Adulteration is the debasement of Genuine materials3. It may be deliberated or accidentally done. In crude drugs, this practice includes the substitution of the original crude drug, fully or partially with sub-stranded substances. Sub-stranded substances include- Inferiority and spoilage. Inferiority- Natural sub-stranded condition whose natural constituent is below the minimum standard. Spoilage- sub-stranded condition produced by microbial or pest infestation.

1. ADULTERATION AND
DETERIORATION
By- Durgashree M D
Advance Pharmacognosy 2
M Pharm, Dept of Pharmacognosy
KLE College of Pharmacy, Bengaluru

2. CONTENT
Introduction
Types of Adulteration/substitution of Herbal Drugs
Causes and Measures of Adulteration
Sampling Procedure
Determination of Foreign Matter
DNA Fingerprinting Techniques in Identification of Drugs of Natural Origin
Detection of Heavy Metals, Pesticidal Residue, Phytotoxin.
Microbial Contamination in herbs and their formulation.

3. INTRODUCTION
• Adulteration is the debasement of Genuine materials3.
• It may be deliberated or accidentally done.
• In crude drugs, this practice includes the substitution of the original
crude drug, fully or partially with sub-stranded substances.

4. INTRODUCTION
• Sub-stranded substances include-
Inferiority and spoilage.
• Inferiority- Natural sub-stranded condition
whose natural constituent is below the
minimum standard.
• Spoilage- sub-stranded condition
produced by microbial or pest infestation.

6. TYPES OF
ADULTERATION /
SUBSTITUTION
Substitution with Inferior Varieties
Adulteration by artificially manufactured
substitutes
Substitution by Exhausted drugs
Substitution by Superficially similar but cheaper
natural Substance.
Adulteration by addition of worthless heavy
materials.
Addition of synthetic principle.
Usage of vegetative material from the same
plant.
Adulteration of Powders

7. 1. Substitution with Inferior Varieties
 Substitution of Alexandrian Senna with Indian
Senna
 Substitution of Capsicum annum fruits with
Capsicum minimum
2. Adulteration by artificially manufactured substitutes:
Pieces of Basswood into correct size & shape of
Nutmeg and sprayed with a volatile oil

8. 3. Substitution by Exhausted drugs
Exhausted Cloves substituted with genuine Cloves
Exhausted Ginger to genuine Ginger
Exhausted Benzoin to genuine Benzoin
4. Substitution by Superficially similar but cheaper natural
Substance.
Addition of Clove stalks to genuine Cloves
Substitution of Digitalis purpurea with leaves of
Digitalis thapsi

9. 5. Adulteration by addition of worthless heavy materials.
Mass of stones mixed with Licorice root
Pieces of limestone in asafoetida
lead shots in opium
6. Addition of synthetic principle.
Addition of synthetic Citral to oil of Lemon
Addition of synthetic balsamic acids to Tolu Balsam

10. 7. Usage of vegetative material from the same plant.
Stems of Buchu
Bark portion with Cascara or Cinchona,
8. Adulteration of powder
In case of powdered drugs color, texture, and density
of the powder are taken into consideration
irrespective of its originetc1,2,3.

11. CAUSES OF ADULTERATION

12. MEASURE OF ADULTERATION
• Procedures are available to detect the Adulteration.
• It is better to obtain confirmatory evidence by using as many means of detection.
• Such as
1. The identity of adulterant
2. Determine the Quality of the drug
• Raw material should be carefully assessed based on Monograph.
• Monographs are available in different official books and the regulatory guidelines as
per WHO guidelines.

13. MEASURE OF
ADULTERATION
• In Herbal Pharmacopoeia of India has come
out in 1st and 2nd Volumes.
• But no statutory guidelines for detection have
been released.
• In volume 3 of EU Guidelines, Guidelines on
Quality, Safety, and Efficacy have been
provided.
• Even UK has guidelines for a product
containing herbal ingredients1.

14. MEASURE OF ADULTERATION
• If adulterated requires detection of nature of adulteration in the identified drug
• Before use of any plant drug
• Its identity should be thoroughly confirmed by comparing morphological and
microscopic characters
• Listed in Pharmacopoeial monograph comparing characters with authentic drug
from the herbarium.
• Considering the wide variations in source
• Crude drugs their chemical nature
• Biological activity
• Standard by different techniques3.

15. SAMPLING PROCEDURE

18. DETERMINATION OF FOREIGN
MATTER
Foreign matter is material consisting of any of the
following –
1. The biological origin of which is the same as that
specified in the monograph concerned but the
appearance or botanical part is different.
2. The biological origin of which differs from that
specified in the monograph concerned.
3. Foreign mineral matters such as stones, sand, and
lumps of soil4.

19. DETERMINATION OF FOREIGN
MATTER
• Method and Procedure –
1. Weigh 100-500 g of sample and spread in a thin layer. Sort the foreign matter into
groups either by visual inspection, using a magnifying lens (5-10×), or with the help
of a suitable sieve.
2. Weigh each group of foreign matter separately, and calculate the percentage of
foreign matter.
• Example- a) Nigella sativa seed- 0.4%w/w
b) Saraca asoca leaf- 1.5%w/w

20. DNA FINGERPRINTING TECHNIQUES IN
IDENTIFICATION
• Herbal formulations are extensively used as therapeutic agents for the treatment of
several diseases.
• Herbal drug technology is used for converting botanical materials into medicines.
• Standardization is a process of evaluating the quality & purity of crude drugs by
various parameters like morphological, microscopical, physical, chemical
&biological observation.
• Fingerprint analysis approach is the most important in quality control of herbal
medicines because of its accuracy and reliability.

21. • DNA fingerprinting is defined as the
application of genetic/ molecular marker
techniques which is used to identify cultivars.
• It is the process that determines the
concentrations of chemical substances in an
herb by several methods,
• like RFLP, AFLP, RAPD, SSR sequencing of the
rDNA-ITS region, and DNA barcoding.
• Development of fingerprints based on DNA
markers is necessary for proper identification
and standardization of plant species5.

22. TYPES OF DNA FINGERPRINT
TECHNIQUE
1. Hybridization based
• Restriction Fragment Length
Polymorphism (RFLP)
• Variable Number Tandem Repeat
(VNTR)
• Probe hybridization with Micro
and Minisatellite
• Random Genomic Clone
• cDNA Clone

23. TYPES OF DNA FINGERPRINT
TECHNIQUE
2. PCR based
• Inter Simple Sequence Repeat (ISSR)
• Random Amplification Polymorphic DNA
(RAPD)/Arbitrary Primed PCR
• Amplified Fragment Length Polymorphism
(AFLP)
• DNA Amplification Fingerprinting (DAF)

24. TYPES OF DNA FINGERPRINT
TECHNIQUE
3. Sequence based
• Simple Sequence Repeats (SSR)
• Sequence Characterized Amplified
Region (SCAR)
• Cleaved Amplified Polymorphic
Sequence (CAPS)
• Single Nucleotide Polymorphism (SNP).

25. STEPS INVOLVED IN DNA
FINGERPRINTING

26. EXAMPLE OF IDENTIFICATION BY
DNA FINGERPRINTING

27. HEAVY METALS
• Heavy metal like cobalt (Co), copper (Cu), iron (Fe), manganese (Mn),
molybdenum (Mo), nickel (Ni), and zinc (Zn) enters in soil from
various sources such as mining, foundries, smelters, combustion, and
agriculture7.
• Studies indicated that high levels of heavy metals were present in
some herbal medicines.
• Particularly as heavy metals are known to have low renal excretion
rates, potentially resulting in adverse effects in humans even at very
low concentrations.

28. DETERMINATION OF HEAVY METALS
• A total of 1773 crude plant extract samples from 2014 to 2019, representing 86
different kinds of commonly used herbal medicines were collected for examination
of heavy metals.
• The resulting solution was analyzed using a Scientific X Series Inductively-Coupled
Plasma Mass Spectrometer (ICP-MS)6.

29. DETERMINATION OF HEAVY METALS
The most predominant techniques used are
• Atomic absorption spectrometry (AAS)
• Atomic emission/ fluorescence
spectrometry (AES/AFS)
• Inductively coupled plasma mass
spectrometry (ICP-MS)
• Inductively coupled plasma optical
emission spectrometry (ICP-OES)
• Neutron activation analysis (NAA),
• X-ray fluorescence (XRF), etc.

30. PESTICIDAL RESIDUE
• Pesticide residue is defined by the World
Health Organization as “any substance or
mixture of substances in food for man or
animals resulting from the use of a pesticide
and includes any specified derivatives, such as
degradation and conversion products,
metabolites, reaction products, and impurities.

33. CAUSES OF PESTICIDAL RESIDUE

34. DETERMINATION OF PESTICIDAL RESIDUE

35. DETERMINATION OF PESTICIDAL RESIDUE
QuEChERS Method for Pesticide
Residue Analysis
• QuEChERS significantly improves
laboratory efficiency and
throughput. This procedure requires
only small quantities of solvent and
is capable of generating recoveries
of 70-120% with RSDs <5% for a
wide range of compounds.

36. PHYTOTOXIN
• Phytotoxins are substances that are
poisonous or toxic to the growth of
plants. Phytotoxic substances may result
from human activity, as with herbicides, or
they may be produced by plants, by
microorganisms, or by naturally occurring
chemical reactions.
• The toxic principles are the adoption of
plants to prevent themselves from being
consumed by predators and ultimately
promoting their survival9.

37. PHYTOTOXICITY
• Plant injury (phytotoxicity) may occur when chemicals are employed to
protect plants from pests, fertilize plants, regulate plant growth, etc.
• Phytotoxicity can occur when: a material is properly applied directly to the
plant during adverse environmental conditions. a material is applied
improperly.
• Phytotoxicity is defined as a delay of seed germination, inhibition of plant
growth or any adverse effect on plants caused by specific substances
(phytotoxins) or growing conditions8.

39. DETERMINATION OF PHYTOTOXINS
• The preliminary tests determine phytotoxins qualitatively (whether present or not)
or sometimes semi-quantitatively (based on the time required for the onset of
reaction and/or intensity of development of color etc.).
• The confirmation of presumptive tests can be done by modern techniques viz. LC-
MS, GC-MS, HPLC, FTIR, etc. act through the separation and identification of
individual molecules.
• However, these confirmatory tests are more expensive, time-consuming, and require
various equipment which is not available or affordable in all circumstances.
• Therefore, the qualitative tests which are simple, fast, and economic play a crucial
role in the presumptive investigation and field analysis of a large number of samples

44. MICROBIAL CONTAMINATION IN HERBS AND
THEIR FORMULATION
• Microbiological contamination refers to the non-intended or accidental
introduction of infectious material like bacteria, yeast, mold, fungi, virus,
prions, protozoa, or their toxins and by-products.

45. MICROBIAL CONTAMINATION IN
HERBS AND THEIR FORMULATION
• Main safety risks related to herbal medicines is contamination by microorganisms of
various kinds
• Which are adherent to leaves, stems, flowers, seeds, and roots from which herbal
medicines are prepared.
• Alternatively, microorganisms can be introduced during harvesting, handling, open-
air drying, preserving, and manufacturing.
• The risk of microbial contamination may have an exacerbated impact in the elderly
population because this population has increased susceptibility to the consumption
of herbal medicines and sensitivity to health complications due to the aging
process.

46. MICROBIAL CONTAMINATION IN HERBS AND
THEIR FORMULATION
• It has exacerbated the impact on the elderly population.
• Because of increased susceptibility and sensitivity to health complications due to
the aging process10.

51. DETECTION OF CONTAMINANTS
Determination
of
contaminants
Total Viable
count method
Membrane
filtration
method
Serial dilution
method.

53. 1. Total viable
count method

59. REFERENCES
1. Quality control of Herbal Drugs- Pulok Mukerjee
2. Pharmacognosy- C K Kokate, A P Purohit, S B Gokhale.
3. Adulteration and Evaluation of Herbal Drugs- gputtawar.edu.in
4. Appendix VIII: Determination of Foreign Matter.
(http://www.cmd.gov.hk/hkcmms/vol6/eng/E_Appendices_EVIII.pdf)
5. Short Review of DNA Fingerprinting and Standardization of Herbal Drug.
(https://ijmbs.info/index.php/ijmbs/article/view/15)
6. Heavy Metal Contaminations in Herbal Medicines: Determination, Comprehensive Risk Assessments,
and Solutions. (https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7883644/)
7. https://www.frontiersin.org/articles/10.3389/fenvs.2016.00069/full

60. REFERENCES
8. Phytotoxicity- An Overview. (https://www.sciencedirect.com/topics/agricultural-and-biological-
sciences/phytotoxicity)
9. Revie of Phytotoxin and Qualitative tests for their Detedtion.
(https://www.researchgate.net/publication/343392642_A_review_on_phytotoxins_and_qualitative_tes
ts_for_their_detection)
10. Microbial contamination in herbal medicines: a serious health hazard to elderly consumers.
(https://bmccomplementmedtherapies.biomedcentral.com/articles/10.1186/s12906-019-2723-1)
11. Microbial and Heavy Metals Contamination of Herbal Medicines.
(https://scialert.net/fulltext/?doi=jm.2008.683.691)
12. Microbial contamination in herbs and their formulation- PPT