time. Mark the l1/2 values on the curve, and project them
onto the time axis. The value thus obtained is the half-life. During the determination of half-
life it is also acceptable if the volume of the sediment column is measured instead of its
height. The last measured value is regarded as the equilibrium height (lE) and the equilibrium
volume (VE) .
2. Redispersibility of the sediment (not an official investigation method in Ph. Hg. VIII.)
Apparatus: resuspension device
Carefully fix the cylinder containing the sedimented suspension into the stand of the
sample holder. Rotate the measuring cylinder until the sediment is uniformly distributed in the
medium. The number of rotations is the value of resuspension. The sediment in the bottom of
the measuring cylinder can be observed better if the device is stopped during the investigation
when the measuring cylinder is with its mouth downwards (make sure that the glass stopper
closes the measuring cylinder).
_______________________________________________________________________________
Compositions to be examined
Instrument: Mixer (stage 1, 1 min of mixing time)
Series „A” A/1 A/2 A/3 A/4

Bismuthi subgallas 5.0 g 5.0 g 5.0 g 5.0 g
Natrii dihydrogenophosphas 0.0 g 0.5 g 10.0 g 20.0 g
dihydricus
Aqua purificata ad 50.0 g ad 50.0 g ad 50.0 g ad 50.0 g
Series „B” B/1 B/2 B/3 B/4
Bismuthi subgallas 5.0 g 5.0 g 5.0 g 5.0 g
Mucilago hydroxyaethylcellulosi 0.0 g 0.5 g 10.0 g 20.0 g
Aqua purificata ad 50.0 g ad 50.0 g ad 50.0 g ad 50.0 g
Attention! Start the preparation of the suspensions in series „A” with the dissolution of
NaH2PO4.
_______________________________________________________________________________
Report
8. Investigation of suspensions
Name: ..................................................... Group: ....................... Date: ............................
1. Investigation of sediment volume (mL) depending on the time:
Time of sedimentation (min)

Sample
5 10 15 20 25 30 45 60 75
A/1
A/2
A/3
A/4
B/1
B/2
B/3
B/4
2. Plot the sediment volume against time (see on the following pages). Calculate and mark
the half-lives on the graph. Complete the table below with the results.
3. Effect of the composition changes of the dispersion medium on the sedimentation of

suspensions and the structure of the sediment:
Note: over 250 rpm, the suspension should be considered unshakeable.
Half-life Value of resuspension

Sample Type of sedimentation
(min) (rpm)
A/1
A/2
A/3
A/4
B/1
B/2
B/3
B/4
_______________________________________________________________________________
Series „A”
_______________________________________________________________________________
Series „B”
Write an evaluation about the results of the investigations. What cause and effect relationship
explains the observed phenomena?
_______________________________________________________________________________
9. Investigation of ointments I. – Investigation of drop point,

solidifying point, oil number and water absorbing capacity
Introduction
The determination of the physical properties of water-free ointments is very important for
cream development. It is essential to know the solidifying and drop points during the
manufacturing process. The oil number gives information about the binding force between the
solid and liquid lipophilic phases. By ascertaining the water absorbing capacity, the maximum
amount of water which can be emulsified into the water-free ointment can be determined.
Tasks
1. Determination of the drop point of ointments

The drop point is the temperature at which the first drop of the melting substance to be
examined falls from a cup under defined conditions.
Apparatus: Ubbelohde-thermometer (Ph. Hg. VIII. 2.2.17., Figure 9.1)
Figure 9.1: Ubbelohde-thermometer

_______________________________________________________________________________
Fill the cup to the brim with the substance to be examined without melting and bubbles.
Remove the excess substance at the 2 ends of the cup with a spatula. When sheaths A and B
have been assembled, press the cup into its housing in sheath B until it touches the supports.
Remove with a spatula the substance pushed out by the thermometer. Place the apparatus in
the water-bath. Heat the water-bath and, when the temperature is at about 10 °C below the
presumed drop point, adjust the heating rate to about 1 C/min. Note the temperature at the fall
of the first drop. Carry out at least 3 determinations, each time with a fresh sample of the
substance. The difference between the readings must not exceed 3 °C. The average of 3
readings is the drop point of the substance.
2. Determination of the solidifying point of ointments

Apparatus: rotating thermometer (Figure 9.2)
Figure 9.2: Determination of the solidification point of ointments with a rotating thermometer
Insert the thermometer through a one-bore cork into a 100-mL, wide-necked conical flask,
which serves as an air-bath. Heat the sample on the water-bath, while stirring, to a
temperature 8-10 ºC higher than the expected solidification point. Heat the air-bath with the
_______________________________________________________________________________
thermometer on the same water-bath and at the same temperature. When the ointment has
reached the desired temperature, remove the thermometer with the cork from the flask, and
immerse its mercury reservoir into the melted ointment so that one drop hangs on its tip.
Move the thermometer into the flask immediately and rotate it carefully together with the
flask, horizontally around its longitudinal axis at a speed of about 2 seconds per rotation. The
solidification point of the sample is temperature at which the drop on the mercury reservoir
makes the first rotation together with the thermometer (the ointment is solidified). Calculate
the average of 3 parallel measurements.
3. Determination of the oil number of ointments (Figure 9.3)
Figure 9.3: Determination of the oil number of ointments
Melt the water-free ointment base, fill it into a glass vessel and let it solidify. Put the vessel
on a 10 x 10 cm filter paper, with its broader mouth downwards. Place the sample pretreated
in this way on a watch-glass and let it stand at 36–37 ºC for 2 hours. After this period,
measure the smallest and largest diameters (mm) of the elliptical fatty stain on the paper. The
arithmetical average of these two values is considered to be characteristic of the oil number.
Calculate the average of 3 parallel measurements.
Attention: start the work with this determination.
_______________________________________________________________________________
4. Investigation of water absorbing capacity
Measure 10.00 g of ointment base with cg accuracy into a tarred metal pot with pestle and
heat it to 35–40 ºC. Add 1.0 g of water of the same temperature. Add more water in small
portions (0.5–1.0 g) emulsifying each portion before a further amount of water is added until
no more water is ready to emulsify. Decant the excess of water and blot the water droplets
cautiously from the surface of the ointment and the pot with filter paper. Determine the
weight of the ointment with 0.01 g accuracy and calculate the water absorbing capacity by
using the following equation. The w/w % water content is the water absorbing capacity of the
ointment. Calculate the average of 3 parallel measurements.
b a
Vf = 100(%) (1)
b
where Vf is the absorbed amount of water, a is the weight of the ointment, b is the weight of
the ointment and the absorbed water together.
_______________________________________________________________________________
Report
9. Investigation of ointments I. – Investigation of drop point, solidifying point, oil
number and water absorbing capacity
Name: ..................................................... Group: ....................... Date: ............................
1. Fill in the table. Calculate the average of the measurements. Evaluate the results.
Name of ointment 1: ................................................................

Name of ointment 2: ................................................................
Name of ointment 3: ................................................................
Investigation Ointment 1. Ointment 2 Ointment 3

1.
2.
Drop point (°C)
3.
average
1.
2.
Solidifying point (°C)
3.
average
1.
2.
Oil number (mm)
3.
average
1.
Water absorbing 2.
capacity (%) 3.
average
Evaluation:
_______________________________________________________________________________
10. Investigation of ointments II. – Investigation of the washability

and rheological characteristics of ointments
Introduction
In the case of ointments and creams applied on the skin surface, it is very important to
know to what extent the ointment layer administered onto the skin surface is able to stay on
the skin surface when coming into contact with water. This information is particularly
important in the case of dermoprotective ointments. The aim of this investigation, besides
determining the rheological parameters, is to model to what extent ointments and creams
serve their application purposes concerning washability.
Tasks
1. Determine the washability coefficient of the creams received.
Apply a thin layer from the cream to be examined onto a plastic plate with a given depth of
notch (100 x 50 x 1 mm). Continuously pour 50 mL of distilled water dropwise from a burette
onto the surface of the prepared ointment. After dropping is finished, dry the plastic plate and
place it on a graph paper. Specify how many mm2 of ointment area is washed off by 50 mL of
distilled water. Knowing this, calculate the value of the washability coefficient (K) from the
results of 3 parallel measurements by using the following formula:
F
K= (1)
V
where F is the area of the ointment film washed off in mm2, V is the used water volume in mL
(Figure10.1).
_______________________________________________________________________________
Figure 10.1: Determination of the washability coefficient
2. Determine with a rheometer the flow curves (0.1-100 1/s) of Unguentum emulsificans
anionicum with three different water content values (10-80 %) and plot the flow curves.
Include the results of the measurements in separate tables.
Fill the measuring device with the ointment to be examined. Measure the shear stress with
increasing and decreasing shear rate (0.1-100 1/s) at 12 points. Plot the flow curves based on
the measurements.
_______________________________________________________________________________
Report
10. Investigation of ointments II. – Investigation of the washability and rheological
characteristics of ointments
Name: ..................................................... Group: ....................... Date: ............................
1. Determine the washability coefficients of the creams received and fill in the table.
Measurement
Cream 1: ..................................................... Cream 2: .....................................................
1.
2.
3.
average
2. Rheological measurements of Unguentum emulsificans anionicum ointments with

different water content. Indicate the water content of the ointment.
Water content: ...........................................
Measuring D I „up” I „down”

B „up” 2
B „down”
degree (1/s) (N/m ) (N/m2)
_______________________________________________________________________________
Water content: ...........................................

B „up” 2
B „down”
Water content: ...........................................

B „up” 2
B „down”
_______________________________________________________________________________
3. Plot the flow curves of Unguentum emulsificans anionicum with different water
content (b–D) and evaluate the rheograms.
Evaluation:
1. Connection between the washability coefficient and the application purpose of the
creams.
2. Evaluation of the rheograms.

_______________________________________________________________________________
11. Investigation of ointments III. – Characterisation of

consistency by viscosity, spreadability and adhesion investigations
Introduction
It is very important to know the consistency of ointments both from the point of view of
the manufacturing process and the field of application. Besides viscosity measurements, the
spreadability and the adhesion of ointments give information about their applicability on the
skin.
Tasks
1. Viscosity measurements
Apparatus: rotational viscometer
Fill the measuring device with the ointment to be examined. Measure the shear stress with
increasing and decreasing shear rate (0.1-100 1/s) at 12 points. Plot the flow and viscosity
curves based on the measurements.
2. Characterization of the spreadability of ointments

Apparatus: extensometer
Put 1.0 g of ointment on a glass plate in a circle of 1 cm radius. Place the glass plate on a
graph paper and cover it with a glass plate of the same size. After 1 minute, read the two
perpendicular diameters of the ointment spot with the help of the graph paper. Place weights
of 10, 20, 50, 100, 200 and 500 g in the centre of ointment spot, and after 1 minute of loading
determine the diameters of the ointment spot in the same way as mentioned above. Investigate
_______________________________________________________________________________
two different ointments with three parallel measurements (Figure 11.1). Calculate the area of
the ointment spots with the average of diameters.
Figure 11.1: Investigation of the spreadability of ointments
Characterisation of spreadability:
d2
A= (1)
4
where A is the area of the ointment spot, d is the average of the perpendicular diameters of the
ointment spot.
3. Investigation of the adhesion of ointments

Apparatus: instrument for adhesion investigation with stand
Spread 0.1 g of ointment in the middle of a glass plate and cover it with another one.
Remove the excess of ointment obtained by pressing the covered plate with 1 kg of weight for
1 minute to form a uniform film from the ointment. Place the sample on the stand and fix the
lower plate. Put the given weights (1-30 g) in the pan and measure the time required for the
upper plate to stop (Figure 11.2). Carry out 5 parallel measurements.
_______________________________________________________________________________
Figure 11.2: Investigation of the adhesion of ointments
Two of the following ointments are investigated:

I. Vaselinum album
II. Pasta zinci oxydata
III. Unguentum emulsificans nonionicum
IV. Unguentum hydrophylicum nonionicum
_______________________________________________________________________________
Report
11. Investigation of ointments III. – Characterisation of consistency by viscosity,
spreadability and adhesion investigations
Name: ..................................................... Group: ....................... Date: ............................
Name of the investigated preparations
Ointment 1: .........................................................
Ointment 2: .........................................................
1. Results of rheological measurements
Ointment 1
Measuring D B „up” I „up” C „up” B „down” I „down” C „down”

degree (1/s) (N/m2) (Pa·s) (N/m2) (Pa·s)
_______________________________________________________________________________
Ointment 2
Measuring D B „up” I „up” C „up” B „down” I „down” C „down”

2 2
degree (1/s) (N/m ) (Pa·s) (N/m ) (Pa·s)
2. Fill the spreadability results in the table.
Weight (g)
Composition Spreadability
0 10 20 50 100 200 500
1.
2.
Ointment 1 3.
d average (cm)
surface (cm2)
1.
2.
Ointment 2 3.
d average (cm)
surface (cm2)
_______________________________________________________________________________
3. Fill the results of the adhesion investigations in the table. Calculate the standard
deviation and the relative standard deviation according to the parallel measurements. If
the measured time is longer than 180 s, consider it infinite.
Weight (g)
Composition Time
(s) 1 5 10 15 20 25 30
1.
2.
3.
4.
Ointment 1 5.
average
deviation
relative
deviation
1.
2.
3.
4.
Ointment 2 5.
average
deviation
relative
deviation
_______________________________________________________________________________
4. Plot the flow curves (b–D) and the viscosity curves (c–D) of the two investigated
ointments.
Flow curves
_______________________________________________________________________________
Viscosity curves
5. What are the flow types of ointments?
Flow type of ointment 1: ...........................................................................

Flow type of ointment 2: ...........................................................................
_______________________________________________________________________________
12. Investigation of the water uptake of tablets
Introduction
Affinity to the solvent – which can be described quantitatively by the amount of water
taken up by the polymer and by the rate of water uptake – is an important preliminary
examination with respect to the technological usability of macromolecules during granulation
and pelleting. This property is also important in the disintegration processes of tablets and
granules. From the technological point of view, the ideal polymer swells rapidly and is able to
take up great quantities of water.
The so-called Enslin number is determined for the measurement and the quantitative
description of the affinity of tablets to fluids. The Enslin number gives the amount of fluid
absorbed by 1 g of solid material, powder or tablet, in a given time. This procedure is not
official either in the Hungarian pharmacopoeia or in foreign pharmacopoeias, yet it is an
important part of preformulation studies.
The following empirical relationship was established between the volume of the fluid
absorbed by the tablet (V) and the time of the examination (t):
V = V0 + m t (1)
where V0 is the volume of the fluid absorbed at time 0. If the quantity of the absorbed water is
plotted against the square root of time, a straight line is obtained. The slope of the line (m)
gives the rate constant of water uptake.
Task
1. Examine the water uptake of the two tablets with different compositions chosen by your
practice leader. Include the data in a table.
Apparatus
The apparatus presented in the figure is used for determining water uptake (Figure 12.1).
The apparatus consists of the following parts: a glass filter with appropriate pore size, in
which the material to be examined is placed; a pipette connected to the filter, which makes it
_______________________________________________________________________________
possible to quantitatively measure the amount of water absorbed by the tablet through the
filter. The amount of water absorbed depends mainly on the affinity of the examined material
to water, but naturally, the result is also influenced by the construction of the apparatus used
in the experiment. For example, the pore size of the filter, the position of the pipette as well as
the mass and diameter of the tablet placed on the filter all influence the results obtained.
Under the same test conditions, the data are well comparable and can be used for the
quantitative description of the water uptake of tablets with different compositions.
Figure 12.1: Apparatus for determining the Enslin number

(1 – tablet; 2 – filter paper; 3 – G filter; 4 – pipette)
Assemble the Enslin apparatus. Fill the pipette – through the filter – with the test fluid
(unless otherwise prescribed, with distilled water) and place it so that the glass filter and the
pipette will be in the same plane. Get a filter paper slightly larger in size than the area of the
glass filter and place it on its surface without leaving gaps. Set the fluid level to zero. Then
place the tablet to be examined on the filter paper concentrically and immediately start
measuring the time.
During the task, take care of the following:
– fill the pipette in a bubble-free way;
– make sure that the filter paper is placed tightly on the glass filter.
Measure the amount of the absorbed water after 10, 20, 30 seconds and 1, 2, 3, 4, 5, 6, 8
and 10 minutes. After finishing the examination, remove the solid material together with the
filter paper. Perform three parallel measurements.
_______________________________________________________________________________
2. Calculate the Enslin number of the tablets from the average.
3. Plot the V = f (t ) and V = f ( t ) functions. From the latter one, calculate the rate constant
of the process (m), the slope of the function.
4. Compare the affinity of the tablets with different compositions to water. Explain the
difference between the Enslin numbers of the tablets.
_______________________________________________________________________________
Report
12. Investigation of the water uptake of tablets
Name: ..................................................... Group: ....................... Date: ............................
1. Examine the water uptake of the two tablets with different compositions chosen by your
practice leader. Include the data in a table.
2. Calculate the Enslin number of the tablets from the average.
Name of tablet 1: .................................................. Average mass: ............................(g)

Name of tablet 2: .................................................. Average mass: ............................(g)
Sample Time (s) 10 20 30 60 120 180 240 300 360 480 600
1. (mL)
2. (mL)
3. (mL)
1.
average (mL)
Enslin
number
1. (mL)
2. (mL)
3. (mL)
2.
average (mL)
Enslin
number
_______________________________________________________________________________
3. Plot the V = f (t ) function.

_______________________________________________________________________________
3. Plot the V = f ( t ) function. Calculate the rate constant of the process (m), the slope of
the function.
Sample m
Tablet 1
Tablet 2
4. Compare the affinity of the tablets with different compositions to water. Explain the
difference between the Enslin numbers of the tablets.
_______________________________________________________________________________
13. Investigation of tablets
Introduction
In the course of tablet formulation, the dosage form must satisfy the requirements specified
in the technological documentation. These are partly the examinations prescribed in Ph. Hg.
VIII, but there may also be other examinations in the documentation. Such is, e.g., the
checking of the geometric parameters of the tablets, which is important for the setting of
automatic packaging machines. So the purpose of these tests is to evaluate the tablets on the
basis of some important examinations.
The Hungarian Pharmacopoeia contains the following requirements about the uniformity
of the mass of coated and uncoated tablets (Ph. Hg. VIII., 2.9.5.):
Tablet mass Accepted tolerances

< 80 mg ± 10.0 %
80 – 250 mg ± 7.5 %
> 250 mg ± 5.0 %
Only 2 individual masses can differ to a greater extent, and the deviation from the average
value cannot be more than two times the given value for any of the individual masses.
According to the Hungarian Pharmacopoeia, tablets must also comply with the
requirements of the uniformity of dosage units described in the examination (Ph. Hg. VIII.,
2.9.40.). The term “uniformity of dosage units” is defined as the degree of uniformity in the
amount of the active substance among dosage units, which can be demonstrated by either of
two methods: content uniformity or mass variation. In order to determine content
uniformity, the contents of ten units are assayed individually using an appropriate analytical
method. To examine mass variation, ten individual tablets are measured precisely. From the
mass of the individual tablets and from the result of the content assay, the active substance
content of the tablets is calculated individually, expressed as the percentage of the label claim.
The test for mass variation is applicable for tablets containing 25 mg or more of an active
substance comprising 25 % or more, by mass, of the dosage unit. The content uniformity
_______________________________________________________________________________
method may be applied in all cases. The requirements of the uniformity of dosage units are
satisfied if the acceptance value calculated from the first ten dosage units is L1 or lower. If it
is higher, another twenty dosage units are tested and the acceptance value is calculated again.
The requirements are satisfied if the final acceptance value calculated for thirty dosage units
is L1 or lower, and if during the calculation of the acceptance values of content uniformity
and mass variation no individual content lower than (1-L2·0.01) M or higher than
(1+L2·0.01) M is found among the dosage units. Unless otherwise prescribed, L1 = 15.0 and
L2 = 25.0.
Tasks
1. Determine the geometric parameters of 20 tablets (diameter, height) with a micrometer

screw. Calculate the average and the relative standard deviation.
2. Perform the mass uniformity test according to the Hungarian Pharmacopoeia (Ph. Hg.
VIII., 2.9.5.). Calculate the average, the standard deviation, the relative standard
deviation and evaluate the results according to the specifications of the Hungarian
Pharmacopoeia.
Measure the mass of each of twenty tablets selected at random. Use an analytical balance
for the measurement.
3. Perform the test of the uniformity of dosage units (Ph. Hg. VIII., 2.9.40.) by using the
method of mass variation, as prescribed by the Hungarian Pharmacopoeia. Evaluate the
tablets according to the specifications of the Hungarian Pharmacopoeia.
In the first step, determine the active substance content of the tablets. Powder 10 tablets in
a mortar and homogenize the powder. Measure an amount of the powdered sample equalling
the mass of 1 tablet with analytical precision and spread it into the flask which contains the
_______________________________________________________________________________
required quantity of dissolution medium. Mix the contents of the flask from time to time, then
30 minutes later take a sample, filter it on a filter paper and measure the active substance
content of the sample spectrophotometrically. Dilute the sampled aliquot quantity with a
filtered dissolution medium. Calculate the active substance content in relation to a tablet with
average mass. Perform three parallel measurements.
Calculate the active substance content of the individual tablets from the individual mass
and from the result of the average active substance content of the first 10 of the tablets
measured in point 2, expressed as the percentage of the label claim, then calculate the
acceptance value. The calculation of the acceptance value (AV) is the following:
AV = M X +k s (1)
where M is the reference value, X is the average value of individual contents as the percentage
of the label claim, k is the acceptance constant (in case of n = 10, its value is 2.4), s is the
standard deviation of the sample.
if 98.5 % g X g 101.5 %, then M = X, AV = k · s

if X < 98.5%, then M = 98.5 %, AV = 98.5 -X + k · s
if X > 101.5%, then M = 101.5 %, AV = X – 101.5 + k · s
Tablets to be examined
Name of tablet Dissolution medium Spectrophotometric data

Tabletta coffeini artificial gastric juice R=277 nm; 0.100 A = 2.0641 µg/mL
Tabletta sulfadimidini artificial gastric juice R=244 nm; 0.100 A = 1.9001 µg/mL
Tabletta paracetamoli artificial intestinal juice R=243 nm; 0.100 A = 1.6260 µg/mL
_______________________________________________________________________________
Report
13. Investigation of tablets
Name: ..................................................... Group: ....................... Date: ............................
1. Determine the geometric parameters of 20 tablets (diameter, height) with a micrometer

screw. Calculate the average and the relative standard deviation.
2. Perform the mass uniformity test according to the Hungarian Pharmacopoeia (Ph. Hg.
VIII., 2.9.5.). Calculate the average, the standard deviation, the relative standard
deviation and evaluate the results according to the specifications of the Hungarian
Pharmacopoeia.
Name of tablet: .......................................................
Tablet Diameter (mm) Height(mm) Mass (g)
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
11.
12.
13.
14.
15.
16.
17.
18.
19.
20.
average
minimum
maximum
standard deviation
relative standard deviation
_______________________________________________________________________________
Evaluation of mass uniformity:
3. Perform the test of the uniformity of dosage units (Ph. Hg. VIII., 2.9.40.) by using the
method of mass variation, as prescribed by the Hungarian Pharmacopoeia. Evaluate the
tablets according to the specifications of the Hungarian Pharmacopoeia.
Measurement A Average active substance content (mg)

1.
2.
3.
average
standard deviation
Tablet Mass (g) Active substance content (mg) Active substance content (%)
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
average
standard deviation
Evaluation of the uniformity of dosage units:

_______________________________________________________________________________
14. Investigation of the disintegration of rectal and vaginal

suppositories
Introduction
The suppository bases and excipients used in manufacturing can significantly modify drug
release, consequently the effect caused by the suppository. The suitable choice of the
suppository base is an important factor in achieving the desired effect produced by the
suppository and in determining the disintegration time of the dosage form. The aim of this
investigation is to determine whether the suppositories disintegrate within the prescribed time
when placed in a liquid medium in the experimental conditions (Ph. Hg VIII. 2.9.2).
Task
In vitro investigation of the disintegration time of suppositories and whether they satisfy
the requirements of the Pharmacopoeia.
Apparatus: Apparatus for the disintegration of suppositories (Figure 14.1)
Figure 14.1: Sample holder of the apparatus for the disintegration of suppositories
_______________________________________________________________________________
Apparatus description
The apparatus consists of a sleeve of glass or suitable transparent plastic, of appropriate

thickness, to the interior of which is attached by means of three hooks a metal device
consisting of two perforated stainless metal discs, each containing 39 holes 4 mm in diameter;
the diameter of the discs is similar to that of the interior of the sleeve; the discs are about
30 mm apart. The test is carried out using three such apparatuses, each containing a single
sample. Each apparatus is placed in a beaker filled with water maintained at 36-37 C. The
apparatuses are placed together in a water bath. The beaker is fitted with a slow stirrer and a
device that will hold the cylinders vertically not less than 90 mm below the surface of the
water and allow them to be inverted without emerging from the water.
1. Switch on the thermostat 20-30 minutes before starting the investigation. Set the knob
on the right side to VAR position, and the one on the left side to the desired
temperature.
2. Measure 3.00 kg of distilled water into 3 glass beakers (3,000 mL) and place them into
the apparatus. Wait until the desired temperature is reached in the acceptor phase (in the
beaker 37 ± 0.5 °C)
3. Switch on the Timer knob. The duration of the examination is 60 min.
4. Switch the knob to ALAP position, the rotation speed will be 1 turn/min.
5. Place the suppositories into the metal baskets and fix them to the rotating device.
6. Press the START knob.
The measurement is started at the moment the suppositories are immersed in the water and
lasts until complete disintegration.
Carry out two parallel measurements and calculate the average of the suppositories of the
same composition. Record the results in the table and evaluate them according to the
requirements of the Pharmacopoeia.
_______________________________________________________________________________
Report
14. Investigation of the disintegration of rectal and vaginal suppositories
Name: ..................................................... Group: ....................... Date: ............................
1. Investigation of the disintegration of rectal and vaginal suppositories
Sample name,
mark ........................................ ........................................ ........................................
Disintegration time min min min
1.
2.
average
conclusion
Evaluation:
_______________________________________________________________________________
III. Investigation of stability

_______________________________________________________________________________
15. Determination of the decomposition (caramelization) of

glucose solution
Introduction
Sugars (glucose, fructose, sucrose, etc.) are known to decompose due to heat (preparation
of syrups, sterilization of infusions containing sugar). During chemical
decomposition/oxidation, yellowish-brownish substances with bitter a taste (so-called
humins) are formed. During the preparation of syrups, local overheating can be prevented by
using a duplicator, thereby preventing the degradation of sugar. In case of infusions, the
solution can be sterilized without degradation if the pH of the glucose infusion is adjusted to
3-4.
Spectrophotometry is a well suitable analytical method for the detection of decomposition
products characterized by light absorption. The purpose of this practice is to investigate
spectrophotometrically the degradation of glucose solutions with different pH values during
sterilization (121 °C, autoclaved for 20 minutes).
Task
Adjust the pH of glucose solutions to different pH values by using a buffer. Determine the
changes occurring during sterilization after appropriate dilution with spectrophotometry.
1. Preparation of buffer stock solutions
Preparation of buffer stock solutions according to Mc Ilvaine. The buffers are prepared by
mixing two buffer stock solutions.
Stock solution Volume Mt

0.2 M Na2HPO4 100 mL (Na2HPO4 12 H20) = 358.14 (Na2HPO4 2 H20) = 177.99
0.1 M citric acid 100 mL (C6H8O 7 H2O) = 210.14
_______________________________________________________________________________
2. Preparation of glucose solutions using different buffers
Dissolve 5 g of glucose in distilled water of a volume of ~70 mL each time, then pipette
the following volumes from the buffer stock solutions into volumetric flasks of 100 mL to
reach the appropriate pH:
0.2 M Na2HPO4 0.1 M citric acid

pH
(mL) (mL)
3.0 4.11 15.89
4.0 7.71 12.29
5.0 9.00 11.00
6.0 12.63 7.37
7.0 16.47 3.53
8.0 19.45 0.55
Complete the flasks with distilled water, then homogenize them. Pour the contents into
infusion bottles, close and sign them properly. The infusions should be sterilized in autoclave
at 121 °C for 20 minutes according to the Hungarian Pharmacopoeia (it is done by assistants)
(Figure 15.1).
Figure 15.1: Colour changes of glucose solutions after sterilization between pH 3-8
_______________________________________________________________________________
3. After appropriate dilution, determine the absorbance of the solutions at the given
wavelength.
The absorbance of the solutions prepared and sterilized the previous week should be
investigated at the absorption maximum of the formed decomposition products, at a
wavelength of 281 nm, against distilled water. If it is necessary, make a dilution from the
solution to be measured, using distilled water.
4. Plotting and evaluation of graph
Plot the results against the pH values.

_______________________________________________________________________________
Report
15. Determination of the decomposition (caramelization) of glucose solution
Name: ..................................................... Group: ....................... Date: ............................
1. Prepare the following stock solutions of buffer solution in a volume of 100 mL each,
then indicate the amount of the substances to be measured in the appropriate place.
The amount of substance needed for the preparation of 100 mL of 0.2 M Na2HPO4 solution:
.......................... g
Indicate which crystal water-containing substance was used. Mt = .....................................
The amount of substance needed for the preparation of 100 mL of 0.1 M citric acid solution:
.......................... g
2. Prepare the 100-100 mL of glucose solution concentration of 5 % using buffer solutions

having different pH-values, and pour them into infusion bottles of 100 mL.
3. Measure the absorbance of the sterilized glucose solutions with a spectrophotometer, at

a wavelength of 281 nm against distilled water. If it is necessary, prepare dilutions to
volume percent with the absorbance value measured between 0.3 and 0.8. Calculate the
product of absorbance and dilution. Include the results obtained in the table below.
pH A dilution A · dilution
3
4
5
6
7
8
_______________________________________________________________________________
4. Plot the results in the last column of the table against the pH (A · dilution).
Evaluate the results obtained. Which pH range can ensure the proper stability of glucose?
Evaluation according to the colour intensity:
Evaluation according to the data:

_______________________________________________________________________________
16. Investigation of the viscosity changes of hydrophilic sols I.
Introduction
Methylcellulose (MC) is a commonly used gel- and sol-forming additive, which dissolves
in cold water colloidally. Its aqueous solution is opalescent because of the not sufficiently
hydrated polymer fibres. Exposure to heat results in a reversible coagulation, which can be
converted back by cooling. Similarly, the reversible coagulation of MC occurs due to larger
amounts of electrolytes (e.g. iodide, chloride and nitrate salts).
Hydroxyethyl cellulose (HEC) is the monoglycol ether of cellulose in its main mass. It is
specifically hydrophilic, therefore it dissolves in water colloidally at a low degree of
substitution. It solvates quickly and completely, the colloid solution obtained is clear and does
not precipitate even when exposed to heat. HEC is used for bonded granulation in the
pharmaceutical industry. The basic compound and especially its derivatives are also used for
film coating. Its colloid solution is one of the most commonly used viscosity enhancing
excipients during the production of suspensions and emulsions. It is incompatible with the
concentrated solutions of inorganic salts and with tannates.
Carmellose sodium (also known as carboxymethyl cellulose sodium, CMCNa) is the
sodium salt of a partially O-carboxymethylated cellulose. In the 8th Edition of the Hungarian
Pharmacopoeia Carmellosum natricum substitutum humile (low-substituted carmellose
sodium) is also official, which may contain short fibrils. The calcium salt of partially O-
carboxymethylated cellulose is also official. It swells in water and becomes opalescent, it
forms a viscous colloidal solution. It is primarily used as a viscosity enhancing excipient
(stabilizer of emulsions and suspensions), as a hydrogel in different concentrations and as a
disintegrant in tablets. It is incompatible with different salts, acidic medium and cationic
surfactants.
These polymers are usually used for preparing colloidal solutions. The resulting gel
structure is very sensitive even to slight changes in the environmental parameters (e.g. pH,
temperature, ion concentration), which is also reflected in the changes of the physical-
chemical properties (e.g. shape, viscosity).
The purpose of the task is to study the viscosity changes of one of the mucilages
(methylcellulose/hydroxyethyl cellulose/carboxymethyl cellulose) due to the effect of sodium
_______________________________________________________________________________
chloride (investigation of a NaCl-free composition and two NaCl compositions containing

different ingredients).
Task
Apparatus: Höppler-type viscometer (Figure 16.1)
Figure 16.1: Höppler-type viscometer
Dissolve an appropriate amount of sodium chloride in the given type of mucilage so that
the total weight will be 100 g and the product will have the given sodium chloride
concentration. Add the salt to the mucilage in portions and apply intensive stirring.
After dissolution, determine the apparent viscosity of the samples with the Höppler-type
viscometer. The salt-free mucilage serves as the basis for comparison. Plot the calculated
_______________________________________________________________________________
viscosity values against the load. Make 3 parallel measurements and always use a new sample
for each measurement.
Process of viscosity measurement
Fill the measuring cup with the colloidal solution to be tested in a bubble-free way. Choose
the ball rod and mount it onto the measuring arm. Make the device ready for measurement.
(levelling, offsetting the buoyancy with sliding weight). Put the smallest weight on the weight
holder plate, release the locking screw, measure the time needed to cover the distance
corresponding to the 0-30 scale. After that, remove the weight, pull back the ball rod carefully
and fix the measuring arm with the eccentric. Wait for a short time and repeat the
measurement with greater load. You can increase the load as long as the descent time is
higher than 4 seconds.
The equation of the calculation of viscosity:

=k p t (1)
where 2 is the apparent viscosity (mPa·s), (Pa = 0.01 g/cm2), k is the instrument constant
(cuvette labelled 01, k = 0.1053), p is the load, shear stress (p = 10, 20, 30, 40 and 50 g/cm2), t
is the descent time (s).
_______________________________________________________________________________
Report
16. Investigation of the viscosity changes of hydrophilic sols I.
Name: ..................................................... Group: ....................... Date: ............................
1. Name of mucilage: ........................................................
Load (g/cm2) 10 20 30 40 50
NaCl conc. Measurement Descent time (s)
1.
2.
0%
3.
Average
1.
2.
....... %
3.
Average
1.
2.
....... %
3.
Average
NaCl conc.
0% ......... % ......... %
(%)
Load t C t C t C
2
(g/cm ) (s) (mPa·s) (s) (mPa·s) (s) (mPa·s)
10
20
30
40
50
2. Plot the viscosity against the load.

_______________________________________________________________________________
3. Evaluation:
Write down what you experienced and find a correlation between the concentration of
sodium chloride and the viscosity of the mucilages based on the results. What kind of
colloidal physical phenomenon is it? Give reasons for your answer.
_______________________________________________________________________________
17. Investigation of the viscosity changes of hydrophilic sols II.
Introduction
Carmellose sodium (also known as carboxymethyl cellulose sodium, CMCNa) is the

sodium salt of a partially O-carboxymethylated cellulose. In the 8th Edition of the Hungarian
Pharmacopoeia Carmellosum natricum substitutum humile (low-substituted carmellose
sodium) is also official, which may contain short fibrils. The calcium salt of partially O-
carboxymethylated cellulose is also official. It swells in water and becomes opalescent, it
forms a viscous colloidal solution. Its application is diverse: for internal use it can be used
only in the form of a colloidal aqueous solution. It is beneficial as a coating agent in the case
of gastro-intestinal disorders. As it can adsorb several substances, it can also be used as an
antidiarrheal drug. It also has an antacid effect, and it is used as a food additive as well (E
466).
In pharmaceutical technology, it is used as a binder agent during the production of
granules. In tablets, Carmellosum natricum conexum (croscarmellose sodium) containing
cross-links can be applied as a disintegrant and it has good swelling properties. In different
concentrations it can also be used as a hydrogel, as the stabilizer of emulsions and
suspensions, furthermore, as a protective colloid, too. It is incompatible with acidic medium,
heavy metal salts, cationic surfactants, alkaloid salts and phenols.
The purpose of the task is to present an incompatibility: to study the viscosity change of 3
% of carmellose sodium mucilage due to the effect of hydrochloric acid.
Task
Apparatus: Höppler-type viscometer
Determine the apparent viscosity of the different available – stored for 24 hours –
carmellose sodium mucilages containing hydrochloric acid of 2 M (1, 2, 3 %) with the
Höppler-type viscometer. The hydrochloric acid-free mucilage serves as the basis for
_______________________________________________________________________________
comparison. Plot the calculated viscosity values against the load. Make 3 parallel
measurements and always use a new sample for each measurement.
Process of viscosity measurement
Fill the measuring cup with the colloidal solution to be tested carefully, in a bubble-free
way. Choose the ball rod and mount it onto the measuring arm. Bring the device state ready to
measure. (levelling, offsetting the buoyancy with sliding weight). Put the smallest weight on
the weight holder plate, release the locking screw, measure the time needed to cover the
distance corresponding to the 0-30 scale. After that, remove the weight, pull back the ball rod
carefully and fix the measuring arm with the eccentric. Wait for a short time and repeat the
measurement with greater load. You can increase the load as long as the descent time is
higher than 4 seconds.
The equation of the calculation of viscosity:

=k p t (1)
where 2 is the apparent viscosity (mPa·s), (Pa = 0.01 g/cm2), k is the instrument constant
(cuvette labelled 01, k = 0.1053), p is the load, shear stress (p = 10, 20, 30, 40 and 50 g/cm2), t
is the descent time (s).
_______________________________________________________________________________
Report
17. Investigation of the viscosity changes of hydrophilic sols II.
Name: ..................................................... Group: ....................... Date: ............................
1. Results of the rheological measurements:
Load (g/cm2) 10 20 30 40
HCl conc. Measurement Descent time (s)
1.
2.
0%
3.
Average
1.
2.
1%
3.
Average
1.
2.
2%
3.
Average
1.
2.
3%
3.
Average
HCl conc.
0 1 2 3
(%)
Load t C t C t C t C
(s) (mPa·s) (s) (mPa·s) (s) (mPa·s) (s) (mPa·s)
10
20
30
40
_______________________________________________________________________________
2. Plot the viscosity against the load.
3. Evaluation:
Write down what you experienced and find a correlation between the concentration of
hydrochloric acid and the viscosity of the mucilages based on the results. What kind of
colloidal physical phenomenon is it? Give reasons for your answer.
_______________________________________________________________________________
18. Influence of humidity on the geometric parameters of tablets
Introduction
Humidity in the air may affect the stability, the physical parameters of tablets, such as their
mass, geometric parameters, mechanical strength, disintegration and also active substance
release. The geometric parameters of tablets (diameter, height) must be within the threshold
limits prescribed in the detailed specifications of the given preparation. The uniformity of size
and shape has great significance when packaging with automatic packaging machines.
The reason for the effect of humidity lies in the hygroscopicity and water uptake capacity
(Enslin number) of the excipients and active substances of the tablets. For example, excipients
used as disintegrants or superdisintegrants, such as starch and sodium starch glycolate, have a
high Enslin number. Because of this, these tablets must be stored carefully away from air, or
with the use of a moisture absorber.
The aim of this investigation is to examine how 100% relative humidity influences the
mass and the geometric parameters of tablets containing various excipients.
Task
1. Measure the mass, diameter and height of the disintegrant- and superdisintegrant-
containing tablets given by your practice leader. Measure the mass with an analytical
balance and the geometric parameters with a micrometer screw.
2. Measure the above parameters also for tablets stored at room temperature, under normal
circumstances and for tablets stored for different periods of time at room temperature at
100% relative humidity (storage time: 24, 48, 72, 96 hours). Perform 10 parallel
measurements for each. Include your results in a table. Calculate the averages, the mass
and volume changes.
3. Plot the volume change against time.

_______________________________________________________________________________
Report
18. Influence of humidity on the geometric parameters of tablets
Name: ..................................................... Group: ....................... Date: ............................
1. Measure the mass, diameter and height of the disintegrant- and superdisintegrant-
containing tablets given by your practice leader. Measure the mass with an analytical
balance and the geometric parameters with a micrometer screw.
2. Measure the above parameters also for tablets stored at room temperature, under normal
circumstances and for tablets stored for different periods of time at room temperature at
100% relative humidity (storage time: 24, 48, 72, 96 hours). Perform 10 parallel
measurements for each. Include your results in a table. Calculate the averages, the mass
and volume changes.
Tablet 1: ..................................................................
Mass (mg)
Tablet 1
0 hours 24 hours 48 hours 72 hours 96 hours
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
Diameter (mm) Height (mm)

Tablet 1 0 24 48 72 96 0 24 48 72 96
hours hours hours hours hours hours hours hours hours hours
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
_______________________________________________________________________________
Volume (mm3)
Tablet 1
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
Average values of the tested parameters of Tablet 1:
Average Average Average Mass change Volume

Storage time Average
diameter height (mm) volume (mg) change
(hours) mass (mg)
(mm) (mm3) (mm3)
0
24
48
72
96
Tablet 2: ..................................................................
Mass (mg)
Tablet 2
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
_______________________________________________________________________________
Diameter (mm) Height (mm)

Tablet 2
0 24 48 72 96 0 24 48 72 96
hours hours hours hours hours hours hours hours hours hours
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
Volume (mm3)
Tablet 2
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
Average values of the tested parameters of Tablet 2:
Average Average Average Mass change Volume

Storage time Average
diameter height (mm) volume (mg) change
(hours) mass (mg)
(mm) (mm3) (mm3)
0
24
48
72
96
_______________________________________________________________________________
3. Plot the volume change against time.
Evaluation:
_______________________________________________________________________________
19. Investigation of the stability of tablets containing

acetylsalicylic acid I.
using (real time) long-term stability test
Introduction
The suitable stability of the dosage form provides the efficacy of the medicine up to the
expiration date. A basic requirement is that neither the effect, nor the side effect of the
medicine is allowed to change. The drug content may change between the values prescribed
by law, the maximum decrease permitted is usually 10%.
The kinetic investigation of drug stability gives information about the time course of drug
decomposition. The main decomposition ways of drugs follow first or zero order kinetics.
In case of zero order kinetics, the reaction rate is independent of the concentration of the
reaction partners (Figure 19.1):
[Ct ] = [C0 ] k t (1)
where Ct is the concentration of the drug in time t, C0 is the initial concentration of the drug, k
is the rate constant.
Figure 19.1: Decomposition of drug following zero order kinetics

_______________________________________________________________________________
According to the above, the reaction rate is constant during the reaction.
d [C ]
v= = k0 (2)
dt
On the basis of equation (1), in case of zero order kinetics, the values of t1/2 (half-time) and
t10% (shelf life) can be calculated as follows:
t1 / 2 =
[C0 ] (3)
2k0
t1 / 2
t10% = (4)
5
In case of first order kinetics, the reaction rate depends on the concentration of the
reaction partners (Figure 19.2):
[Ct ] = [C0 ]e k t 1
(5)
d [C ]
v= = k [C ] 1 (6)
dt
On the basis of equation (5), in case of first order kinetics, the values of t1/2 and t10% can be
calculated as follows:
ln 2 0.693
t1 / 2 = = (7)
k1 k1
2.303 100
t10% = lg (8)
k1 90
Figure 19.2: Decomposition of drug following first order kinetics

_______________________________________________________________________________
Task
The aim of the investigation is to determine the half-time and the shelf life (t1/2, t10 %) of
tablets containing acetylsalicylic acid (ASA).
Measure the ASA content of tablets stored in well-sealed containers for different periods of
time by means of spectrophotometry. Make reaction kinetics calculations on the basis of 3
parallel measurements. Determine the order of the reaction (zero or first order). Knowing the
rate constant, calculate the values of t1/2 and t10 %.
Pulverize 5 tablets from each batch of tablets stored for a given time. Weigh the amount
corresponding to 1 tablet from the pulverized sample by using an analytical balance and wash
it into a 100-mL volumetric flask with 15 mL of 96% alcohol. Dilute it with purified water,
shake it for 5 minutes and complete it to 100.00 mL with purified water. Filter the solution
immediately and prepare a dilution in the given way.
Make 3 parallel measurements, preparing 3 dilutions from each filtered solution. Measure
the absorbance of the solutions (A) at R = 276 nm. The reference solution is purified water.
On the basis of the calibration curve, 0.100 A corresponds 25.66 µg/mL of ASA.
_______________________________________________________________________________
Report
19. Investigation of the stability of tablets containing acetylsalicylic acid I.
– Investigation of decomposition kinetics, calculation of shelf life using (real time) long-
term stability test
Name: ..................................................... Group: ....................... Date: ............................
Nominal weight of 1 tablet: ................................... g

ASA content of fresh tablet: ................................... g
Tablet Storage time

number (month)
2
10
30
60
Pulverize 5 tablets from each batch of tablets stored for a given time. Weigh the amount
corresponding to 1 tablet from the pulverized sample by using an analytical balance and wash
it into a 100-mL volumetric flask with 15 mL of 96% alcohol. Dilute it with purified water,
shake it for 5 minutes and complete it to 100.00 mL with purified water. Filter the solution
immediately and dilute 5 mL of the filtered solution to 100 mL with purified water. Prepare 3
dilutions from each filtered solution.
1. Results of the spectrophotometric measurements:
Measurement 1 Measurement 2 Measurement 3 ASA

conc. conc. conc. Average content
Sample (Mg/mL) (Mg/mL) (Mg/mL) conc. in 1
A A A
(Mg/mL) tablet
(mg)
_______________________________________________________________________________
2. Calculation of reaction kinetics parameters:
sample C0 (%) Ct (%) k1 k2 t1/2 t10%
AVERAGE:
Evaluation:
_______________________________________________________________________________
20. Investigation of the stability of tablets containing

acetylsalicylic acid II.
using stress test
Introduction
Tablets are usually stored at room temperature. Investigations lasting for months or years
are often needed to determine how long they remain stable and contain the effective drug
concentration (>90%) when stored at room temperature. Stress tests can be performed in order
to shorten these investigations.
There is proportionality between the decomposition rate of the drug and the storage
temperature, which is known as the Arrhenius equation:
Ea
lg k = lg A (1)
2.303RT
where k is the rate constant of decomposition, A is the pre-exponential factor/frequency factor,
Ea is the activation energy (kJ/mol), R is the universal gas constant (8.314 J/mol·K), T is the
temperature (K).
If log k is plotted against 1/T, it will result in a linear correlation, which is called Arrhenius
plot (Figure 20.1). The slope of this line is -Ea/2.303·R, from which the activation energy can
be calculated.
Figure 20.1: Arrhenius plot

_______________________________________________________________________________
Knowing the value of the activation energy (Ea) and the rate constant (k1) at a given
temperature (T1), the rate constant (k2) at a desired temperature (T2) can be calculated and the
shelf life (t10%) can also be determined at this temperature.
k2 Ea 1 1
log = (2)
k1 2.303R T2 T1
0.105
t10%,T2 = (3)
k2
In case of first order kinetics, if the drug concentrations are known, the rate constant can be
calculated as follows:
2.303 C
k= log 0 (4)
t Ct
where C0 is the initial drug concentration, Ct is the drug concentration in time t, t is the time
(days).
Task
The aim of the investigation is to determine the half-time and the shelf life (t1/2,) of tablets
containing ASA by using stress test. The tablets were stored at different temperatures (40, 50,
60, 70, 80 °C) and the drug content was measured after 1, 2 and 3 days.
1. Calculate the average drug content of the tablets from the data at the given time and
temperature. On the basis of equation (4), calculate the rate constant of the reaction as a
first order reaction at each measuring point, by using the substitution method. Calculate
the rate constant for each temperature.
2. Draw the Arrhenius plot from the mean value of the rate constants belonging to the
given temperature. Calculate the activation energy from the slope of the plot.
3. Calculate the rate constant for 25 °C on the basis of equation (2). Knowing the rate
constant, calculate the shelf life of the tablet referring to 25 °C.
_______________________________________________________________________________
Report
20. Investigation of the stability of tablets containing acetylsalicylic acid II.
– Investigation of decomposition kinetics, calculation of shelf life using stress test
Name: ..................................................... Group: ....................... Date: ............................
1. The ASA concentrations in tablets with different storage times and temperatures are
listed below. Calculate the rate constant for each measuring point and for each
temperature.
Temp. 1/T Storage ASA concentration (%)

(T) time (t) 1 2 3 mean k kmean log kmean
(°C) (K-1) (days)
40 1 99.4 99.9 99.8
2 99.1 98.4 98.9
3 97.8 98.1 98.7
50 1 98.9 99.2 98.9
2 97.0 96.9 96.5
3 95.4 95.4 95.7
60 1 96.4 96.3 95.9
2 92.8 92.9 92.7
3 89.9 89.9 92.0
70 1 88.7 89.1 89.2
2 82.0 82.2 81.8
3 78.9 78.7 78.2
80 1 76.2 75.9 75.9
2 60.5 59.9 59.6
3 47.2 46.8 47.0
_______________________________________________________________________________
2. Arrhenius plot, determination of activation energy:
Ea = ...................................................................
3. Rate constant and shelf life of tablet referring to 25 °C:
k25°C = ...............................................................
t10%, 25°C = .........................................................
Evaluation:
_______________________________________________________________________________
IV. Biopharmaceutical investigations

_______________________________________________________________________________
21. Investigation of the drug release of emulsions by static

diffusion method
Introduction
Although the excipients used in formulation do not have an effect on their own, they can
influence the release of the API, and thereby the efficacy of the drug significantly. The type of
the emulsion is chosen on the basis of the physical-chemical properties of the API. Based on
these factors, the proper choice of the type of the emulsions and the emulsifiers applied is
very important. In the case of emulsions for external use, both the o/w and the w/o types are
used, depending on the extent of penetration into the deeper layers of the skin and on the
duration of drug release.
Polysorbate 20 is a nonionic, water-soluble surfactant, which can be used in o/w emulsions
and emulsion ointments. It increases the solubility of poorly water-soluble drugs. In water it
forms micelles belonging to the colloidal size range and it encloses the non-soluble solid and
liquid components (e.g. essential oils). It is used as a wetting agent in suspensions and
suppositories. It is applied in a concentration of 1-10 % as an emulsifier and 0.1-3 % as a
wetting agent.
Span 80 is a nonionic, practically water-insoluble surfactant, which can be used to produce
w/o type emulsions by itself. It can increase the water absorption of creams. It is also applied
as a solubilizer and a wetting agent in lipophilic medium.
The static diffusion method is a model by which the release of the drug can be determined.
It consists of three main parts: a donor and an acceptor phase, and a semipermeable dialysis
membrane separating the two phases. The donor phase contains the emulsion to be tested,
from which only the API can get through the dialysis membrane into the acceptor phase.
Concerning the latter one, the physical-chemical properties of the API should be considered
and the appropriate temperature of the phase is also an important aspect (it is not an official
method in the Hungarian Pharmacopoeia).
Because the HLB-value significantly influences drug release, during the task the rate of the
release of the API incorporated in different types of emulsions is determined by the static
diffusion method under in vitro conditions, using an emulsifier with a lower (Span 80) and a
higher (Polysorate 20) HLB-value.
_______________________________________________________________________________
Figure 21.1: Investigation of the drug release of emulsions by static diffusion
Task
The release and the membrane diffusion of lidocaine hydrochloride, as a model API, is
determined using different types of emulsions (o/w and w/o). The API is dissolved in the
aqueous phase in every case. During the investigations, the drug release of the emulsion is
measured, which means the diffusion of the API through the dialysis membrane into the
acceptor phase in such a way that the amount of lidocaine hydrochloride is determined
spectrophotometrically in the acceptor phase (Figure 21.1).
For the investigations, the students have to make the emulsions and arrange the membranes
themselves.
1. Preparation of the diffusion tests
Cut a 10-cm piece from the membrane used, soak it in water for 15 minutes and affix the
plastic closures to the membrane. The two emulsions to be prepared are investigated
_______________________________________________________________________________
simultaneously, so altogether 6 membranes should be arranged for the 3 parallel

measurements each. Put the membranes into 6 beakers of 250 mL, previously filled with 100
mL of distilled water of room temperature (~ 25 °C).
2. Preparation of the emulsions to be examined
Prepare the emulsions with the given ingredients properly by using 2 medicine bottles of
100 g.
Sample "A" weight (g)

Lidocaini hydrochloridum 5.00
Polysorbatum 20 1.00
Mucilago hydroxyaethylcellulosi 6.00
Paraffinum liquidum 20.00
Aqua purificata 18.00
Preparation: dissolve the lidocaine hydrochloride in the aqueous phase, mix the emulsifier,
then emulsify the oily phase into this in portions. Shake vigorously.
Sample "B" weight (g)

Lidocaini hydrochloridum 5.00
Span 80 1.50
Mucilago hydroxyaethylcellulosi 5.00
Paraffinum liquidum 20.00
Aqua purificata 18.50
Preparation: dissolve the lidocaine hydrochloride in the aqueous phase. Mix the emulsifier
into the oily phase, then emulsify the previously prepared aqueous phase in portions. Shake
vigorously.
_______________________________________________________________________________
3. The carrying out of dialysis
Carefully measure out 5 mL of the emulsions prepared into each dialysis membrane with a
digital pipette. Make sure that the dialysis membranes are closed bubble-free.
When you have finished the insertion of the sample, the investigation is started. Start the
first measurements after 15 minutes by taking out 5 mL of each sample. Filter it on a filter
paper, then determine spectrophotometrically the amount of lidocaine hydrochloride dissolved
in the acceptor phase and replace the amount of the sample taken out (5 mL) with distilled
water.
The spectrophotometric evaluation is made possible by the light absorption maximum of
the model API at 263 nm. At this wavelength, the values of the calibration curve were
recorded previously, and by using them the measured extinction values can be recalculated
into the concentration of the API (0.100 A = 154.262 µg/mL). During the measurements,
make dilutions if necessary and consider them when making the final calculations. Repeat the
sampling at the times given (15, 30 and 60 minutes).
_______________________________________________________________________________
Report
21. Investigation of the drug release of emulsions by static diffusion method
Name: ..................................................... Group: ....................... Date: ............................
1. Fill in the table below with the results of the investigation and calculate the appropriate
values. Give the total amount (in mg!) of the API dissolved after 60 minutes.
API in the
Time c API in the dissolution Total API
Sample A dilution sample medium dissolved
(min) (mg/mL) (mg/5 mL) (mg/95 (mg)
mL)
B
_______________________________________________________________________________
2. Show your results graphically by plotting the total amount of the API diffused against
diffusion time.
Evaluation:
_______________________________________________________________________________
22. Investigation of drug release by agar plate diffusion method
Introduction
The ointment bases and excipients used in formulation can modify drug release, and
thereby the efficacy of the drug significantly. Their proper choice is very important with
regard to effect.
During the experiment the rate of the release of the incorporated API depending on the
properties of the ointment bases and drug concentration is determined under in vitro
conditions.
Task
Investigate the drug release from ointments containing salicylic acid. The iron (III)
chloride dissolved previously in the agar medium will form a coloured complex with salicylic
acid after its diffusion from the ointment samples. The size and the intensity of this violet-
coloured ring around the samples are proportional to the amount of salicylic acid released
(Figure 22.1).
Figure 22.1: Salicylic acid diffusion in iron (III) chloride containing agar gel
_______________________________________________________________________________
1. Preparation for the diffusion investigation
For the investigation, the students get the ready agar gels in Petri-dishes and the different
ointment samples in marked tubes. In each Petri dish six cylindrical holes should be punched
with a special puncher, spaced at equal distances from each other and not too close to the wall
of the Petri dish. Carry out 2 parallel measurements with the same ointment with different
salicylic acid contents. Mark the holes on the bottom of the Petri dish to avoid mixing up the
samples. Use the short markings on the tubes.
2. Measurement of drug (salicylic acid) release from different ointments
Carry out the experiments with 4 different ointments containing 3 different salicylic acid
concentrations (1 w/w %, 5 w/w %, 10 w/w %). Fill each hole completely with the samples.
Use a small spatula to help the filling. Take care to avoid the ointment getting onto the surface
of the agar gel. The samples must be in contact with the agar gel on the side of the holes.
Start measuring the release time as one Petri dish is filled. The first reading time is after 30
minutes. Measure 2 perpendicular diameters of the coloured ring by means of a graph paper.
Be careful to read the Petri dishes in the same order as they were filled. Measure the size of
the ring by putting a graph paper under the Petri dish and reading the diameter of the coloured
ring through the transparent agar gel.
The reading times of the coloured rings are: 30, 60, 90 and 120 minutes.
_______________________________________________________________________________
Report
22. Investigation of drug release by agar plate diffusion method
Name: ..................................................... Group: ....................... Date: ............................
1. Fill in the table below and calculate the average of the two parallel measurements.
30 min. 60 min. 90 min. 120 min.

API average diameter average diameter average diameter average diameter
Ointment
% (mm) (mm) (mm) (mm)
1. 2. 3. average 1. 2. 3. average 1. 2. 3. average 1. 2. 3. average
Evaluation:
_______________________________________________________________________________
2. Plot the results.
Ointment: ...............................................................
_______________________________________________________________________________
Ointment: ...............................................................
_______________________________________________________________________________
Ointment: ...............................................................
_______________________________________________________________________________
Ointment: ...............................................................
_______________________________________________________________________________
23. Investigation of drug release from suppositories by dynamic

diffusion method
Introduction
Suppository masses do not have an effect on their own, but they can modify drug release –
and thereby therapeutic effect – significantly. In this practice the rate of the release of the API
incorporated in different suppository masses and in different concentrations is determined by
dynamic diffusion.
Task
The drug release of readily water-soluble chloroquine phosphate and its diffusion through a
membrane from different suppository masses are determined. During the investigation the
drug release from the suppositories through a membrane is measured by determining the
chloroquine phosphate content in the acceptor phase spectrophotometrically.
Investigation method:
1. Measure the 3 suppositories received from your practice leader with mg accuracy and
mark them as Sample A, B, and C. You have 3 Petri dishes for the storage and the
identification of the suppositories. The base and the API content are the same in all 3
samples.
2. Cut off a piece from the membrane tube equalling the suppository length + 2 cm. Place
the suppositories into the tubes and close them with the closing device.
3. Place the samples prepared in this way into the apparatus.
4. Start the apparatus. The rotational speed is 1 turn / 30 s.
5. Take samples after 15, 30 and 60 minutes in a volume of 20 mL from the acceptor
phase with the rubber tube connected to the equipment. You will need 3 x 3 pieces of
Erlenmeyer flasks for the sampling.
6. Use an aliquot of the 20-mL samples and make a dilution to measure the value of
quenching. The spectrophotometric evaluation is made possible by the light absorption
maximum of chloroquine phosphate at 220 nm. At this wavelength, the values of the
_______________________________________________________________________________
calibration curve were recorded previously, and by using them the measured quenching
values can be recalculated into the concentration of the API. Consider the dilutions
when making the calculations.
7. The volume of the acceptor phase taken during sampling was supplied; do not forget
about it when you make the final concentration calculation.
Apparatus: Apparatus for the disintegration of suppositories
Use of apparatus
1. Switch on the thermostat, set the knob on the right side to VAR position, and the one on
left side to the desired temperature.
2. Measure 3.00 kg of distilled water into 3 glass beakers (3,000 mL) and place them into
the apparatus. Wait until the desired temperature is reached in the acceptor phase.
3. Place the wrapped suppositories into the metal baskets, immerse them into the acceptor
phase and fix the rotating device.
4. Switch on the timer.
5. Switch the knob to ALAP position.
6. Press the START knob (to set the rotating device to the right position).
7. Press the ‘STOP’ knob.
8. Switch the knob to ‘STOP’ position.
9. Set the timer: 00, 30. (In ALAP position the two numbers mean hours and minutes, in
TIMER position minutes and seconds.)
10. Press the LOAD knob.
11. Switch the knob to ‘TIMER’ position.
12. As the switch is turned to ‘TIMER’ position, the investigation is started. Half a
rotation is made, repeated every 30 seconds.
The apparatus should work continuously during the 1 hour of investigation time. Take the
samples after 15, 30 and 60 minutes. When the experiment is finished, switch off the
apparatus and the thermostat.
_______________________________________________________________________________
Start the work by switching on the thermostat and measuring the acceptor phase. You have
time to prepare the suppositories for investigation till the acceptor phase reaches the desired
temperature.
_______________________________________________________________________________
Report
23. Investigation of drug release from suppositories by dynamic diffusion method
Name: ..................................................... Group: ....................... Date: ............................
1. Give the following data.

Number of composition: .............................................
API concentration: ...................................................... %
Weight of suppository A: ............................................ mg
Weight of suppository B: ............................................ mg
Weight of suppository C: ............................................ mg
2. Fill in the table with the results of the three parallel measurements (aliquot, dilution,
quenching values).
Sampling time (min)

Supp. 15 30 60
Aliquot Dilution Quenching Aliquot Dilution Quenching Aliquot Dilution Quenching
A
B
C
3. Calculate the released amount of API in lg/mL. Use the values of quenching, and the
slope and the intersection of the calibration curve. Give the released amount of API in
mg and w/w%, too. During the calculation, take care to consider the dilution, the
starting amount of the acceptor phase and the supplied amount of the sampling volume.
Equation of the calibration curve:

y = a x+b (1)
where y is the quenching, a is the slope (0.0618), x is the concentration (lg / mL), b is the
intersection (- 0.0776).
The absorption maximum of chloroquine phosphate is at 220 nm.
_______________________________________________________________________________
Sampling time Released API (mg)

(min) Supp. A Supp. B Supp. C Average
15
30
60
Sampling time Released API (%)
(min) Supp. A Supp. B Supp. C Average
15
30
60
3. Plot the released API % against sampling time.
Evaluation:
_______________________________________________________________________________
24. Investigation of the mucoadhesivity of hydrogels

(Calculation task)
Introduction
The mucoadhesivity of a pharmaceutical preparation means the adhesion between the

preparation and the mucosa, in the course of which physical and chemical interactions can
form. Physical interactions mean the diffusion of the polymer chains into the mucin chains
and their interpenetration. Chemical interactions mean the formation of primary (e.g.
covalent) and secondary (e.g. hydrogen, van der Waals) bonds between the entangled chains
(Figure 24.1).
Figure 24.1: Mechanism of mucoadhesion

A: contact stage; B: consolidation stage;
C: formation of primary and secondary chemical bonds between the mucin and polymer chains
As mucoadhesion is a complex mechanism, numerous theories exist to explain it, such as:
– electric;
– adsorption;
– wetting;
– diffusion and
– fracture theory.
_______________________________________________________________________________
Because of these complexities, there is no standardized method of measurement; numerous

methods are known and developed to investigate mucoadhesive behaviour. One of the most
accepted methods is rheology. The rheological synergism between the polymer and the mucin
chains can be regarded as an in vitro parameter of mucoadhesion. The base of the
methodology is that the rheological parameters change after mixing the polymer gels with
mucin. This change is greater than it could be expected from the sum of the rheological
parameters.
Hassan and Gallo were the first to describe the relationship which uses a viscosity
parameter (2b) to characterize the strength of mucoadhesivity:
b = t m p (1)
where, 2t is the viscosity of gels containing polymers and mucin, 2p is the viscosity of the
polymer gels without mucin, and 2m is the viscosity of mucin dispersion.
Besides the equation above, the relative synergism parameter can be also used, where the
changes in viscosity refer to the original viscosity:
b ,rel = b
(2)
m + p
Task
The mucoadhesivity of polymer gels containing different amounts of hydroxyethyl

cellulose polymer (HEC) is investigated by means of rotational viscometry. From the
measured data listed below, the synergism and relative synergism parameters can be
calculated, and the changes of mucoadhesivity can be characterized by increasing the polymer
concentration.
The flow and viscosity curves of the mucin (porcine stomach type II) dispersion, and gels
containing HEC alone and HEC and mucin together were recorded in the range of 0.1-100 1/s
shear rate.
_______________________________________________________________________________
The composition of the samples is listed in the table below:
Sample Mucin HEC

(%) (%)
1 4 0
2.1 0 1
2.2 0 2
2.3 0 3
2.4 0 5
3.1 4 1
3.2 4 2
3.3 4 3
3.4 4 5
_______________________________________________________________________________
Report
24. Investigation of the mucoadhesivity of hydrogels
(Calculation task)
Name: ..................................................... Group: ....................... Date: ............................
1. Shear stress (D) and viscosity data (2) (at 100 1/s) derived from the flow and viscosity
curves can be seen in the table below. Three parallel measurements were made.
Calculate the mean values.
Measurement 1 Measurement 2 Measurement 3 Mean

Sample I C I C I C I C
(Pa) (mPa·s) (Pa) (mPa·s) (Pa) (mPa·s) (Pa) (mPa·s)
1 1.5 15.1 1.6 15.5 1.3 13.2
2.1 2.1 21.1 2.1 21.3 2.1 20.8
2.2 14.3 143 14.4 144 14.3 143
2.3 43.5 435 45.5 455 50.7 507
2.4 163 1630 182 1820 148 1480

3.1 7.32 73.2 7.6 75.7 8.9 88.8
3.2 35.6 356 35.1 351 33 330
3.3 89.6 896 88.1 881 90.8 908
3.4 244 2440 306 3060 296 2970
_______________________________________________________________________________
2. Plot the values of the shear stress against the HEC concentration both for the sample
containing HEC alone and for the sample containing HEC and mucin together.
3. Evaluation:
_______________________________________________________________________________
4. Calculate the value of 2b and 2b rel.
HEC conc. (%) Cb Cb rel

(mPa*s) (-)
1
2
3
5
5. Plot 2b and 2b rel against the HEC concentration. Use a different y-axis for the 2b (left
side y-axis) and the 2b rel (right side y-axis) values.
Evaluation:
_______________________________________________________________________________
Appendix
_______________________________________________________________________________
A.1. Helios Alpha UV-Vis spectrophotometer
The spectrophotometer is an optical instrument, which measures the intensity of

monochromatic light and its changes.
Spectrophotometers can be classified on the basis of:

– applied wavelength:
ultraviolet
visible
infrared
– operation way:
single beam
double beam
The Helios Alpha UV-Vis spectrophotometer (Figure A.1.1) is a double beam

spectrophotometer, which means the optical system divides the light from the light source into
two paths, one of them goes through the reference, and the other one goes through the sample.
This operation way allows the simultaneous measurements of light intensity passing through
the sample and the reference.
Figure A.1.1: Unicam Helios Alpha UV-Vis spectrophotometer

_______________________________________________________________________________
Parts list:
– light source (wolfram, deuterium lamp, R = 190-1100 nm, stepwise: 2 nm)
– sample cell with holder (quartz cuvettes)
– monochromator
– detector
– display
Operations:
– scanning mode (SCAN)
absorbance (and its 1st-4th derivative)
transmittance
intensity
– at fixed wavelength
absorbance
transmittance
concentration (knowing the correlation between the concentration and the absorbance).
– plotting of the calibration curve
_______________________________________________________________________________
A.2. Anton Paar RHEOLAB MC 1 rheometer
The Anton Paar Physica RHEOLAB MC 1 rheometer is a rotational rheometer, which can be
used to measure the flow curve, yield point, structure breakdown and recovery (CREEP test),
viscosity of Newtonian and non-Newtonian systems. The instrument can be used in CSR
(controlled shear rate) and CSS (controlled shear stress) mode, too (Figure A.2.1).
Figure A.2.1: Anton Paar Physica RHEOLAB MC 1 rheometer
The measuring device is a concentric cylinder. The sample is placed in a gap between a fixed
cup and a rotating cylinder (Searle system). The measuring controller developed for the
device is based on a dynamic system which consists of a drive unit and an optical encoder
without gear wheel and mechanical power converter, therefore it measures the rotary torque
without deformation, so without loss. The measuring controller is such that the revolving
body rotates at the desired speed and it measures the rotary torque – which comes from the
flow resistance of the sample – acting on the measuring body. The investigation of shear
stress or the CREEP test is also possible by setting the desired shear stress, and the shear
deformation of the sample is measured through the angle rotation.
Controlled shear investigations (CSS) for the determination of the flow properties of
plastic materials can also be performed by the Anton Paar Physica RHEOLAB MC 1 device,
_______________________________________________________________________________
which allows the accurate measurement of the flow point without deformation, so without the
shearing of the internal structure of the sample (rotation).
Handling of the device, measurement:

1. In case of the investigation of emulsions, sample holder "Z3" is filled hermetically with
the sample up to the inner mark (Figure A.2.2).
1. In case of the investigation of the gel-forming of polymers and the investigation of
ointments, sample holder "Z4" is filled hermetically with the sample up to the inner
mark (Figure A.2.3).
2. The cylinder measuring head is fitted and fixed with the help of the retaining ring.
3. The device is switched on at the back.
4. The "PROG" function is chosen and the "OK" button is pressed.
5. In case of the investigation of emulsions, the "PROG 02" function is chosen and the
"OK" button is pressed.
5. In case of the investigation of the gel-forming of polymers, the "PROG 01" function is
chosen and the "OK" button is pressed.
6. The measurement is started by pressing the "ST" (START) button.
7. A piping sound will indicate the end of the measurement.
Figures A.2.2 and A.2.3: Measuring heads "Z3" and "Z4"

Analysis of Pharmaceutical Ingredients, Excipients and Dosage Forms (Practical Workbook)

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University of Szeged

ANALYSIS OF PHARMACEUTICAL INGREDIENTS,

1. Determination of the dissolution rate of the active substance ....................................7

II. Investigation of dosage forms ...................................................................................28

5. Determination of the drop-weight of dosage forms used dropwise ..........................29

III. Investigation of stability ...........................................................................................88

15. Determination of the decomposition (caramelization) of glucose solution ..............89

IV. Biopharmaceutical investigations ...........................................................................117

21. Investigation of the drug release of emulsions by static diffusion method.............118

Helios Alpha UV-Vis spectrophotometer ...............................................................143

Szeged, 1st June, 2015

Dr. Mária Budai-Sz"cs Dr. Piroska Szabó-Révész Dr. Zoltán Aigner

1. Determination of the dissolution rate of the active substance

Measure theobromine samples for three parallel examinations. 0.2500 g of theobromine

Figure 1.1: Basket and paddle dissolution apparatus

2. Calculate the dissolution rate constant of theobromine (k).

3. Plot the average of c* against sampling time.

4. Write a short evaluation of the results of the test.

Results of the dissolution test of theobromine

3. Plot the average of c* against sampling time.

4. Write a short evaluation of the results of the test.

2. Determination of the spectral transmission of glass containers

Apparatus: UV-VIS spectrophotometer, in transmittance mode

Limits (Ph. Hg. VIII.; 3.2.1.)

Maximum percentage of spectral

Glass sample (code, characteristic):

Measurement Thickness (mm)

Transmittances referring to a thickness of 1 mm:

3. Investigation of the gel formation of Carbomera

3. Prepare again Carbomera containing systems according to point 2. Adjust the pH up to

3. Prepare again Carbomera containing systems according to point 2. Adjust the pH up to

4. Investigation of the consistency of suppository bases –

The investigation of the consistency and hardness of suppositories is important with

1. Determination of resistance to rupture

3. Determination of consistency by penetrometry

the depth of penetration (cm):

Figure 4.2: Penetrometer

1. Determination of resistance to rupture

Letter symbol of series: ..........................................................

Mark Measurement Measurement Measurement Measurement Measurement Average

2. Determination of the flow point of suppository

II. Investigation of dosage forms

5. Determination of the drop weight of dosage forms used

Figure 5.1: Standard dropping tube

Name of the excipient used: .....................................................

Drop Drop Drop Drop Drop Drop

Name of the excipient used: .....................................................

Drop Drop Drop Drop Drop

Name of the excipient used: .....................................................

Name of the excipient used: .....................................................

Alcoholum 96% Amount of Mucilago methylcellulosi Amount of

Polysorbate 20 Amount of Tinctura aromatica Amount of

6. Determination of the viscosity of dextran infusion and the

Figure 6.1: Capillary viscometer

20°C =k 20°C t (2)

1. The determination of the yield properties and viscosity of emulsions

Apparatus: Anton Paar Rheolab MC 1 Rheometer

2. Investigation of spontaneous separation (not an official investigation method in Ph. Hg.

Materials I. II. III. IV. V.

Apparatus: Mixer (stage 1, 1 min of mixing time)

2. The effect of the concentration of emulsifiers on the properties of the emulsion:

3. Rheological characterization of the compositions tested (determination of flow type):

I. II. III. IV. V.