Pharmaceutical Analytical

Chemistry (1216213)

Prof. Saleh Abu-Lafi

Faculty of Pharmacy
Al-Quds University

September 2023

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Textbook

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 Profiles of physico-chemical properties of
some drug molecules

Drug type: analgesic.

Functional groups:
• A – Carboxylic acid, weak acid, pKa 3.5.
• B – Phenolic ester, particularly unstable.
Half-life in water: 52 h at pH 7.0 and 25oC; 40 days at pH 2.5 and 25oC.

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Additional information: The rate of HO- catalyzed hydrolysis of esters is > the
rate of H+ hydrolysis of esters. Solid aspirin absorbs water from the atmosphere
and then hydrolyses (Fig. 2.13).

Aspirin is mostly absorbed in the stomach! Why?

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Drug type: analgesic.
Functional groups:
• A – Amide group, neutral.
• B – Phenolic hydroxy group, very weak acid, pKa 9.5.
Half-life in water: 21.8 years at pH 6 and 25oC. Most amides are very stable to
hydrolysis (Fig. 2.11).

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Drug type: antibiotic.
Functional groups:
• A – Amide, neutral, relatively stable, see paracetamol.
• B – Thioether, neutral, can be oxidized at high levels of oxygen stress.
• C – Lactam ring, neutral, particularly susceptible to hydrolysis (Fig. 2.15).

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 What Is Analytical Science?
Analytical Chemistry provides the methods and
tools needed for answering 4 basic questions
about a sample:
•What? qualitative analysis
•Where? proper sampling, spatial analysis
•How much? quantitative analysis
•What arrangement, structure or form? determine
the species present chemical speciation
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 Analytical Chemistry
Science of Measurements

Questions:
1. Is this water safe to drink?
2. Is there (THC) in his urine sample?
3. What is the composition of this rock?
4. What is the ‘assay’ in this analgesic drug?
5. Does this blood or urine contains a banned
substance (stimulants, steroids, etc.)?

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 As analyst, if somebody put a bottle on your disk and
asked: what is in there? or is this safe? there are more than
20 million compounds!
 The concept of safe or (zero/nothing) is hard to define.
Don’t say the water is safe to drink, but it falls within the
specs!
 Never report an answer of zero but less than < detection
limits (DL) of the instrument.
 Medical cannabis contains tetrahydrocannabinol (THC)
which is psychoactive compound while cannabidiol
(CBD) is a nonpsychoactive compound. CBD oil is used
to help various conditions, such as: epilepsy, pain,
inflammation, depression, anxiety.
 Rocks are naturally occurring solid mass of minerals
Si, Al, Fe, Ca, Na, K, Mg, O, etc. 11
 Assay of Active Pharmaceutical Ingredients (API) in
Pharmaceutical dosage forms (finished products)
 The list of prohibited substances (banned substances at
the Olympic Games) includes about 500 different active
constituents: stimulants, steroids, beta-blockers, diuretics,
narcotics, analgesics, local anesthetics, and sedatives.
Some are detectable only as their metabolites. Many
athletes must be tested rapidly.
 There are three phases in the analysis:
1. Fast-screening phase: immunoassays, GC, LC analysis
2. Identification phase: identified by GCMS, LCMS
3. Possible quantification: since they are present at low
levels (traces).
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What do Chemical Analyst Do?
Analyst:
•Applies known measurement techniques (usually
using SOP) to well defined analytical questions.
Senior Analyst:
•Develops new measurement methods on existing
principles to solve new analysis problems.
Research Analytical Chemist:
•Creates and /or investigates novel techniques for
chemical measurements.

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 Areas of Chemical Analysis
Answers 4 basic questions about a material sample?

1. Separation:
– How can the species of interest be separated from the
sample matrix for better quantitation and identification?
2. Detection:
– Does the sample contain substance X?
3. Identification:
– What is the identity of the substance X in the sample?
4. Quantitation:
– How much of substance X is in the sample?

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An
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operationswhich
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depend on:
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The
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involved in every step

Steps in analysis
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The analytical process involves a sequence of logical
events including:

1. Defining the problem:

This means that the analyst (problem solver) should
know what information is required, the type and
amount of sample, the sensitivity and accuracy of the
results, the analytical method which can be used to
achieve these results, etc...

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2. Obtaining a representative sample:
It is very important to collect a representative
sample for analysis (sampling). One should
take several samples from different locations
and depths, mix them well and then take a
sample for analysis.
(Bulk of material)
 Gross sample (few g to Kg)
 Lab sample (few g)
 Analysis sample what is actually analyzed
(few mg or few ml)
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3. Preparing the sample for analysis:
 Most analytical methods require a solution of the sample
rather than solid. Therefore, samples should be dissolved
quantitatively and may be diluted to the concentration
range of the method.
 Blood can be analyzed as serum: the liquid from clotted
blood or as plasma: liquid from un-clotted blood (by
adding heparin or citrate salt) followed by centrifuge to
get the red cells separated from the plasma
 Enzymes and proteins must be analyzed fast to prevent the
denature
 Urine samples are unstable, keep it acidic at pH 4.5 by
adding 1-2 ml of glacial acetic acid per 100 ml sample to
prevent precipitation of calcium phosphate which entrap
metal ions or other substances

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4. Chemical separations:
The sample may contain solutes which interfere with the
determination of the analyte. If this is the case, analytes
should be separated from the sample matrix by an accepted
procedure. Separation steps may include precipitation,
distillation and chromatography
 Dry ashing: the sample is slowly combusted in a furnace at ~
400-700Co, leaving behind inorganic residues that is soluble
in diluted acids.
 Wet digestion: destroying the organic matter by heating with
oxidizing acids (mixture of HNO3+H2SO4)
 Biological fluids precipitation of proteins by methanol or
acetonitrile solvents
 Always run a blank! A blank consists of all chemicals except
the analyte
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5. Performing the measurement:
This implies conducting the analytical procedure and
collecting the required data.
-Calibration
-Validation
-Replicates
6. Calculations:
The final event in the analytical process is to perform the
calculations and present the results in an acceptable
manner.
• Statistical analysis

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Range: what is the size of the sample?
The size of the sample can be used to describe the class of a
method where
Meso if the sample size is >100 mg or 100 microliters (l).
A semimicro if the sample size is from 10-100 mg or 50-100
microliters (l).

When the sample size is in the 1-10 mg or <50 l, the

method is said to be a micro method
while a sample size <1 mg denotes an ultramicro method.

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 The
Thesample
samplesize
sizedictates
dictateswhat
whatmeasurement
measurementtechniques
techniquescan
canbe
beused.
used.

©Gary Christian, Analytical Chemistry, 6th Ed. (Wiley)

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An analyte in a sample can be classified as:
1. a major constituent if it constitutes more than >1% of
the sample or
2. a minor in the range from 0.1-1.0 %.
3. It is classified as trace if it constitutes less than < 0.1%.
Analyze versus Determine
These terms are sometimes misused.
Always use the term ‘analyze’ with the sample,
while use the term ‘determine’ with specific analyte.
Therefore, a sample is ‘analyzed’, while an analyte is
’determined’.

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 History of Analytical Methods

1. Classical methods: Early years (separation of analytes) via

precipitation, extraction or distillation

Qualitative: recognized by color, boiling point, solubility, taste

Quantitative: gravimetric or titrimetric measurements
also called ‘wet chemistry’ or ‘bench chemistry’ very precise ~ 0.1%
but need large solute quantity

2. Instrumental Methods: newer, faster, more efficient and more

sensitive, selective and automated but less precise ~1-5 %
Physical properties of analytes: conductivity, electrode potential,
light emission, absorption, mass to charge ratio and fluorescence,
many more…
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 Summary of analytical methods

Some of ‘classical methods’ are still serve as standard

pharmaceutical methods of analysis in USP and BP.
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 Different
Differentmethods
methodsprovide
provideaarange
rangeof
ofprecision,
precision,sensitivity,
sensitivity,selectivity,
selectivity,
and speed capabilities.
and speed capabilities.

©Gary Christian, Analytical Chemistry, 6th Ed. (Wiley)

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 Chemical Analysis Affects
Many Fields
• Pharmacy: analysis of active ingredients, pharmaceuticals
USP, BP, JP, EP, etc.
• Medicine: clinical lab tests to diagnose disease, etc.
• Industry: test raw material, finished products, paints, etc.
• Environmental: monitor levels of pollutants in the
atmosphere, rivers, etc. “EPA”
• Food and agriculture: Nutrient analysis, protein, fat,
carbohydrates, mineral and vitamins, additives such as sweeteners,
preservatives and colors, toxic metal contaminants, etc.
• Forensics: e.g. analysis of prohibited substances, firearms
identification, DNA analysis, etc. 27
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 Calibration and measurments
Y = aX + b
Limit of linearity
 Limit of detection (LOD) and LOQ: Slope relates to
the lowest amount (concentration or

Detector response
sensitivity
mass) of an analyte that can be
detected (or quantified) at a known LOQ
confidence level respectively LOD
 Linearity: the degree to which a Dynamic range

response of an analytical detector to

analyte concentration/mass Concentration
approximates a linear function
 Linear dynamic range: the range over which measurements can be made
(usually the linear range), often defined by detector dynamic range
 Selectivity: the degree to which a detector is free from interferences
(including the matrix or other analytes)
 Sensitivity: the slope of the curve, the larger the slope, the more sensitivity
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Example: Caffeine content in a chocolate
bar?

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• The first step is to obtain a representative sample
to measure, a process called sampling.
• A pure chocolate bar is homogeneous, which
means its composition is the same everywhere.
• Chocolate with nuts in it is an example of a
heterogeneous, composition differs from place to
place.

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 Measuring the Amount of
Analyte (As) in Deer Kidney
• Spectrophotometer: Highly colored
complex of arsenic (As) was found to
absorb light at a wavelength of 535 nm.

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Conc. Absorbance
ppm
0 0 Absorbance vs Concentration
5 0.16
10 0.28
15 0.41
20 0.595 0.8 y = 0.0282x + 0.005
25 0.7
0.6 R2 = 0.9961
deer 1 0.61
deer 2 0.43
Absorbance 0.4
0.2
0
0 5 10 15 20 25 30
Conc., ppm

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 Calculating the Concentration
• ppm = (Absorbance - 0.005)/0.0282
• Deer 1: (0.61 - 0.005)/0.0282 = 22 ppm
• Deer 2: (0.43 -0.005)/0.0282 = 15 ppm
• Arsenic in the kidney tissue of animals is
toxic at levels above about 10 ppm.

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 Homework 1
• Construct a calibration curve using
spreadsheet excel to measure the amount of
analyte (As) in Deer Kidney
• Hint: You can learn how to do it by
refereeing to page 118 of the textbook.

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