The document discusses types and sources of impurities in active pharmaceutical ingredients, including genotoxic impurities. It describes how impurities can arise from starting materials, byproducts, reagents, and degradation of the drug substance over time. Impurities are classified as organic, inorganic, residual solvents, or genotoxic. A five-class system is presented for categorizing genotoxic impurity risk based on structural properties and existing toxicity data.

1. Presenting by:
Mr. PURSHOTHAM K N
Asst. Professor,
Dept. of Pharmaceutical Chemistry
SAC College of Pharmacy.
2022-2023
IMPURITIES IN API’s INCLUDING GENOTOXIC IMPURITIES –
TYPES AND SOURCES

2. CONTENTS:
• Introduction
• Sources of impurities
• Types of impurities
• Sources of organic impurities
• Genotoxic impurities
• References

3. IMPURITIES:
• Impurities are chemical substances composed of liquid, gas, or solid,
which differ from the chemical composition of the material or
compound.
• They are either naturally occurring or added during synthesis of a
chemical/commercial product.
• During production, impurities may be purposely, accidentally, added
into the substance.
• Inorganic contaminants are not considered as an impurity unless they
are toxic, such as heavy metal or arsenic.

4. Sources and Route of impurities
A) SOURCE:
Most API are produced by organic chemical synthesis.
Various components used in the synthesis including starting materials,
by-products, residual solvents, trace amounts of inorganic, and organic
components can be generated during such a process.
 Those components remaining in the final API are considered as
impurities

5. B) ROUTES
 Degradants formed during the process and long-term storage.
 Contaminants from packaging components and other drug products
manufactured in the same facility.
 Impurities could be forming from the impact of heat, light, and
 Oxidants (including air) on the drug, Changes in the ph of the
formulation.
 Interactions with packaging components, excipients and other active
ingredients, in the case of combination products.

6. Common terminologies for Impurities:
a) Intermediate, Penultimate Intermediate and By-products:
• The compounds produced during synthesis of the desired material are
called intermediates, especially if they have been isolated and
characterized.
• The penultimate intermediates are the last compound in the synthesis chain
prior to the production of the final desired compound.
• Byproducts are unplanned compounds produced in between the reaction.
b) Transformation Products:
• Similar to byproducts which relates to theorized and non-theorized
products that may be produced in the reaction.

7. c) Interaction products:
• Interaction products that could occur between various involved chemicals
intentionally or unintentionally.
d) Related Products: These products have similar chemical structure and
potentially similar biological activity.
e) Degradation products: These compounds are products due to
decomposition of the active ingredient or the material of interest

8. 1. Organic impurities (process- and drug-related).
2. Inorganic impurities.
3. Residual solvents.
4. Genotoxic impurities.
CLASSIFICATION:

9. 1. Organic impurities:
• Most of the drug substances (low molecular weight) are chemically
synthesized.
• Chemical entities, other than the drug substance, that are involved or
produced in the synthetic process can be carried to the final drug
substance as trace level impurities.
• These chemical entities include raw materials, intermediates, solvents,
chemical reagents, catalysts, by-products, impurities present in the
starting materials- referred to as process impurities.
• The goal of process impurities identification is to determine the
structures and origins of these impurities.
• This knowledge is critical for improving the synthetic chemical
process, in order to eliminate or minimize process impurities

10. • Starting materials and intermediates: chemical building blocks used to
construct the final form of a drug substance.
• Unreacted starting materials and intermediates can potentially serve as
impurities.
• For example, synthesis of tipranavir, aniline is the intermediate in the last
step of the synthesis.
• Due to the similarity between the structures of aniline and the final product,
it is difficult to totally eliminate it in the subsequent purification step.
Consequently, it appears in the drug substance at around 0.1%.
• 2-methyl analogue present as a trace impurity in tolperisone, due to the
presence of 2methylpropiophenone in the starting material,
4methylpropiophenone.

11. • Reagents, ligands and catalysts: Chemical reagents, ligands, and catalysts
used in the synthesis of a drug substance can be carried over to the final
products as trace level impurities.
• For example,
carbonic acid chloromethyl tetrahydro-pyran-4-yl ester (CCMTHP),
which is used as an alkylating agent in the synthesis of a ß lactam drug
substance, was observed in the final product as an impurity.
Beta-Lactum API

12. • Byproducts of the synthesis: All chemical reactions are not 100%
selective; the side-reactions are common during the synthesis of drug
substances.
• By-products can be formed through a variety of side reactions, such as
incomplete reaction, over reaction, isomerisation, dimerisation,
rearrangement, or unwanted reactions of starting materials or intermediates
with chemical reagents or catalysts.
• Products of over-reaction: In many cases the previous steps of the
syntheses are not selective enough and the reagents attack the intermediate
not only at the desired site.
• For e.g. in the synthesis of nanodralone decanoate, the last step of the
synthesis is the decanoylation of the 17-OH group.
• In the course of overreaction the reagents also attack the 4-ene- 3 oxo group
leading to an enol ester- type impurity

13. • Impurities from degradation of the drug substance: Impurities
can also be formed by degradation of the end product during
manufacturing of bulk drugs.
• Degradation products resulting from storage or formulation to different
dosage forms or aging are common impurities in the medicines.
• ICH guidelines, is a molecule resulting from a chemical change in the
substance brought about by overtime or due to the action of light,
temperature, pH or water or by reaction with excipient and/or the
intermediate container closure system.
• For example in the case of aspartame, in the presence of moisture,
hydrolysis occurs to form the degradation products L- aspartyl- L
Phenyalanine and 3-benzyl-6-carboxymethyl 2, 5-diketopierazine. A
third degradation product β-L- aspartyl-L-phenylalanine methyl ester is
also known to form. Aspartame degradation also occurs during
prolonged heat treatment.

14. • A third degradation product β-L- aspartyl-L-phenylalanine methyl ester is
also known to form.
• Aspartame degradation also occurs during prolonged heat treatment.

15. 2. Inorganic impurities:
• The main sources of heavy metals are the water used in the processes
and the reactors (if stainless steel reactors are used), where acidification
or acid hydrolysis takes place.
• Leaching from the process equipment or storage containers.
• These impurities of heavy metals can easily be avoided using
demineralized water and glass-lined reactors.
• Many chemical reactions are promoted by metal based catalysts. For
instance,
1. a Ziegler-Natta catalyst contains titanium,
2. Grubb’s catalyst contains ruthenium,
3. and Adam’s catalyst contains platinum.

16. 3. Residual solvents
• Residual solvents are potentially undesirable substances.
• They either modify the properties of certain compounds or may be
hazardous to human health.
• Residual solvents are potentially undesirable substances. They either
modify the properties of certain compounds or may be hazardous to
human health.
• The residual solvents also affect physicochemical properties of the bulk
drug substances such as crystallinity of bulk drug, which in turn may
affect the dissolution properties, odor and color changes in finished
products.

17. a. Class I solvents:
Class I solvents and their permissible concentration limits given in
the Table 1. These solvents not employed in the manufacture of drug
substances, excipients and formulations because of their unacceptable toxicity
or their deleterious effects. If use of these solvents is unavoidable, then their
usage should be restricted.
Residual solvent Concentration limit (ppm)
Benzene 2(Carcinogenic)
Carbon tetrachloride 4(Toxic)
1,1 Dichloro ethene 8(Toxic)
1,2 Dichloro ethene1,1 5(Toxic)
1,1,1 Trichloro ethane 1500(Environmental hazard)
Table 1: Class I Residual Solvents.

18. b) Class II solvents:
Class II solvents usage should be limited in pharmaceutical
products because of their inherent toxicity.
c) Class III Solvents:
These are less toxic and possess lower risk to human health than
class I or class II solvents Long-term toxicity or carcinogenicity not
reported, which is evident from the available data for the solvents under this
category. The use of class III solvents in pharmaceuticals does not have any
serious health hazard.

19. Genotoxicity:
• The property of chemical agents that damages the genetic information
within a cell causing mutations, which may lead to cancer.
• While genotoxicity is often confused with mutagenicity, all mutagens are
genotoxic, whereas not all genotoxic substances are mutagenic.
• The alteration can have direct or indirect effects on the DNA: the
induction of mutations, and direct DNA damage leading to mutations.
• The permanent, heritable changes can affect either somatic cells of the
organism or germ cells to be passed on to future generations.

20. • Cells prevent expression of the genotoxic mutation by either DNA repair
or apoptosis; however, the damage may not always be fixed leading to
mutagenesis.
• Formation of defective Protein, Create Disorder in Metabolic processes,
Affect the DNA repair Mechanism.
• Examples: Alkylating agents, nitroso groups, lead, arsenic etc.

21. Types of DNA damage:
1. Single-base alteration
a. Depurination
b. Deamination of cytosine to uracil
c. Deamination of adenine to hypoxanthine
d. Alkylation of base
e. Insertion or deletion of nucleotide
f. Base-analog incorporation.
2. Two-base alteration
a. UV light–induced thymine-thymine (pyrimidine) dimer

22. 3. Chain breaks
a. Ionizing radiation
b. Radioactive disintegration of backbone element
c. Oxidative free radical formation.
4. Cross-linkage
a. Between bases in same or opposite strands
b. Between DNA and protein molecules (eg, histones).

23. A FIVE-CLASS SYSTEM FOR CATEGORIZING GENOTOXIC
IMPURITIES:
CLASS 1:
• Impurities known to be genotoxic (mutagenic) and carcinogenic.
• It includes known animal carcinogens with reliable data for a genotoxic
mechanism, and human carcinogens.
• The genotoxic nature of the impurity is demonstrated using publisheddata
on the chemical structure.

24. CLASS 2:
• Impurities known to be genotoxic (mutagenic), but with unknown
carcinogenic potential. It includes impurities with demonstrated
mutagenicity based on testing of the impurity in conventional
genotoxicity tests.
CLASS 3:
• Impurities that have an alerting structure unrelated to the structure of the
API, and of unknown genotoxic (mutagenic) potential.
• It includes impurities with functional moieties that can be linked to
genotoxicity based on structure.
• These moieties have not been tested as isolated compounds and are
identified based on chemistry and using SAR

25. CLASS 4:
• Impurities with an alerting structure related to the API and impurities that
contain an alerting functional moiety that is shared with the structure of
the API.
CLASS 5:
• No alerting structure or indication of genotoxic potential.