Analysis of Hair in Forensic Toxicology

Analysis of Hair in Forensic Toxicology

Hair is the commonly found evidence on the crime scene related to assault cases or the cases where some sort of struggle happened at the crime scenes. Matching and identification of hair serve a major role in forensic biology and serology. But also, hair serves great importance in forensic toxicology [1,2]. It is an ideal matrix for the identification of drugs and chronic poisoning. Along with the routine analysis of other visceral organs in forensic toxicological analysis, hair must also consider. It has several advantages over the other biological matrices [2].

Advantages of Hair as a Matrix:

  1. The drug can be detected up to 90 days
  2. Indefinite stability
  3. Metabolites can also be detected
  4. A hair sample is quick and easy for a live subject
  5. Noninvasive techniques can be employed for the sample collection
  6. No need for preservatives to preserve the sample for further analysis.
  7. Segmental analysis of hair helps to study the exposure level.
  8. Manipulation of the result is not easy

Disadvantages of Hair as a Matrix:

  1. Amount of drug present is very less detected by only some of the instrumental techniques
  2. Medical history of the patient is required
  3. Require more precautions to handle the hair sample

Toxicological analysis of hair involves four major steps [3,4]:

  1. Collection and preparation of hair sample
  2. Decontamination of the hair sample
  3. Extraction of drug from hair
  4. Analysis of the extract using the analytical technique

Collection, Preparation of Hair Sample

For the analysis of drugs, head hair is preferably collected from near the scalp of the posterior vertex region of head due to uniformity in the growth of hair [3-5]. Around 10-80mg of hair is required for the analysis depending on the method and techniques used. Growth of hair is approximately 1cm/month; thus, the segmental analysis of hair plays an important role to find out the exposure level of a drug (condition is a medical history of the patient should be known). The sample should be collected by trained individuals only without external contamination. Hair samples need not be required to preserve in some preservatives or refrigeration. A hair sample can simply keep in a paper envelop or wrap in aluminum foil at room temperature, at a dry place [3,5]. During the collection of samples ethical, legal, and human rights of the person should be respected [5].

After the collection aim is the extraction of drugs from the hair, it takes lots of time in the extraction and high precision to extract the complete drug without the loss of analyte.

Decontamination Procedure for Hair

Decontamination procedure of hair is done to find out the external contamination in hair it includes dust, oil, sweat, hair care products, lipids, and loosely bound drugs from the environment, which may interfere with the final result and may produce a false-positive result. The decontamination of hair is done by using organic solvents for two to three times. The first wash of hair should be kept for the analysis [3-5].

After decontamination hair strands should be air-dried and then segmented as fine as possible and grind it into a fine powder, this fine powder is subject to extract the drug.

Extraction Procedure for Hair

The extraction of drugs requires high precision accuracy because the amount of drug present in hair is at picogram and nanogram level [3-5]. There are three methods to extract the drug from hair.

  1. Acidic extraction
  2. Basic extraction
  3. Enzymatic extraction

For the extraction of desirable analyte (acidic/basic drug), hair sample is kept overnight in solution (acidic/basic or solution). Followed by extraction procedure such as solid-phase microextraction (SPE/SPME), liquid-liquid extraction (LLE). In enzymatic extraction, digestion of matrix is done by using the enzyme-like β-glucuronidase/arylsulfatase which results in the extraction of drugs from the hair [5]. The extracts thus collected is subject to analysis using the various analytical technique.

Analysis of Analytes Extracted from Hair

The final step for drug identification from hair is the analysis. The analysis of analyte extracted from hair requires very sophisticated and sensitive techniques because the analyte present in hair is in a very little small amount. In the last few years, many researches have been done to analyze the drug in nanogram and picogram level. Gas chromatography- electron ionization/ mass spectrometry (GC-EI/MS), Liquid chromatography-tandem mass spectrometry (LC-MS/MS), LC-MS, Ultrahigh-Performance Liquid Chromatography-Tandem Mass Spectrometry (UHPLC-MS/MS) are the some analytical analyze used to detect an analyte in such small quantity [6-9]. Also, the quantification of drugs present in hair is also possible by applying these techniques.


In the field of forensic toxicology, hair as a matrix should also consider along with the other biological matrix and visceral samples. It has several advantages over the other biological matrices. There are some factors which affect the analysis, these factors should also consider during hair analysis such as external decontamination, hair care product use, hair color, cosmetic treatments, the concentration of dose. There are several studies on hair analysis and methods have been developed, but still, there is a need for more research and study in the same field.


  1. T. Oien, C. A. R. R. Y. (2009, April). Forensic Hair Comparison: Background Information for Interpretation. Forensic Science Communications.
  2. Usman, Muhammad & Naseer, Abid & Baig, Yawar & Jamshaid, Tahir & Shahwar, Muhammad & Khurshuid, Shazia. (2019). Forensic toxicological analysis of hair: a review. Egyptian Journal of Forensic Sciences. 9. 17. 10.1186/s41935-019-0119-5.
  3. Society of Hair Testing (2004). Recommendations for hair testing in forensic cases. Forensic science international, 145(2-3), 83–84.
  4. Cooper, G.A. &Kronstrand, R. &Kintz, P. & T, Society. (2012). Society of hair testing guidelines for drug testing in hair. Forensic Sci. Int. 20.
  5. Khajuria, H., Nayak, B. P., & Badiye, A. (2018). Toxicological hair analysis: Pre-analytical, analytical and interpretive aspects. Medicine, Science and the Law, 58(3), 137-146. doi:10.1177/0025802418768305
  6. Kronstrand, R., Forsman, M., & Roman, M. (2013). A screening method for 30 drugs in hair using ultrahigh-performance liquid chromatography time-of-flight mass spectrometry. Therapeutic drug monitoring, 35(3), 288–295.
  7. Khajuria, H., Nayak, B. P., & Badiye, A. (2018). Toxicological hair analysis: Pre-analytical, analytical and interpretive aspects. Medicine, science, and the law58(3), 137–146.
  8. Cuypers, E., & Flanagan, R. J. (2018). The interpretation of hair analysis for drugs and drug metabolites. Clinical toxicology (Philadelphia, Pa.)56(2), 90–100.
  9. Di Corcia, D., Salomone, A., & Gerace, E. (2018). Analysis of Drugs of Abuse in Hair Samples by Ultrahigh-Performance Liquid Chromatography-Tandem Mass Spectrometry (UHPLC-MS/MS). Methods in molecular biology (Clifton, N.J.)1810, 107–114.
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