journal article·2025-12-07Open access

Study of the methodological system for blood residues with different degrees of degradation on artefacts

Yawen Huang, Guang Huang, Zixuan Wang, Bingbing Yan, Yan Liu, An Du, Rui Wen, Kun Zhang, Fuwei Yang, Xinnan Chen

npj Heritage Science· Volume 13· Issue 1

Abstract

This study aims to develop a methodology for the detection of blood residues in archaeological contexts, providing valuable information about religious tradition, social development, and daily life of ancient society. The testing method system using iron and porphyrin compounds as blood markers is developed. The results show that, for well-preserved blood residues, the non-destructive techniques including Raman spectroscopy (for porphyrin compounds) and micro-area X-ray fluorescence spectroscopy (for iron) are adequate; for blood residues suffering from severe deterioration, a combination of micro-area X-ray fluorescence spectroscopy (for detecting iron), Ultraviolet and visible absorption spectroscopy and reduced phenolphthalein test (for porphyrin compounds) is necessary. The method systems in this study are meaningful and highly applicable for the confirmation of blood with different degrees of degradation on the archaeological artefacts.
Source: openalex_inverted_index · Quality: baseline

Research context

OpenAlex topics: Cultural Heritage Materials Analysis · Forensic and Genetic Research · Building materials and conservation

SciVX discovery tags are terminology-based. OpenAlex topics are source-provided classifications.

References (51)

Show source-provided references
1. Wang, L. et al. Study on the analytical methods of component materials and making techniques of painting and coloured drawing in ancient architecture, Relics Museol. 451−454 (2009).
2. Wang, L. et al. Determination of contents of protein and starch in plaster materials used in coloured drawing on Chinese ancient building. J. Lanzhou Univ. 48, 136–140 (2012).
3. Fu, Y. C., Chen, Z. F., Zhou, S. H. & Wei, S. Y. Comparative study of the materials and lacquering techniques of the lacquer objects from Warring States Period China. J. Archaeol. Sci. 114, https://doi.org/10.1016/j.jas.2019.105060 (2020).10.1016/j.jas.2019.105060 ↗
4. Wang, N. et al. Scientific research on the materials of mortars used for lacquerwares collected in the Palace Museum. Sci. Conserv. Archaeol. 35, 101–110 (2023).
5. Pires, E., Carvalho, L. D. C., Shimada, I. & McCullagh, J. Human blood and bird egg proteins identified in red paint covering a 1000-Year-Old Gold Mask from Peru. J. Proteome Res. 20, 5212–5217 (2021).10.1021/acs.jproteome.1c00472 ↗
6. Charlier, P. et al. Metaproteomic analysis of King Ghezo tomb wall (Abomey, Benin) confirms 19th century voodoo sacrifices. Proteomics 24, 2400048 (2024).10.1002/pmic.202400048 ↗
7. Loy, T. H. Prehistoric blood residues: detection on tool surfaces and identification of species of origin. Science 220, 1269–1271 (1983).10.1126/science.220.4603.1269 ↗
8. Loy, T. H. & Hardy, B. L. Blood residue analysis of 90,000-year-old stone tools from Tabun Cave, Israel. Antiquity 66, 24–35 (1992).10.1017/S0003598X00081011 ↗
9. Seeman, M. F. et al. Evaluating protein residues on Gainey phase Paleoindian stone tools. J. Archaeol. Sci. 35, 2742–2750 (2008).10.1016/j.jas.2008.05.001 ↗
10. Wei, Y.-Y. et al. Haplogroup structure and genetic variation analyses of 60 Mitochondrial DNA Markers in Southern Shaanxi Han Population. Biochem. Genet. 58, 279–293 (2020).10.1007/s10528-019-09942-0 ↗
11. Splendiani, A. et al. The effects of paleoclimatic events on mediterranean trout: preliminary evidences from ancient DNA. PLOS ONE 11, e0157975 (2016).10.1371/journal.pone.0157975 ↗
12. Jin, X., Pan, C. M. & Gao, Q. Research on the influencing factors of benzidine determination by high performance liquid chromatography. Guangdong Chem. Ind. 49, 173–175+178 (2022).
13. Cox, M. A study of the sensitivity and specificity of four presumptive tests for blood. J. Forensic Sci. 36, 1503–1511 (1991).10.1520/JFS13170J ↗
14. Morris, G., Keith, W. & L., K. P. Simplified preliminary blood testing. an improved technique and a comparative study of methods. J. Crim. Law, Criminol. Police Sci. 42, 95 (1951).10.2307/1140307 ↗
15. Higaki, R. S. & Philp, W. M. S. A study of the sensitivity, stability and specificity of phenolphthalein as an indicator test for blood. Can. Soc. Forensic Sci. J. 9, 97−102 (2013).10.1080/00085030.1976.10757252 ↗
16. Zhang, K., Corti, C., Grimoldi, A., Rampazzi, L. & Sansonetti, A. Application of different Fourier Transform Infrared (FT-IR) methods in the characterization of lime-based mortars with oxblood. Appl. Spectrosc. 73, 479–491 (2019).10.1177/0003702818815693 ↗
17. Fraser, D., Deroo, C. S., Cody, R. B. & Armitage, R. A. Characterization of blood in an encrustation on an African mask: spectroscopic and direct analysis in real time mass spectrometric identification of haem. Analyst 138, 4470–4474 (2013).10.1039/c3an00633f ↗
18. Cai, J., Li, W., Shi, M. & Hu, W. B. Interdisciplinary application of chemistry in archaeology: dominant element identification via p-XRF in Buddhist blood scriptures from the Tang Dynasty. Univ. Chem. 37, 223–231 (2022).
19. Xie, J. et al. Spectral properties of blood cell. Anal. Test. Technol. Instrum. 6, 178–181 (2000).
20. Mazel, V. et al. Identification of ritual blood in African artifacts using TOF-SIMS and synchrotron radiation microspectroscopies. Anal. Chem. 79, 9253–9260 (2007).10.1021/ac070993k ↗
21. Miklin-Kniefacz, S., Griesser, M., Parson, W., Pitthard, V. & Stanek, S. Searching for blood in Chinese lacquerware. Stud. Conserv. 59, S252–S253 (2014).10.1179/204705814X13975704320477 ↗
22. Qiu, L., Zhuang, X., Huang, X., Liu, P. & Zhao, X. Study on heavy metals’ influence on buried cultural relics in soil of an archaeological site. Environ. Technol. 42, 1955–1966 (2021).10.1080/09593330.2019.1684569 ↗
23. Tomasini, E. et al. Identification of pigments and binders of a 17th century mural painting (Bolivia). New report on pigments associated with Andean minerals. J. Cult. Herit. 62, 206–216 (2023).10.1016/j.culher.2023.05.030 ↗
24. Vadrucci, M. et al. Characterisation of ancient Roman wall-painting fragments using non-destructive IBA and MA-XRF techniques. X-Ray Spectrom. 49, 668–678 (2020).10.1002/xrs.3178 ↗
25. Young, P., Silence, P. & Lam, T. Microfadeometry of Miss Breme Jones Watercolor with Iron-gall Ink Inscriptions. Microsc. Microanal. 24, 2170–2171 (2018).10.1017/S1431927618011339 ↗
26. de la Codre, H. et al. The use of XRF imaging for the chemical discrimination of iron-gall ink inscriptions: a case study in Stradivari’s workshop. X-Ray Spectrom. 50, 244–252 (2021).10.1002/xrs.3160 ↗
27. Valério, P., Sequeira, S., Dias, V., Encarnação, G. & Araújo, M. F. The composition of roman metals from Moinho do Castelinho and Quinta da Bolacha (Portugal). J. Cult. Herit. 64, 275–281 (2023).10.1016/j.culher.2023.10.018 ↗
28. Qiu, J. et al. UV-Visible absorption spectra and FTIR of healthy people and it spectroscopic analysis. Spectrosc. Spectr. Anal. 40, 1425–1430 (2020).
29. Jung, F., Stopp, G. & Ruckpaul, K. Ultra-violet absorption spectra of haemoglobins from various vertebrates. Nature 207, 990–991 (1965).10.1038/207990a0 ↗
30. Karabaliev, M., Tacheva, B., Paarvanova, B. & Georgieva, R. Change in osmotic pressure influences the absorption spectrum of hemoglobin inside red blood cells. Cells 13, 589 (2024).10.3390/cells13070589 ↗
31. Santos-Velázquez, L. & López-Garriga, J. Insights into Sulfhemoglobin Detection: UV-Vis and Fluorescence Spectroscopy Correlations. J. Chem. 2023, 8854277 (2023).10.1155/2023/8854277 ↗
32. Fu, Y. C. et al. Identification and classification of six kinds of proteinaceous binding media by THM-Py-GC/M. Sci. Conserv. Archaeol. 34, 22–31 (2022).
33. Wang, N. et al. Identification of protein binding media used in chinese cultural relics by pyrolysis-gas chromatography/mass spectrometry. Chin. J. Anal. Chem. 48, 90–96 (2020).
34. Downs, E. F. & Lowenstein, J. M. Identification of archaeological blood proteins: A cautionarynote. J. Archaeol. Sci. 22, 11–16 (1995).10.1016/S0305-4403(95)80160-X ↗
35. Kooyman, B., Newman, M. E. & Ceri, H. Verifying the reliability of blood residue analysis on archaeological tools. J. Archaeol. Sci. 19, 265–269 (1992).10.1016/0305-4403(92)90016-V ↗
36. Leach, J. & Mauldin, R. P. Additional comments on blood residue analysis in archaeology. Antiquity 69, 1020–1022 (1995).10.1017/S0003598X00082570 ↗
37. Tahoun, M., Engeser, M., Svolacchia, L., Sander, P. M. & Müller, C. E. Molecular Taphonomy of heme: chemical degradation of hemin under presumed fossilization conditions. Molecules 28, 4887 (2023).10.3390/molecules28134887 ↗
38. Zhang, K., Zhang, B. & Fang, S. The application history and scientific nature of blood-based materials in traditional Chinese mortar. Sci. Conserv. Archaeol. 25, 94–102 (2013).
39. Kabała, C., Musztyfaga, E., Gałka, B., Łabuńska, D. & Mańczyńska, P. Conversion of Soil pH 1:2.5 KCl and 1:2.5 H2O to 1:5 H2O: conclusions for soil management, environmental monitoring, and international soil databases. Pol. J. Environ. Stud. 25, 647–653 (2016).10.15244/pjoes/61549 ↗
40. Li, X. et al. Rapid determination of chlorophyll and pheophytin in green tea using fourier transform infrared spectroscopy. Molecules 23, https://doi.org/10.3390/molecules23051010 (2018).10.3390/molecules23051010 ↗
41. Mozzi, A., Forni, D., Clerici, M., Cagliani, R. & Sironi, M. The diversity of mammalian hemoproteins and microbial heme scavengers is shaped by an arms race for iron piracy. Front. Immunol. 9, https://doi.org/10.3389/fimmu.2018.02086 (2018).10.3389/fimmu.2018.02086 ↗
42. Wood, B. R., Tait, B. & McNaughton, D. Micro-Raman characterisation of the R to T state transition of haemoglobin within a single living erythrocyte. Biochim. Biophys. Acta 1539, 58–70 (2001).10.1016/S0167-4889(01)00089-1 ↗
43. Wood, B., Caspers, P., Puppels, G., Pandiancherri, S. & McNaughton, D. Resonance Raman spectroscopy of red blood cells using near infrared excitation. Anal. Bioanal. Chem. 387, 1691–1703 (2007).10.1007/s00216-006-0881-8 ↗
44. Rao, S., Bálint, Š, Cossins, B., Guallar, V. & Petrov, D. Raman study of mechanically induced oxygenation state transition of red blood cells using optical Tweezers. Biophys. J. 96, 209–216 (2009).10.1529/biophysj.108.139097 ↗
45. Aguayo, T. et al. Raman and surface enhanced Raman scattering of a black dyed silk. J. Raman Spectrosc. 44, 1238–1245 (2013).10.1002/jrs.4348 ↗
46. Xie, J., F. Identification of Leather by Infrared and Raman Spectroscopy. Prog. Textile Sci. Technol. 36–39 (2018).
47. Holtum, T., Reichenauer, T., Kumar, V. & Schlücker, S. Picosecond UV Kerr-gated Raman spectroscopy with 280 nm laser excitation. J. Raman Spectrosc. 54, 288–295 (2023).10.1002/jrs.6482 ↗
48. Zhou, Z., Cao, C., Liu, Q. & Jiang, R. Hybrid Orbital Deformation (HOD) effect and spectral red-shift property of nonplanar porphyrin. Org. Lett. 12, 1780–1783 (2010).10.1021/ol100434w ↗
49. Wood, B. R., Hammer, L., Davis, L. & McNaughton, D. Raman microspectroscopy and imaging provides insights into heme aggregation and denaturation within human erythrocytes. J. Biomed. Opt. 10, 14005 (2005).10.1117/1.1854678 ↗
50. Dybas, J., Bokamper, M. J., Marzec, K. M. & Mak, P. J. Probing the structure-function relationship of hemoglobin in living human red blood cells. Spectrochim. Acta Part A: Mol. Biomol. Spectrosc. 239, 118530 (2020).10.1016/j.saa.2020.118530 ↗
51. Zhao, L., Yang, X., Min & Huang, T. Synthetic method improvement of Tetra (4-Aminophenyl) Porphyrin and Metallo-porphyrin Complex. Chem. Reag. 37, 843–847 (2015).