Illuminating Blood Evidence: ATR FT-IR Spectroscopy and its Role in Forensic Science

Forensic analysis is a hot topic in spectroscopy. Because of the ongoing need to analyze evidence and its spectral data in criminal investigations, spectroscopy is being used regularly, and its applications are only increasing.

What is also furthering this trend is the development of portable instrumentation. Spectrometers are being built smaller and smaller, while still containing the necessary capabilities of their larger counterparts (1,2). Many of the more common spectroscopic techniques, including near-infrared (NIR), mid-infrared (mid-IR), and Raman spectroscopy, as well as elemental techniques, such as X-ray fluorescence (XRF) and laser-induced breakdown spectroscopy (LIBS), are becoming miniaturized, and this has created many benefits for researchers, including the ability to conduct analysis on-site (2).

A close-up view of a wet street with a yellow crime scene tape and red and blue lights reflecting in a puddle. Generated by AI. | Image Credit: © kinara art design - stock.adobe.com

This is especially important for crime scenes, as many forensic analysts prefer to conduct their analysis on-site to prevent possible contamination or alteration during transport. A recent review article touched upon this trend by exploring how IR spectroscopy and attenuated total reflectance Fourier transform infrared (ATR FT-IR) spectroscopy is shaping modern forensic science by offering precise, non-destructive methods for examining blood evidence (3).

Bloodstains often serve as critical evidence in solving crimes, providing information about the events surrounding a crime scene. This review article examined the role of ATR FT-IR spectroscopy in analyzing the biochemical composition of blood, differentiating it from other substances, identifying species, and estimating the time since deposition. Drawing on studies published from 2008 to the present, the authors provide a detailed account of the advancements, challenges, and future directions for this promising technology (3).

ATR FT-IR spectroscopy uses the principles of IR light absorption to identify molecular compositions (3). When applied to forensic blood analysis, ATR FT-IR spectroscopy can detect key biochemical markers such as proteins, lipids, and carbohydrates (3). What makes this technique useful for this type of analysis is that is highly sensitive, which makes it simple for forensic scientists to distinguish blood from other biological and non-biological substances (3).

That is not the only benefit that ATR FT-IR spectroscopy provides. It can also identify the species of origin for bloodstains, which is a critical factor in criminal investigations involving multiple parties or non-human sources (3). As a result, ATR FT-IR is more versatile than other techniques for this type of analysis.

The researchers also discuss in their review how integrating chemometrics with ATR FT-IR spectroscopy has also helped advance forensic science. Chemometrics applies advanced statistical and mathematical tools to extract meaningful insights from complex data sets (3). By leveraging multivariate statistical methods, forensic analysts can achieve highly accurate qualitative and quantitative assessments of blood evidence (3).

Despite the advantages of ATR FT-IR spectroscopy, the review identifies several obstacles that the industry continues to face. Key challenges include issues with sample preparation, such as interference from substrates like fabrics or other materials that may alter spectroscopic readings (3). Additionally, the lack of standardized protocols for ATR FT-IR applications in forensic investigations remains a significant barrier. Standardization is essential to ensure consistent, reproducible results across different laboratories and practitioners (3).

There are several proposed strategies to overcome these limitations. The authors stated that developing robust sample preparation techniques and minimizing substrate interference will be important in solving these challenges (3). Additionally, creating standardized protocols and databases for forensic ATR FT-IR spectroscopy will enhance the reliability of the technique (3).

The review also calls for further research into combining ATR FT-IR spectroscopy with other vibrational spectroscopic techniques, such as Raman and NIR spectroscopy. These hybrid approaches could overcome current limitations and expand the technique’s forensic capabilities (3).

Recent advances in vibrational spectroscopy, including ATR FT-IR, Raman, and NIR techniques becoming portable, have helped advance forensic research by offering cost-effective, non-destructive methods for analyzing evidence. With its ability to provide reliable, detailed insights into blood evidence, ATR FT-IR spectroscopy has great potential in crime prevention and forensic analysis (3).

References

1. Wetzel, W. Portable NIR Spectrometers Enhance Flexibility in Forensic Science. Spectroscopy. Available at: https://www.spectroscopyonline.com/view/portable-nir-spectrometers-enhance-flexibility-in-forensic-science (accessed 2025-01-06).
2. Crocombe, R. Spectrometers in Wonderland: Shrinking, Shrinking, Shrinking. Spectroscopy Suppl. 2022, 37 (s11), 6–11. Available at: https://www.spectroscopyonline.com/view/spectrometers-in-wonderland-shrinking-shrinking-shrinking (accessed 2025-01-06).
3. Al-Hetlani, E.; Al-Qalfas, M.; Amin, M. O. The Application of Attenuated Total Reflectance-Fourier Transform Infrared Spectroscopy in Forensic Bloodstain Analysis: A Review of Recent Advances. Microchem J. 2025, 112655. DOI: 10.1016/j.microc.2025.112655