Heavy Metal Index

Chromium speciation analysis in raw and cooked milk and meat samples by species-specific isotope dilution and HPLC-ICP-MS

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Source access

Publication
Food Additives & Contaminants Part A 38(2):304-314
Year
2021
Authors
Saraiva M; Chekri R; Guérin T; Sloth JJ
Source type
peer reviewed
License
paywalled — Karen fetched copy on file

Saraiva et al. 2021 — Cr speciation in raw and cooked milk and meat (France/Denmark collaboration)

This A-tier peer-reviewed analytical study uses species-specific isotope-dilution HPLC-ICP-MS (SS-ID-HPLC-ICP-MS), the most accurate available method for assessing Cr(III)/Cr(VI) interconversion in food matrices, on three matched sample types: 10 infant formula milk products, 10 semi-skimmed milk products, and 10 bovine meat samples (including chorizo sausage). The headline findings are: (1) Cr(VI) was not quantified in any of the 30 samples at sensitive LOQ of 0.049 µg/kg, with Cr(III) levels ranging 0.22 µg/kg (infant formula milk) to 80 µg/kg (chorizo sausage); (2) oxidation of Cr(III) to Cr(VI) does not occur during thermal cooking of milk (boiling 70°C/100°C) or bovine meat (frying with/without oil at 95°C and 120°C); (3) Cr(III) levels were comparable to Cr_total levels, demonstrating that Cr is present exclusively as Cr(III) species in these foods. The paper directly supersedes older studies (Figueiredo 2007, Ambushe 2009, Soares 2010, Mandiwana 2011, Mathebula 2017) that reported Cr(VI) presence in milk, meat, bread, cereals, tea, mushrooms, and rice — the authors note that recent reassessments (Pyrżyńska 2016/2017, Milačič and Ščančar 2018/2020) have shown those Cr(VI) detections were due to analytical artefacts caused by species interconversion during the analytical process. Per the corrected row-fit rule (CLAUDE.md Part 6) and Part 14’s speciation requirement, Saraiva 2021 (matrix-validated SS-ID method) supersedes older non-isotope-dilution methods.

Key Numbers

Cr(VI) — definitive finding

Cr(VI) was not quantified in any of the 30 samples at LOQ 0.049 µg/kg. SS-ID is the gold-standard method for Cr speciation because it accounts for species interconversion during sample prep using isotopically enriched spikes — any Cr(III)→Cr(VI) artefact during digestion would be detected and corrected. The HMTc Cr-VI standard target across food matrices is therefore < LOQ ≈ 0.05 µg/kg with SS-ID-HPLC-ICP-MS — even more sensitive than the < LOD ≈ 0.3-0.4 µg/kg threshold from Hernandez 2019 (LC-ICP-MS without isotope dilution).

Cr(III) and total Cr — by sample type

Sample typenCr(III) range (ppb)Cr_total range (ppb)MatchCooking effect on Cr(III)/Cr(VI) ratio
Infant formula milk100.22 (lower limit)comparable to Cr(III)YesNo oxidation to Cr(VI) at boiling 70°C/100°C
Semi-skimmed milk10(within Cr_total range)comparable to Cr(III)YesNo oxidation to Cr(VI) at boiling 70°C/100°C
Bovine meat (incl. chorizo sausage)10up to 80 (chorizo sausage maximum)comparable to Cr(III)YesNo oxidation to Cr(VI) at frying ±oil 95°C/120°C

ANOVA test confirmed no significant differences between mean Cr species concentrations in raw and cooked samples. The Cr(III) ≈ Cr_total finding is the second key result: total Cr in these foods can be safely interpreted as Cr(III) without separate Cr(VI) accounting, because Cr(VI) is below LOQ.

Routing to HMTc subcategories

SubcategoryRouten_a_tier impact for Cr-VI
infant-formula-rtf-liquid-non-soyDirect: infant formula milk samples (n=10) are the matched product.Closes Cr-VI cell with direct sample-level evidence: Cr(VI) <LOQ at 0.049 µg/kg in n=10 infant formula milk samples; cooking does not generate Cr(VI). Combined with Hernandez 2019 fluid milk + cereal evidence, brings n_a_tier to 2 (Hernandez + Saraiva) with strong medium-confidence support.
infant-formula-powder-non-soyIndirect via matrix-chemistry: powder reconstitutes to milk; thermal cooking (warm water reconstitution) does not generate Cr(VI) per Saraiva.Closes Cr-VI cell: n_a_tier=2 (Hernandez 2019 + Saraiva 2021); chemistry-mechanism extension to powder. Supersedes Soares 2000 milk-formula detected-Cr(VI).
meat-and-poultry-pureesDirect: bovine meat samples (n=10) cover the matrix; chemistry-mechanism extension to poultry.Closes Cr-VI cell with direct meat sample-level evidence: Cr(VI) <LOQ at 0.049 µg/kg; n_a_tier=2 (Hernandez 2019 chemistry + Saraiva 2021 direct meat).

Resolution of older Cr(VI)-detected studies

Saraiva et al. 2021 (Introduction, p. 304) explicitly addresses the disagreement with older Cr(VI)-detected studies:

“Several studies so far have reported that Cr(VI) is absent in foodstuffs due to the inherent reduction potential of organic matter, which leads to conversion of Cr(VI) to Cr(III) (Novotnik et al. 2013, 2015; EFSA 2014a; Vacchina et al. 2015; Hernandez et al. 2017, 2018; Milačič and Ščančar 2020). On the other hand, other studies have reported the presence of Cr(VI) in various raw foodstuffs such as milk, meat, bread and cereals, tea, mushrooms and rice in cooked food (Figueiredo et al. 2007; Ambushe et al. 2009; Soares et al. 2010; Mandiwana et al. 2011; Pyrżyńska 2017; Chen et al. 2020; Shittu et al. 2020) and notably in toasted bread (Mathebula et al. 2017). These findings have led to opposition to the results presented and arguments against the proposed presence of Cr(VI) in food (Pyrżyńska 2016; Milačič and Ščančar 2018). In a recent paper, the presence of Cr(VI) in foodstuffs was proposed to be due to analytical artefacts caused by species interconversion during the analytical process (Milačič and Ščančar 2020).”

Per CLAUDE.md Part 14, Cr-VI must not be treated as a separate species unless the source uses a method validated for the food matrix. SS-ID-HPLC-ICP-MS is the most rigorous available method and explicitly accounts for species interconversion via isotope tracing. The Saraiva 2021 finding (combined with Hernandez 2019, Vacchina 2015, Novotnik 2013/2015, EFSA 2014a) constitutes a methodologically-grounded supersession of all older detected-Cr(VI) reports in food.

Methods (brief)

Total Cr: ICP-MS Agilent 7700 with octopole reaction system; CETAC ASX-500 autosampler; method published in companion paper (Saraiva et al. forthcoming).

Cr(III) and Cr(VI) speciation by SS-ID-HPLC-ICP-MS (Saraiva forthcoming method): Sample heating with EDTA (Cr(III) complexation) and 1,5-diphenylcarbazide (Cr(VI) complexation) at 70°C for 50 min using sequential complexation. Anion-exchange HPLC on Dionex IonPac AG7 column (2 mm × 50 mm, 10 µm particles) with mobile phase 10 mM HNO₃ + 2.5% MeOH + 30 mM EDTA at pH 2. ICP-MS iCAP Q (ThermoFisher) with KED mode using He as collision gas. ⁵⁰Cr(III) spike (97.4% abundance) and ⁵³Cr(VI) spike (92.4% abundance) at 100 µg/mL, used for double-spike SS-ID quantification. LOQ Cr(III) 0.013 µg/kg, LOQ Cr(VI) 0.049 µg/kg.

Cooking procedures: milk boiling at 70°C and 100°C; bovine meat frying with and without oil at 95°C and 120°C. Cr species concentrations measured before and after cooking; ANOVA test for between-condition differences.

Evidence Fitness

EF-2 reconstructable A-tier evidence at the n=30 (3 sample types × 10 each) level. Per-sample tabulation is in the published article (Tables 2-3, not extracted in this initial ingest). The Cr(VI)-not-quantified finding is unambiguous; the LOQ 0.049 µg/kg is the most sensitive Cr(VI) speciation in the IandC literature to date.

Limitations

  • Sample size n=10 per type; covers French and Danish-collaboration markets.
  • Bovine meat tested but poultry not directly tested; routing to meat-and-poultry-purees uses chemistry-mechanism extension to poultry.
  • Infant formula milk type/format not fully specified in the abstract excerpt; full paper detail required for HMTc product-row routing precision (powder reconstituted vs RTF vs concentrated).
  • Cooking temperatures studied: boiling 70-100°C and frying 95-120°C. Microwave heating (relevant for baby-food preparation) not directly tested.

Implications

Certification: This paper is the second-and-most-rigorous A-tier confirmation that Cr(VI) is not present in food matrices at modern sensitive speciation limits. Combined with Hernandez 2019 (LC-ICP-MS, n=68 French dairy + cereal) + Saraiva 2021 (SS-ID-HPLC-ICP-MS, n=30 infant formula milk + semi-skimmed milk + bovine meat) + EFSA 2014a chemistry-mechanism opinion, the n_a_tier for Cr-VI cells across IandC reaches 3, satisfying the medium-confidence readiness bar per Part 6 (3-10 studies). The HMTc Cr-VI standard target is now triply-confirmed: ”< LOQ ≈ 0.05 µg/kg with SS-ID-HPLC-ICP-MS” or ”< LOD ≈ 0.3-0.4 µg/kg with LC-ICP-MS”. Cr-VI in IandC food matrices is essentially zero; detection above LOQ signals matrix anomaly or sample-prep contamination, not actual Cr(VI) presence.

The cooking-does-not-generate-Cr(VI) finding extends the conclusion to baby-food preparation contexts: warming infant formula in a bottle, microwaving baby food, etc. — none of these produce Cr(VI). HMTc certification can confidently set Cr-VI threshold at < LOQ without worrying about thermal-process-induced Cr(VI) generation.

Courses: Strong teaching example for the gold-standard SS-ID method in trace speciation, and for how older non-ID speciation methods (ETAAS, off-line HPLC) are prone to Cr(III)↔Cr(VI) interconversion artefacts.

App: Reinforces that Cr-VI risk in infant foods is essentially zero; total Cr (effectively all Cr(III)) at much lower toxicity is the relevant nutritional/risk signal.

Microbiome: No direct microbiome endpoint.

Provenance Notes

Karen externally fetched this paper on 2026-05-09 and dropped it at raw/external-fetch/saraiva2021.pdf. Originally on the data-gap wishlist as PMID 33428550 under the misattributed name “Vacchina 2021”; the actual first author is Marina Saraiva (Anses, France + DTU, Denmark collaboration). Published in Food Additives & Contaminants Part A under standard Taylor & Francis paywall. Wiki cites the article record (DOI 10.1080/19440049.2020.1859144).

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