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International Journal of Hygiene and Environmental Health xxx

(xxxx)

xxx

Contents lists available at

ScienceDirect

International Journal of Hygiene and Environmental Health

journal homepage:

www.elsevier.com/locate/ijheh

The SAM-Krom biomonitoring study shows occupational exposure to

hexavalent chromium and increased genotoxicity in Denmark

Anne Thoustrup Saber

a,*

, Marcus Levin

, Pete Kines

, Kukka Aimonen

, Lucas Givelet

Christina Andersen

, Anja Julie Huusom

, Tanja Carøe

, Niels Erik Ebbehøj

Frans Møller Christensen

, Zheshun Jiang

, Thomas Lundh

, Håkan Tinnerberg

Maria Albin

h,j

, Malin Engfeldt

h,k

, Karin Broberg

a,h

, Julia Catalan

c,l

, Katrin Loeschner

Karsten Fuglsang

, Ulla Vogel

a,d

National Research Centre for the Working Environment, 105 Lersø Parkall

, DK-2100, Copenhagen

Ø,

Denmark

FORCE Technology, 345 Park All

, DK-2605, Brøndby, Denmark

Finnish Institute of Occupational Health, 00250, Helsinki, Finland

National Food Institute, Technical University of Denmark, DK-2800, Kgs. Lyngby, Denmark

Department of Occupational and Environmental Medicine, Copenhagen University Hospital, Bispebjerg and Frederiksberg, Denmark

Department of Occupational and Social Medicine, Holbæk University Hospital, DK-3400, Holbæk, Denmark

COWI A/S, Parallelvej 2, DK-2800, Kgs. Lyngby, Denmark

Division of Occupational and Environmental Medicine, Department of Laboratory Medicine, Lund University, Lund, Sweden

Occupational and Environmental Medicine, School of Public Health and Community Medicine, Institute of Medicine, University of Gothenburg, Sweden

Unit of Occupational Medicine, Institute of Environmental Medicine, Karolinska Institutet, Sweden

Department of Occupational and Environmental Medicine, Region Skåne, SE-22381, Lund, Sweden

Department of Anatomy, Embryology and Genetics, University of Zaragoza, 50013, Zaragoza, Spain

A R T I C L E I N F O

Keywords:

Hexavalent chromium

Bath platers

Welding

Biomarkers

Occupational exposure limits

Vocational schools

A B S T R A C T

Background:

Hexavalent chromium (Cr(VI)) is a carcinogen. Exposure to Cr(VI) may occur in different industrial

processes such as chrome plating and stainless steel welding. The aim of this study was to assess occupational

exposure to Cr(VI) in Denmark.

Methods:

This cross-sectional study included 28 workers and 8 apprentices with potential Cr(VI) exposure and 24

within company controls, all recruited from six companies and one vocational school. Use of occupational safety

and health (OSH) risk prevention measures were assessed through triangulation of interviews, a questionnaire

and systematic observations. Inhalable Cr(VI) and Cr-total were assessed by personal air exposure measurements

on Cr(VI) exposed participants and stationary measurements. Cr concentrations were measured in urine and in

red blood cells (RBC) (the latter reflecting Cr(VI)). Genotoxicity was assessed by measurement of micronuclei in

peripheral blood reticulocytes (MNRET).

Results:

At announced visits, a consistent high degree of compliance to OSH risk prevention measures were seen

in ‘chromium bath plating’ for both technical devices (e.g. ventilation, plastic balls, sheet coverings) and in the

use of personal protective equipment (e.g. gloves, respirators), yet a lesser degree of compliance was observed in

‘stainless steel welding’. The geometric mean of the air concentration of Cr(VI) was 0.26

g/m

(95% confidence

interval (CI): 0.12–0.57) for the Cr(VI)-exposed workers and 3.69

g/m

(95% CI: 1.47–9.25) for the Cr(VI)-

exposed apprentices. Subdivided by company type, the exposure levels were 0.13

g/m

(95% CI: 0.04–0.41)

for companies manufacturing and processing metal products, and 0.81

g/m

(95% CI: 0.46–1.40) for bath

plating companies. Workers with occupational exposure to Cr(VI) had significantly higher median levels of

urinary Cr (2.42

g/L, 5th-95th percentile 0.28–58.39), Cr in RBC (0.89

g/L, 0.54–4.92) and MNRET (1.59

‰,

0.78–10.92) compared to the within company controls (urinary: 0.40

g/L, 0.16–21.3, RBC: 0.60

g/L,

0.50–0.93,MNRET: 1.06

‰,

0.71–2.06). When sub-dividing by company type, urinary Cr (4.61

g/L, 1.72–69.5),

Cr in RBC (1.33

g/L, 0.95–4.98) and MNRET (1.89

g/L, 0.78–12.92) levels were increased for workers with

potential Cr(VI) exposure in bath-plating companies, and when subdividing by work task, workers engaged in

* Corresponding author.

E-mail address:

[email protected]

(A.T. Saber).

https://doi.org/10.1016/j.ijheh.2024.114444

Received 11 July 2024; Received in revised form 19 August 2024; Accepted 20 August 2024

(

http://creativecommons.org/licenses/by/4.0/

Please cite this article

as:

Anne Thoustrup

https://doi.org/10.1016/j.ijheh.2024.114444

Saber

al.,

International

Journal

Hygiene

and

Environmental

Health,

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A.T. Saber et al.

International Journal of Hygiene and Environmental Health xxx

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process operation had increased levels of urinary Cr (8.51

g/L, 1.71–69.5), Cr in RBC (1.33

g/L, 0.95–4.98)

and MNRET (1.89

g/L, 0.82–12.92) levels.

Conclusion:

This biomonitoring study shows that bath platers were highly exposed to Cr(VI), as suggested by

relatively high levels of urinary Cr, Cr in RBC and increased levels of micronuclei. The urinary Cr concentrations

were high when compared to the French biological limit value of 2.5

g Cr/L, corresponding to the Danish

occupational exposure limit of 1

g/m

. This, in turn, indirectly suggests that additional exposure routes than via

air may contribute to the exposure. For welders, no statistically significant increases compared to within com-

pany controls were observed, however, the observed urinary Cr levels were similar to the levels observed in a

European study (HBM4EU), and were higher than the levels observed for welders in Sweden (SafeChrom). In

spite of a high degree of self-reported and observed compliance to OSH risk prevention measures during

announced visits, the biomarkers of exposure reflecting recent exposure (urinary Cr) or exposure during the last

four months (Cr in RBC) may point to variation in compliance to OSH risk prevention measures in general.

Reduced occupational exposure to Cr(VI) may be achieved by applying the hierarchy of controls in eliminating or

substituting Cr(VI), and the use of more effective technical solutions (e.g. automation).

Abbreviations

BOEL

Biological occupational limit

Confidence interval

Chromium

Cr(III)

Trivalent chromium

Cr(VI)

Hexavalent chromium

Tot

Total amount of chromium

DISCO-08 Danish International Standard Classification of

Occupation 2008

The European Union

EPA

Environmental Protection Agency

G-EQUAS German External Quality Assessment Scheme

Geometric mean

GSP

Gesamtstaubprobenahme sampler

HBM4EU The European Human Biomonitoring Initiative

IARC

International Agency for Research on Cancer

ICP-MS Inductively coupled plasma mass spectrometry

LEV

LOD

LOQ

MAG

MIG

MMA

MNRET

NACE

OSH

OEL

P95

PPE

RBC

RBC-Cr

RPE

SOP

TIG

Local exhaustion ventilation

Limit of detection

Limit of quantification

Metal active gas

Metal inert gas

Manual metal arc

Micronuclei in peripheral blood reticulocytes

National version of EU’s nomenclature

Occupational safety and health

Occupational exposure limit

5th percentile

95th percentile

Personal protective equipment

Red blood cells

Chromium concentration in red blood cells

Respiratory protective equipment

Standard operating procedure

Tungsten inert gas

1. Introduction

Hexavalent chromium (Cr(VI)) is a carcinogen (IARC Group 1)

(IARC,

2012).

Exposure to Cr(VI) may occur at different industrial

processes such as chrome plating and stainless steel welding (IARC,

2012).

The main routes of occupational exposure to Cr(VI) are inhala-

tion and dermal contact (IARC,

2012).

In addition, hand to mouth

contact exposure may result in gastrointestinal tract exposure (Beattie

et al., 2017).

In the European Union (EU), occupational exposure to Cr(VI) is

regulated both at the European level and by national occupational

exposure limits (OEL) in some of the member states. The current EU OEL

for Cr(VI) is 10

g/m

(IFA,

2024)

which will be further reduced to 5

g/m

in 2025 (Santonen

et al., 2022).

Sweden has an OEL of 5

g/m

and France, the Netherlands and Denmark have implemented an OEL of

g/m

(IFA,

2024).

The Danish OEL is expected to be further lowered

to 0.25

g/m

in 2025 if deemed technically and economically feasible

(Beskæftigelsesministeriet,

2020).

Chromium (Cr) in the urine is often used as a biomarker of internal

exposure to Cr(VI) (Verdonck

et al., 2021; Viegas et al., 2022).

addition to the regulation regarding airborne exposure to Cr(VI), some

European countries have also implemented biological occupational

exposure limits (BOELs) using Cr in urine as a biomarker of Cr(VI).

France and Finland have implemented BOELs of 2.5

g/L and 10

g/L,

corresponding to their OELs of 1 and 5

g/m

, respectively (Santonen

et al., 2022).

However, Cr in urine reflects both exposure to Cr(VI) and

trivalent chromium (Cr(III)).

The chromium content in the red blood cells (RBC) has been used as a

specific marker for the Cr(VI) exposure because only Cr(VI) and not Cr

(III) can cross the erythrocyte membrane (Devoy

et al., 2016).

The

content of Cr in the erythrocyte reflects the accumulated exposure over

the previous 4 months, corresponding to the lifetime of erythrocytes

(Ndaw

et al., 2022).

Besides internal exposure biomarkers, some early biological effects

can also be assessed from the same human samples, i.e., blood. Flow

cytometric analysis of micronuclei in peripheral blood reticulocytes

(MNRET) is a sensitive high-throughput method for detection of geno-

toxicity in biomonitoring studies (Abramsson-Zetterberg

et al., 2000).

MNRET reflect genotoxicity in bone marrow approximately three days

prior to sample collection. The lifespan of human reticulocytes in blood

circulation is only 1–4 days and micronucleated reticulocytes are effi-

ciently removed by the spleen. Recently, MNRET was successfully used

within the European Human Biomonitoring Initiative (HBM4EU) as a

short-term biomarker of genotoxicity from low Cr(VI) exposure, and

together with other effect biomarkers it contributed to identifying

occupational subgroups that are at increased cancer risk (Tavares

et al.,

2022).

MNRET has also been used for studying the potential genotox-

icity of pesticides (Costa

et al., 2011),

disinfection by-products

(Font-Ribera

et al., 2019),

industrial pollution (Montero-Montoya

et al., 2020),

and polycyclic aromatic hydrocarbons (Andersen

et al.,

2021).

The current SAM-Krom biomonitoring study was initiated in

response to concern in Denmark regarding the risk of occupational

exposure to Cr(VI) in the working environment. The overall purpose of

the project was to assess occupational exposure to Cr(VI) and investigate

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whether occupational safety and health (OSH) risk prevention measures

aiming at minimizing workers’ exposure to Cr(VI) were present. The

present paper describes the results of this Danish Cr(VI) study and adds

to a systematic review of biomonitoring data on occupational exposure

to Cr(VI) (Verdonck

et al., 2021),

the recent European HBM4EU Cr(VI)

study (Santonen

et al., 2022)

and the Swedish SafeChrom project (Jiang

et al., 2024).

2. Materials and methods

2.1. Study design and study participants

The study is a cross-sectional study carried out in Denmark. The

study and the questionnaire were designed to be as similar and com-

parable as possible to the two recently published chromium studies: 1)

the HBM4EU study with workers from eight European countries

(Santonen

et al., 2022),

and 2) the SafeChrom study with workers in

Sweden (Jiang

et al., 2024).

The present study contains: 1) Interview

data on company OSH strategy, 2) questionnaire data on self-reported

information from workers and apprentices on work tasks, use of per-

sonal protective equipment (PPE) and life style habits, 3) data from

systematic observations of the use of OSH risk prevention measures, 4)

biomonitoring of biomarkers of exposure and effect in blood and urine,

as well as personal air exposure measurements carried out in five com-

panies and a vocational school, and 5) personal air exposure measure-

ments on workers from one additional workplace and

apprentices/teachers from a vocational school for whom no biological

sampling was performed. The vocational school offers both educations

with and without Cr(VI) exposure. Thus, apprentices with possible Cr

(VI) exposure were recruited for air measurements of Cr(VI) exposure

while apprentices in educations without Cr(VI) exposure were recruited

at a later stage as within company controls.

2.1.1. Recruitment of companies and processes

In the initial stage of the project, a mapping of existing knowledge on

exposure to Cr(VI) in Denmark was performed (Højriis

et al., 2020).

Based on the mapping and a pilot study performing air measurements at

different companies and at different processes, we selected companies

and processes meeting the following criteria: 1. The work entails a risk of

exposure to Cr(VI) and 2. The work task is present at a larger number of

Danish workplaces or exposure occurs repeatedly for the workers

involved. Four work tasks were identified: 1) chromium-containing steel

welding, 2) production of metal-containing products and work with

chromates, 3) thermal spraying, and 4) bath plating (Koponen

et al.,

2021).

Regarding welding type, it was decided to focus on metal active

gas (MAG) welding and

–

to the extent possible

–

manual metal arc

(MMA) welding, while omitting tungsten inert gas (TIG) welding, as the

measurements on TIG welding in the pilot study confirmed the

assumption that TIG welding does not entail heavy exposure to Cr(VI)

due to its low mass emission rate and Cr(VI) conversion factor (Fuglsang

et al., 2011; Serageldin and Reeves, 2009).

For the main study, a total number of 29 companies (including four

bath-plating companies and 25 companies with manufacturing/pro-

cessing activities) and one vocational school were contacted by phone.

For the companies that met the inclusion criteria and showed immediate

interest in the study, this was followed up by an e-mail with more details

on the study. Twelve of the 25 manufacturing companies contacted were

omitted, as they were only doing TIG welding in stainless steel. Fifteen of

the remaining, relevant companies (3 bath plating and 12 manufacture/

processing) and the vocational school showed preliminary interest in

taking part in the study. However, due to the COVID 19 pandemic, for

some of the companies there was a delay of 1–2 years from the initial

contact with the companies to the time point when it was possible to

perform the measurements. Meanwhile, the COVID 19 pandemic had

forced some of the companies to change their production or to reduce

their number of workers. Six of the companies (three bath-plating and

three manufacture/processing companies) and the vocational school

(MMA welding, black smiths and electricians) agreed to participate in

the study.

2.1.2. Participants recruited for biomonitoring and personal air exposure

assessment

For the biomonitoring part of the study, potentially Cr(VI) exposed

participants were recruited from five companies with potential Cr(VI)

exposure-related activities such as welding or machining in stainless

steel or chromium bath plating. Unexposed participants were recruited

from the same companies among office workers with no activities

related to Cr(VI) exposure (“within company controls”) and from a

vocational school with fields of study not involving activities with Cr(VI)

exposure (“vocational school controls”). An overview of the recruited

exposed and controls stratified by work task on the day the measurement

was performed is presented in

Table 1.

The biomonitoring study was approved by The Scientific Ethics

Committee for the Copenhagen Capital Region (H-20077777). All par-

ticipants were informed orally and received an information folder about

the study before signing a written consent.

2.1.3. Participants only with personal air exposure assessment

In addition to the biomonitoring part of the study, personal air

exposure measurements were performed on the following two groups: 1)

Apprentices from a stainless steel blacksmith training vocational school,

and, 2) Workers from a company with MAG welding activities. The

reasons for only performing personal air exposure assessment and no

collection of biological samples for this group were that: 1) we did not

have an ethical permission for recruiting apprentices for a bio-

monitoring study at the time of these measurements, and 2) the group at

the company consisted almost entirely of non-Danish speaking staff. Our

information material accepted by the ethical committee was in Danish.

2.2. Categorisation of companies and work tasks for exposed workers

The companies were categorized according to the Danish Industrial

Classification (In Danish

“Dansk

Branchekode DB07” (Statistics

Denmark, 2014)

which is a 6-digit classification:

“Dansk

Branchekode

DB07 is the National version of EU’s nomenclature (NACE). The first four

digits refer to NACE rev. 2, while the last two represent the Danish subdivi-

sion”

(Statistics

Denmark, 2014).

The SAM-Krom classification of the

companies was based on the first 3-digits (Supplementary

Table 1).

The job functions for exposed workers were classified according to

Statistics Denmark’s Classification of Occupations (DISCO-08), v1:2010

(Statistics

Denmark, 2010).

DISCO-08 is a 6-digit classification, but we

chose to use only the 3 first digits for our classification (Supplementary

Table 2).

2.3. Interview and questionnaire data

Company leaders with responsibility for OSH were interviewed

regarding company OSH strategy, instruction and training, and pro-

cedures for handling Cr(VI). On the day of the biological sampling, most

participants answered a questionnaire regarding lifestyle habits (e.g.

smoking), work tasks, and use of PPE. Questionnaire items included the

respondents’ use of PPE during various work tasks (chrome [bath]

plating; grinding; processing of surfaces with Cr(VI) content;

manufacturing of metal products containing Cr(VI); office worker visit

to the shop floor) both during the past week and the past three months,

and included fresh air fed respirators (independent), powered air pur-

ifying respirators (without fresh air), reusable (e.g. half and full face

masks) and disposable (single use) respirators, coveralls, gloves and

aprons. For tasks involving welding and hard chromium plating/thermal

spraying, the PPE included welding helmets with or without respirators

as well as fire retardant clothing.

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Table 1

Categorisation in SAM-Krom of companies/vocational school (n

7) and work tasks for individuals (n

36, exposed group and n

24, control group) for whom biological and/or air sampling were performed.

Total # of individuals

2.4. Systematic safety observations

Systematic observations of exposure control initiatives were regis-

tered in the free app

“Safety

Observer” (nfa.dk/Safetyobserver), where a

tailored list of mandatory chemical exposure control initiatives was

drawn up for visual observation during worksite walk-arounds (Kines

et al., 2010, 2013; Kirkegaard et al., 2018; Nielsen et al., 2015).

The

systematic observations were performed by one member of the research

team at the same time as the initial biomonitoring measurements. The

observation list consisted of Cr(VI) control initiatives in regards to six

topics: 1) order and tidiness (workspaces, disposal areas, access and

escape routes); 2) physical conditions for working with Cr(VI) (safety

signs, labelling, storage, waste); 3) welfare measures (changing, bathing

and dining rooms); 4) first aid (signs and supplies); 5) personal protec-

tive equipment (airway, skin, eyes, ears); 6) technical devices (ventila-

tion, bubble dispensers, mist suppressants, screens and curtains). One

observation marking per object, area (max 50 m

), machine, tool, per-

son, etc. was scored as either ‘correct’ or ‘not correct’. Notes and photos

could be added to the observations. Upon completion of the

walk-around in the areas where there were ongoing activities, the app

generated a report and a safety index (%) based on the percent of correct

observations from the total number of observations.

2.5. Personal air sampling and Cr analysis

Measurement of the inhalable fraction of dust was carried out in

accordance with FORCE Technology’s accreditation no. 51 from

DANAK, and following CEN/TS 15230 (CEN,

2005)

and EN 689.

Particles were collected on a PTFE filter (PTFE Membrane Disc Filters

- TF 1000, 1

m, 37 mm) using GSP-3.5 Conical Style Inhalable Dust

Samplers. Collection of inhalable dust was carried out with a sample

flow of 3.5 l/min. Pumps with automatic regulation for constant flow

were used for all measurements, and sample flow was adjusted and

calibrated to 3.50 L/min before each measurement.

The air measurements were carried out at each measurement site

using two identical parallel sampling systems, and the sampling inlets

were both placed in the worker breathing zone outside of any used PPE.

One of the two filters exposed in parallel was analyzed for total chro-

mium (Cr

tot

) and the other for Cr(VI). Blank filters were included for

each measurement site and analyzed together with the samples.

Measurements were carried out during normal work activities. The

collection time varied based on the duration of the relevant work

activities.

Analysis of Cr(VI).

Cr(VI) was extracted with a basic EDTA solution to

avoid reduction of Cr(VI) to Cr(III). Quantification was based on the

addition of the isotope-enriched 50 Cr(VI) using LC-ICP-MS technique.

Extraction and analysis was carried out by Eurofins according to USEPA

SW-846 METHOD 6800 (2007) (U.S,

2007),

and according to Eurofins’

DANAK accreditation no. 168. Eurofins reported the relative expanded

uncertainty of the analysis to be 20%.The LOD was 0.02

g/sample,

which with an airflow of 3.5 L/min and a minimum sampling time of 2 h,

results in a maximum LOD of 0.048

g/m

. Thus, the LOD of the air

sampling is well below 0.1

g/m

, 10% of the Danish OEL.

Analysis of Cr

Tot

. Extraction and analysis of total chromium followed

ISO15202. Chromium and chromium-containing compounds were

extracted with strong acid and analysis was carried out by Eurofins by

ICP-MS (ISO,

2004).

The LOD was 0.2

g/sample. Eurofins reported the

relative expanded uncertainty of the analysis to be 20%.

The calculation of mean, standard deviation etc. was done by

including all results below the limit of detection (LOD). This was done by

dividing the calculated LOD by 2, and using LOD/2 as the input result in

all calculated statistics.

2.6. Blood and urine sample collection

The biological sampling was performed on the same day as the

Total

Office work/non-metal

work

Others

Machining

Process operation

Work task on sampling day

Welding

Total number of participants

Controls

Within company controls: Manufacture/processing of metal

products

Within company controls: Bath plating

Within company controls: Vocational school (blacksmiths

welding and electricians)

Exposed

Manufacture/processing of metal products

Bath plating

Vocational school (stainless steel welding)

Categorisation

Total

EB: Participants with personal air exposure and biological sampling.

E: Participants only with personal air exposure.

B: Participants only with biological sampling.

A: Participants with any measurement (E or B or (EB).

Companies

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measurement of inhalable Cr

Tot

and Cr(VI) except for three participants

at one of the bath plating companies, where the air measurements were

performed approximately five months after the biological sampling for

logistical reasons. The blood and spot urine samples were collected on

Wednesdays and Thursdays between 10 a.m. and 2 p.m. (after the

workers had worked for at least 3–5 h). The participants collected a spot

urine sample in an acid-washed cup. Urine samples were transferred to

an acid-washed tube. The urine was kept at 4

◦

C on the sampling day,

and stored at 20

◦

C after transport to NFA. The blood was collected in

two vacutainer tubes: 1) one sodium-heparin for micronuclei, 2) one

sodium-heparin acid-washed tube for analysis of Cr content. The blood

was kept at 4

◦

C on the sampling day. In the afternoon on the sampling

day, after transport to NFA, the erythrocytes were isolated as previously

described (Jiang

et al., 2024).

In brief, the erythrocytes were separated

by centrifugation for 10 min at 1300 g and washed three times with

isotone saline. The washed erythrocytes were kept at 20

◦

C until the

tubes were used for Cr content analysis. On the day after sample

collection, the second sodium-heparin tubes were sent to FIOH for

MNRET analysis.

2.7. Measurement of creatinine and density in urine

Cr content in urine was adjusted for urinary dilution in two different

ways: 1) density of urine, and 2) creatinine. The density of the urine was

measured by weighing. Adjustment of the density was performed ac-

cording to (Jiang

et al., 2024)

using the following formula: C

(densi-

ty-adjusted)

(1-

mean

)/(1-

sample density

), where C

the determined Cr

concentration in the sample,

mean

the mean of the urinary density of

all participants, and

sample density

the density of the urine sample. The

urine density of three samples was below 1.00 g/mL suggesting that the

urine had been diluted with water. The samples were excluded from the

analysis of density-adjusted Cr content in urine. Creatinine was

measured as previously described (Hansen

et al., 2008).

The molecular

weight of creatinine (113.1 g/mol) was used.

2.8. Measurements of Cr in urine and red blood cells

Tubes and tips were washed with acid (5% HCl and 5% HNO

) to

remove metal background contamination. Analysis of Cr content in

urine and blood was performed essentially according to (Jiang

et al.,

2024).

The samples were stored at 4

◦

C prior to the analysis and

thawed at room temperature on the day of analysis. Sample volumes of

250

L (urine) or 100

L (blood) were diluted 20 times in disposable

polypropylene tubes (Sarstedt AG

Co. KG, Germany) with an alkaline

solution containing 0.5 g/L EDTA disodium salt dihydrate (Merck,

Darmstadt, Germany), 0.5 g/L Triton X-100 (Sigma-Aldrich, St Louis,

USA) and 5 g/L ammonia (25 %, suprapur, Merck KGaA, Darmstadt,

Germany). The samples were prepared in duplicates (R1 and R2). The

mean values were used in subsequent statistical analyses.

The total mass concentration of Cr in the samples was determined by

inductively coupled plasma-mass spectrometry (ICP-MS) using an iCAP

TQ ICP-MS (Thermo Fisher Scientific, Bremen, Germany) equipped with

an ASX-560 autosampler and a ASXpress PLUS valve equipped with the

1 mL sample loop (Teledyne CETAC Technologies, Omaha, NE, USA).

The analysis was performed in single quadrupole mode with helium as

collision gas in the collision cell (kinetic energy discrimination).

Instrumental configuration and parameters are listed in

Supplementary

Table 3.

For matrix matching of the calibration standards, whole blood

(SERONORM, SERO AS, Billingstad, Norway Whole blood L-1, Lot:

2011920, Cr: 0.61

g/L) and urine (The German External Quality

Assesment Scheme, Erlangen Germay) (G-EQUAS R64/2019 8 A, Cr:

0.25

0.09

g/L, were prepared in the same way as the samples and

spiked with Cr concentrations of 0.5, 1, 5 and 10

g/L.

Internal standard correction was performed by spiking 1

g/L

rhodium (Rh) to all samples and calibration standards. Calibration and

internal standard were prepared from standard solutions that contained

1000 mg/L of Cr or Rh (PlasmaCAL, SCP Science, Baie D’Urf

, QC,

Canada).

The analytical accuracy was verified towards certified reference

materials from G-EQUAS and Seronorm. The results (

g/L, mean

SD)

obtained for G-EQUAS (Lot. R64/2019 1 A) were for Cr in blood 2.0

0.1 vs. range 1.1–2.3 and for Seronorm (Lot, 2011920) in blood 0.70

0.12 vs. range 0.48–0.75. For G-EQUAS Cr in urine (Lot. R64/2019 2 A

and 8 A) the results obtained were 3.5

0.1 vs. range 2.8–4.0 and 0.22

0.02 vs. range 0.16–0.34 respectively. The relative standard deviation

based on triplicate analysis of the certified reference materials was 3 %

for urine and 3.4% for blood (G-EQUAS).

The limit of detection (LOD) and limit of quantification (LOQ),

calculated as 3- and 10-times the standard deviation of the blank sam-

ples, were 0.05 and 0.16

g/L, respectively, for the urine and 0.2 and

0.6

g/L, respectively, for the RBC.

2.9. Analysis of micronuclei frequency

Chromosomal damage was assessed by MNRET as previously

described (Andersen

et al., 2021; Tavares et al., 2022).

Briefly, 2 mL

whole blood samples collected with sodium-heparin were stored at 4

◦

and processed within seven days after collection. Immunomagnetic bead

separation was performed according to the instructions of the CELLec-

tion™ Pan Mouse IgG Kit (Invitrogen, Thermo Fisher Scientific, Wal-

tham, MA, USA) to isolate transferrin-positive (+CD71) reticulocytes

using a FITC Mouse Anti-human CD71 antibody (BD Biosciences, San

Jose, CA, USA). Samples were fixed in 2% paraformaldehyde in PBS with

g/mL of sodium dodecyl sulfate (SDS; Sigma-Aldrich, Merck KGaA,

Darmstadt, Germany) and stored refrigerated (4

◦

C) until analysis. Prior

to the analysis, each sample was divided into two replicates and DNA

was stained with Hoechst 33 342 (Invitrogen, Thermo Fisher Scientific,

Walthamt, MA, USA). The samples were analyzed with CytoFlex S flow

cytometer (Beckman Coulter, Brea, CA, USA) using blue (488 nm) laser

for the identification of

+CD71

reticulocytes and near UV (375 nm) laser

for the detection of DNA-containing micronuclei. CytExpert software

version 2.3 was used for data acquisition and analysis of 20 000–150 000

+CD71

reticulocytes per replicate sample. The micronuclei frequency

was quantified as per-mille (‰) of micronucleated reticulocytes from all

analyzed

CD71 reticulocytes.

2.10. Statistical analysis

Statistical testing was performed using GraphPad Prism 8.0.2. For

continuous variables, statistically significant differences between

exposed and controls were determined using the Kruskal–Wallis test

with Dunn’s post-hoc. Statistically significant differences between

exposed stratified by company and work task and controls were deter-

mined using Kruskal-Wallis test with Dunn’s post-hoc. For categorical

variables, the Chi-square test and Fisher’s exact test were used to

compare differences between exposed and controls. Correlations be-

tween variables were assessed by Spearman’s correlation.

3. Results

3.1. Characteristics of the study population

In total, the study involved 60 individuals including 36 classified as

Cr(VI) exposed and 24 classified as controls. The numbers of participants

with biological sampling and/or personal air exposure assessment across

different company types/vocational schools and work tasks are shown in

Table 1.

The volunteers from the exposed group were recruited from six

companies (three companies performing manufacture/processing of

metal products and three companies performing bath plating) and one

vocational school, where apprentices were taught to perform stainless

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steel welding. The exposure measurements and biological sampling

were performed between January 2022 and October 2023.

The volunteers from the control group (n

24) were recruited from

the same six companies and the same vocational school as the exposed

group. At the companies, office workers were recruited, and at the

vocational school, apprentices at other educations (electricians, black-

smiths welding) were recruited. Stationary exposure measurements at

relevant locations within the primary working area of the controls were

used as proxies for the personal air exposure for the control group.

The characteristics of the 49 participants (25 exposed and 24 con-

trols) with biological samples are presented in

Table 2.

Both types of controls (within company controls from the six com-

panies and vocational school controls) were regarded as within company

controls, as both are in occupational settings with possible bystander

exposures to Cr(VI). The two types of within company controls differed

from the group of Cr(VI) exposed for some lifestyle characteristics (re-

sults not shown), but much less, when the two types of controls were

merged into one group. For this reason, and to increase the statistical

power, the two types of controls were merged into one within company

control group.

3.2. Interviews, questionnaire and observations regarding OSH risk

prevention

In the interviews with OSH leaders from the six companies (three

bath plating, three manufacture/processing of metal products) and one

vocational school), all were highly aware of the OSH risks of working

with Cr(VI), and had either written or verbal procedures for handling

and working with Cr(VI). Some collaborated with external OSH experts

in carrying out yearly internal OSH meetings, establishing standard

operating procedures (SOPs), carrying out regular air measurements,

etc. Onboarding, instruction and training in Cr(VI) work tasks were most

often done through a learning-by-doing ‘buddy’ system

–

although most

workers had received some general OSH vocational training. Company

sizes (micro to large) and the degree of systematic OSH work varied

greatly, from a very informal verbal culture, to one company having

attained the ISO4500 international standard for their formal OSH

management system.

In regards to use of PPE and local exhaust ventilation systems,

questionnaire responses were received from 9 of the 11 bath plating

respondents, 12 of the 14 metal manufacturing respondents (welding

and grinding) and 9 of the 13 apprentices serving as within company

controls. For the bath plating workers (in both the previous week and

previous three months) the most often used form of respiratory

Table 2

Characteristics of the study group in Sam-Krom.

Cr(VI) Exposed n

Age, median (P5, P95)

Female, n (%)

Body Mass Index, median (P5,

P95)

Smoking, n (%)

Never smoker

Previous smoker

Current smoker

Coffee drinking, n (%)

Tea drinking, n (%)

Diet (mix, vegetarian, vegan),

n (%)

Supplement, n (%)

Implant, n (%)

Leisure activity with Cr, n (%)

protection were disposable respirators (single use P2/P3 masks), fol-

lowed by powered air purifying respirators (dependent, without fresh

air). Interviews with the OSH leaders and systematic observations of the

work processes revealed that all three bath plating companies used local

exhaust ventilation systems attached to the bath plating equipment, and

also used plastic balls and plastic sheets or cardboard coverings to

reduce aerosol generation for most of the baths. In the three welding and

grinding companies and the vocational school education entailing po-

tential Cr(VI) exposure, they all used local exhaust ventilation with

moveable capture hoods, and the workers/apprentices primarily used

welding helmets without further respiratory protection. However, two

of the workers, who did not fill out the questionnaire, were observed

using air-fed (independent) respirators. Safety observation indexes were

relatively high in both the three bath plating cases (94% average) and

four welding/grinding cases (86% average). The lower indexes in the

welding/grinding cases were partly due to incorrect use of the moveable

capture (ventilation) hoods. Appropriate gloves for the various purposes

(e.g. welding, bath plating) were worn by all workers, and coveralls/

aprons were worn by most workers. The latter were used over a period of

time (shifts) before being washed or replaced.

3.3. Cr(VI) in air

Concentrations of inhalable Cr(VI) measured in the exposed group

and stratified by company and work task are shown in

Table 3

and

Fig. 1.

The geometric mean of the air concentration of Cr (VI) was 0.26

g/m

for the workers with potential Cr(VI) exposure and 3.69

g/m

for the

apprentices with potential Cr(VI) exposure. When subdividing by com-

pany type, the exposure levels were 0.13

g/m

(95% CI: 0.04–0.41) for

companies manufacturing and processing of metal products, 0.81

g/m

(95% CI: 0.46–1.40) for bath plating companies and 3.69

g/m

(95%

CI: 1.47–9.25) at vocational schools during welding education.

For the workers with potential Cr(VI) exposure, 19 out of 24 (79%)

personal air measurements of inhalable Cr(VI) were below the Danish

OEL of 1

g/m

while 10 out of 24 (42%) measurements were below the

expected future Danish OEL of 0.25

g/m

(Fig.

2).

When subdividing by company type, two of fifteen air measurements

for companies in manufacturing and processing of metal products

exceeded the Danish OEL of 1

g/m

, as did 3/9 measurements for bath

plating companies and 6/8 measurements during welding exercises at

the vocational school. Five of fifteen air measurements for companies in

manufacturing and processing of metal products exceeded the expected

future Danish OEL of 0.25

g/m

, as did 9/9 measurements for bath

plating companies and 8/8 measurements during welding exercises at

the vocational school.

3.4. Cr in urine

Controls n

53 (18.5, 69.8)

4 (16.7)

28.0 (18.6,

58.4)

10 (42)

7 (29)

18 (75)

8 (33)

24/0/0 (100/0/

6 (25)

5 (21)

0 (0)

0.69

0.03

0.80

0.50

50 (26.3, 66.8)

0 (0)

27.0 (17.5, 46.0)

11 (44)

4 (16)

10 (40)

22 (88)

6 (24)

25/0/0 (100/0/0)

8 (32)

5 (20)

4 (16)

0.24

0.47

0.59

>0.99

0.04

Mann-Whitney test.

Chi-square test.

Fisher’s exact test.

No statistical testing, all participants are on a mixed diet.

Urine content of Cr was used as a biomarker of recent exposure to Cr

(VI), while bearing in mind that urinary Cr reflects exposure to Cr(III)

and Cr(VI). Urinary Cr content is presented in three different ways as Cr

content in urine, Cr content normalised to creatinine, and density

adjusted (Table

4).

Urinary Cr was increased in workers with potential

Cr (VI) exposure as compared to controls for all three measures of uri-

nary Cr (Table

and

Fig. 3).

Thus, workers with potential occupational

exposure to Cr(VI) had a median Cr of 1.52

g Cr/g crea (5th-95th

percentile 0.25–100) as compared to the controls with a median of 0.31

g Cr/g crea (0.14–4.1). Urinary Cr levels did not differ between within

company controls and vocational school controls (results not shown).

When sub-dividing by company or work tasks, urinary Cr was increased

for workers with potential Cr(VI) exposure at bath-plating companies

(median 3.12

g Cr/g crea (1.72–137.1) compared to controls and when

subdividing by work task, workers engaged in process operation had

significantly increased urinary Cr levels (median Cr level: 3.95

g Cr/g

crea (1.72–137.1)

g Cr/g crea) compared to controls.

The French BOEL of 2.5

g/L Cr was exceeded for 9 out of 11 workers

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A.T. Saber et al.

Table 3

Upper panel: concentrations of inhalable total chromium (Cr

Tot

;

g/m

) and hexavalent chromium (Cr(VI);

g/m

) measured in the exposed group (vocational school apprentices and workers) and within company controls

and stratified by company and work task. Lower panel: stationary measurements of CrTot and Cr(VI) at control locations and locations with potential Cr exposure at the companies.

AIR (PERSONAL)

Cr(VI)

n (n

LOD)

GM (95% CI)

Median (P5, P95)

Mean (95% CI)

Tot

n (n

LOD)

GM (CI)

Median (P5, P95)

Mean (95% CI)

AIR (STATIONARY)

Cr(VI)

n (n

LOD)

GM (95% CI)

Median (P5, P95)

Mean (95% CI)

Tot

n (n

LOD)

GM (CI)

Median (P5, P95)

Mean (95% CI)

Controls

Vocational school Apprentices exposed

8 (0)

3.69 (1.47; 9.25)

3.80 (0.80; 20.0)

5.98 (0.69; 11.26)

8 (0)

9.99 (5.09; 19.61)

11.85 (2.70; 31.40)

12.95 (4.95; 20.95)

Workers exposed

24 (5)

0.26 (0.12; 0.57)

0.42 (0.02; 21.74)

1.69 ( 0.70; 4.07)

24 (1)

4.49 (2.46; 8.16)

4.03 (0.28; 61.08)

10.76 (3.82; 17.69)

Cr companies

8 (1)

0.22 (0.05; 0.90)

0.30 (0.011; 1.28)

0.48 (0.09; 0.87)

8 (1)

1.73 (0.61; 4.89)

1.40 (0.25; 14.18)

3.39 ( 0.47; 7.26)

COMPANY - TYPE

Manufacture/processing

15 (5)

0.13 (0.04: 0.41)

0.08 (0.02; 27.96)

2.08 ( 1.89; 6.05)

15 (0)

6.86 (3.50; 7.03)

7.47 (1.60; 61.40)

14.14 (3.23; 25.05)

Manufacture/processing

3 (1)

0.05 (0.00; 4.56)

0.01; 0.36)

0.13 ( 0.35; 0.62)

3 (0)

3.03 (0.11; 83.85)

1.40 (1.40; 14.18)

5.66 ( 12.67; 23.99)

Bath plating

9 (0)

0.81 (0.46; 1.40)

0.71 (0.28; 3.10)

1.03 (0.36; 1.70)

9 (1)

2.21 (0.69; 7.10)

2.29 (0.25; 19.20)

5.11 (0.27; 9.95)

Bath plating

5 (0)

0.53 (0.18; 1.54)

0.82 (0.19; 1.28)

0.69 (0.11; 1.26)

5 (1)

1.23 (0.28; 5.47)

1.01 (0.25; 5.00)

2.03 ( 0.46; 4.53)

COMPANY - WORK TASK

Welding

9 (1)

0.24 (0.05; 1.28)

0.22 (0.02; 28.96)

3.33 ( 3.77; 10.43)

9 (6)

12.87 (5.74; 28.88)

11.20 (3.00; 61.40)

21.17 (3.44; 38.89)

Welding

Process operation

8 (0)

0.86 (0.47; 1.60)

0.79 (0.28; 3.10)

1.10 (0.35; 1.86)

8 (1)

1.67 (0.54; 5.31))

1.95 (0.25; 9.10))

3.35 (0.29; 6.41)

Process operation

Machining

6 (3)

0.09 (0.01; 0.53)

0.06 (0.02; 1.01)

0.27 ( 0.15; 0.69)

6 (0)

3.85 (1.24; 12.01)

2.55 (1.60; 19.20)

6.45 ( 1.17; 14.07)

Machining

Control location

11 (9)

0.017 (0.01; 0.03)

0.011 (0.01; 0.13)

0.03 (0.002; 0.05)

11 (7)

0.35 (0.20; 0.59)

0.39 (0.10; 1.60)

0.46 (0.18; 0.75)

Vocational school

Endpoints stratified by company type.

Endpoints stratified by company work task. Results from

“Others”

not shown for GDPR reasons (only one participant).

No personal air measurements performed among controls.

Stars indicates statistically significance following a Mann-Whitney test, stationary control measurements versus vocational school.

Stars indicates statistically significance following a Mann-Whitney test, stationary control measurements versus all company exposed.

Stars indicate statistically significance by Dunnet’s multiple comparisons test, stationary control measurements versus company types, as a post hoc analysis following a significant Kruskal-Wallis test.

Stars indicate statistically significance by Dunnet’s multiple comparisons test, stationary control measurements versus work tasks, as a post hoc analysis following a significant Kruskal-Wallis test.

Stationary air measurements performed at control locations (companies and vocational school).

No stationary measurements were performed in exposed locations on the vocational school.

Stars indicates statistically significance following a Mann-Whitney test, stationary control measurements versus all company exposed.

Dunnet’s multiple comparisons test, controls versus work tasks, as a post hoc analysis following a significant Kruskal-Wallis test.

No measurements.

Number of measurements (number of measurements below LOD.

0.05.

0.01.

0.001.

0.0001.

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Fig. 1. Inhalable hexavalent chromium (Cr(VI)) in exposed workers across company and work task and in exposed students at a vocational school

A. Inhalable hexavalent chromium (Cr(VI)) in exposed workers across company; B. Inhalable hexavalent chromium (Cr(VI)) in exposed workers across work task; C.

Inhalable hexavalent chromium (Cr(VI)) in exposed apprentices at a vocational school; OEL: Occupatinal exposure limit, Manu./Proces: Manufacture/Processing of

metal products, Process oper.: Process operation; Dunn’s post-hoc test, *P

0.05, **P

0.01. The data are presented as geometric mean and 95% CI.

3.6. Micronuclei

MNRET was used as a biomarker of recent genotoxic exposure. The

median MNRET frequency was 1.59 (5th-95th percentile: 0.78; 10.92) in

Cr(VI) exposed and 1.06 (0.72; 2.064) in within company controls

(Table

4).

When subdividing by company or work task (Fig.

4),

Cr (VI)-

exposed workers at bath plating companies had increased median levels

of MNRET (1.89 (0.78–12.92)) as compared to controls (1.06

(0.71–2.06) (p

0.004), while the level among Cr(VI) exposed workers

at manufacture/processing companies (1.31 (0.79–2.58) was not

increased compared to controls. Similarly, subdividing by work task

revealed that the median level of MNRET for the work task process

operation (1.89 (0.82–12.92)) was significantly increased compared to

controls. The medians for work tasks welding (1.49 (0.79–2.38)) and

machining (0.95 (0.78–6.25)) were not significantly increased

compared to controls.

3.7. Correlation between effect and exposure biomarkers

Correlations between exposure and effect biomarkers are presented

Fig. 5

and

Supplementary Fig. 1.

There were strong correlations be-

tween inhalable Cr(VI) and the urinary Cr (unadjusted: Spearman’s rank

correlation coefficients (r

)

0.75; creatinine adjusted: r

0.71 and

density adjusted: r

0.76)). Inhalable Cr(VI) correlated moderately

with the Cr content in RBC (r

0.52). There were also strong to very

strong correlations between RBC-Cr and all three ways of presenting

urinary Cr (unadjusted: r

0.77; creatinine adjusted: r

0.80 and

density adjusted: r

0.76). The effect biomarker for genotoxicity,

MNRET, correlated moderately to the creatinine adjusted urinary Cr

content (r

0.40) and moderately and negatively to Cr

Tot

in air (r

0.51).

4. Discussion

This is the first study on Cr(VI) exposure and toxicity in Denmark in

30 years and shows that although the Cr(VI) levels in air have decreased,

particularly bath platers still have an elevated Cr(VI) exposure. Overall,

Fig. 2.

Frequency distribution histogram for personal measurements of inhal-

able Cr(VI) at companies.

with potential Cr(VI) exposure at bath plating companies, for 2 out of 14

workers within manufacture and processing and for one out of 24

within-company control. The Finnish BOEL of ca. 10

g/L Cr was

exceeded for 3 out of 11 workers with potential Cr(VI) exposure at bath

plating companies, for none of the workers within manufacture and

processing and for one out of 24 within-company control.

3.5. Cr in red blood cells

Cr in RBC was used as a biomarker of the cumulative exposure to Cr

(VI) the last 4 months (Table

4).

Cr(VI) content in RBC was increased in

workers with potential Cr(VI) exposure (median level: 0.89

g/L (95%

CI: 0.54–4.9) as compared to controls (median: 0.60 (0.50–0.93) (P

0.0001). Levels of Cr(VI) in red blood cells did not differ between within

company controls and vocational school controls (results not shown).

When subdivided by company type and work task, workers with po-

tential Cr(VI) exposure at bath plating companies had significantly

increased blood levels of Cr(VI) (P

0.0001), as had process operators

0.0001) when subdivided by work task (Fig.

and

Table 4).

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Table 4

Chromium (Cr) concentration in urine and red blood cells (RBC) and micronucleus assay in reticulocytes (MNRET).

URINE

Unadjusted

Median (P5, P95)

Mean (95% CI)

Creatinine adjusted

Median (P5, P95)

Mean (95% CI)

Density adjusted

Median (P5, P95)

Mean (95% CI)

BLOOD

RBC-Cr

Median (P5, P95)

Mean (95% CI)

MNRET

Median (P5, P95)

Mean (95% CI)

Unit

Controls

n¼24

0.40 (0.05; 6.44)

0.78 (0.10; 1.46)

Workers exposed

d,e

n¼25

1.47 (0.2; 50.7)

5.66 (0.29; 11.0)

n¼25

1.52 (0.25; 100)

7.74 ( 3.46; 18.93)

n¼24

2.42 (0.28; 58.39)

6.98 (0.85; 13.10)

Workers exposed

d,e

n¼25

0.89 (0.54; 4.92)

1.41 (0.86; 1.96)

n¼25

1.59 (0.78; 10.92)

2.27 (1.22; 3.32)

COMPANY

Manufacture/processing

Bath plating

n¼11

4.68 (1.47; 64.6)

11.69 ( 059; 23.96)

n¼11

3.12 (1.72; 137.1)

16.6 ( 10.37; 43.56)

n¼11

4.61 (1.72; 69.5)

13.55 (0.23; 26.87)

Bath plating

n¼11

1.33 (0.95; 4.98)

2.28 (1.16; 3.41)

n¼11

1.89 (0.78; 12.92)

3.35 (0.96; 5.74)

n¼14

0.56 (0.14; 3.91)

0.94 (0.36; 1.52)

n¼14

0.50 (0.20; 2.0)

0.78 (0.48; 1.07)

n¼13

0.69 (0.28; 5.89)

1.41 (0.41; 2.42)

Manufacture/processing

n¼14

0.71 (0.52; 1.00)

0.72 (0.65; 0.80)

n¼14

1.31 (0.79; 2.58)

1.42 (1.07; 1.76)

WORK TASK

Welding

n¼8

0.55 (0.10; 2.30)

0.80 (0.22; 1.38)

n¼8

0.65 (0.44; 2.0)

0.87 (0.42; 1.32)

n¼7

1.15 (0.35; 5.89)

1.79 (0.03; 3.56)

Welding

n¼8

0.74 (0.52; 1.00)

0.74 (0.63; 0.85)

n¼8

1.49 (0.79; 2.38)

1.41 (0.98; 1.84)

Process operation

n¼9

4.70 (1.50; 64.6)

13.51 ( 1.78; 28.8)

n¼9

3.95 (1.72; 137.1)

19.7 ( 14.3; 53.7)

n¼9

8.51 (1.71; 69.5)

15.80 ( 0.69; 32.32)

Process operation

n¼9

1.33 (0.95; 4.98)

2.49 (1.11; 3.88)

n¼9

1.89 (0.82; 12.92)

3.32 (0.45; 6.18)

Machining

n¼7

0.90 (0.40; 5.0)

1.89 (0.19; 3.59)

n¼7

0.95 (0.38; 3.1)

1.3 (0.32; 2.23)

n¼7

0.69 (0.30; 4.21)

1.77 (0.20; 3.34)

Machining

n¼7

0.84 (0.59; 1.43)

0.89 (0.57; 1.20)

n¼7

0.95 (0.78; 6.25)

2.06 (0.22; 3.90)

g Cr/L

g Cr/g crea

n¼24

0.31 (0.14; 4.1)

0.62 (0.19; 1.04)

g Cr/L

¼22

0.40 (0.16; 21.3)

1.72 ( 0.57; 4.00)

Controls

n¼24

0.60 (0.50; 0.93)

0.63 (0.58; 0.68)

g Cr/L

‰

n¼24

1.06 (0.71; 2.06)

1.14 (0.99; 1.30)

P5: 5th percentile; P95: 95th percentile; 95% CI: 95% confidence interval.

Endpoints stratified by company type.

Endpoints stratified by company work task. Results from

“Others”

not shown for GDPR reasons (only one participant).

All controls (within company controls

vocational school controls).

All company exposed.

Stars indicates statistically significance following a Mann-Whitney test, controls versus all company exposed.

Manufacture/procession used as abbreviation of Manufacture/procession of metal products.

Stars indicate statistically significance by Dunnet’s multiple comparisons test, controls versus company types, as a post hoc analysis following a significant Kruskal-Wallis test.

Stars indicate statistically significance by Dunnet’s multiple comparisons test, controls verus work tasks, as a post hoc analysis following a significant Kruskal-Wallis test.

Three samples were taken out due to a negative density (2 from the control group and 1 from the exposed group).

Chromosomal damage assessed by the micronucleus assay in reticulocytes (MNRET).

0.05.

0.01.

0.001.

0.0001.

International Journal of Hygiene and Environmental Health xxx

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Fig. 4. Red blood cell Cr (RBC-Cr) and micronuclei in peripheral blood

reticulocytes (MNRET) in controls and exposed workers across company

and work task

A. RBC; B. MNRET; Manu./Proces: Manufacture/Processing of metal products,

Process oper.: Process operation; Dunn’s post-hoc test, ****P

0.0001. The

data are presented as median and interquartile range.

Fig. 3. Urinary Cr in controls and exposed workers across company and

work task

A. Urinary Cr (unadjusted); B. Urinary Cr (density adjusted); C. Urinary Cr

(creatinine adjusted); Manu./Proces: Manufacture/Processing of metal prod-

ucts, Process oper.: Process operation; Dunn’s post-hoc test, **P

0.01, ****P

0.0001. The data are presented as median and interquartile range.

the SAM-Krom biomonitoring study recruited 60 subjects including 36

with possible occupational exposure to Cr(VI) and 24 without occupa-

tional exposure to Cr(VI). The company types included were selected

based on a report reviewing the current knowledge on occupational

exposure to Cr(VI) in Denmark (Højriis

et al., 2020).

The report

concluded that manufacturing and processing was the company cate-

gory with the highest number of workers with possible Cr(VI) exposure,

while bath plating represented companies with possible high occupa-

tional exposure to Cr(VI) (Højriis

et al., 2020).

The report estimated that

there are 5–7 Danish bath plating companies with 10–49 workers with

possible occupational exposure to Cr(VI) and ca. 5.000–20.000 workers

in welding, thermal cutting and sanding in stainless steel who are

potentially occupationally exposed to Cr(VI) (Højriis

et al., 2020).

For

welding, we recruited companies doing MAG and MMA welding in

stainless steel as these are the primary types of welding entailing Cr(VI)

exposure (Højriis

et al., 2020).

We included three bath-plating com-

panies encompassing 11 participants with possible occupational expo-

sure to Cr(VI) and 6 within company controls. Thus, the SAM-Krom

study included a substantial fraction of the bath-plating companies and

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Fig. 5. Scatter plot and Spearman correlation analysis between inhalable Cr(VI), red blood cells Cr (RBC-Cr), urine Cr/density, urine Cr/Crea and

micronuclei in peripheral blood reticulocytes (MNRET) in the exposed group.

A. Urine Cr/density versus inhalable Cr(VI); B. RBC-Cr versus inhalable Cr(VI); C. RBC-Cr versus urine Cr/density; D. MNRET versus Urine Cr/Creatinine.

bath platers in Denmark. In

Supplementary Table 4,

results from

SAM-Krom are compared to the Swedish SafeCrom and the European

HBM4EU results.

4.1. Cr(VI) in air

Air concentrations of Cr(VI) were measured for 3.0 h on average,

thus reflecting 37.5% of an 8-h work day. Assuming that the air mea-

surements are representative for an 8-h working day, the air concen-

tration of Cr(VI) in the working areas of the within company controls

was low (0.017

g/m

, (95% CI: 0.01; 0.03) and most of the measure-

ments were below the limit of detection. For workers with possible

occupational exposure to Cr(VI), the geometric mean for Cr(VI) expo-

sure was significantly increased and more than 10 times higher; 0.26

, (95% CI:0.12; 0.57), while still below the Danish OEL. When sub-

diving by company type, bath plating companies had significantly

higher Cr(VI) concentrations in the air (geometric mean (GM): 0.81

(95% CI: 0.46; 1.40) as compared with within company offices and as

compared to air levels within manufacturing (GM: 0.13

g/m

(95% CI:

0.04: 0.41). We measured rather high Cr(VI) exposure levels (GM: 3.69

g/m

, (95% CI: 1.47; 9.25) for trainees performing MMA welding at a

vocational school, highlighting the need to increase the focus on OSH

risk preventive measures and training in using the preventive measures

in the education and training processes.

Knudsen et al. (1992)

performed a biomonitoring study of occupa-

tional exposure to Cr(VI) among 127 welders and 80 reference persons

in Denmark in 1987 (Knudsen

et al., 1992).

Knudsen et al. reported air

exposure levels to chromates as geometrical means of 0.9

g/m

(0.4

Cr(VI)/m

), 1.4

g/m

(0.6

g Cr(VI)/m

) and 1.5

g/m

(0.7

g Cr

(VI)/m

) for TIG, MMA

TIG, and metal inert gas (MIG) welders,

respectively, thus reporting 2–3 times higher Cr(VI) air concentrations

than in the current study (GM for welding: 0.24

g/m

(95% CI: 0.05;

1.28).

Bonde and Christensen (1991)

assessed Cr exposure in 60 welders

and 45 reference persons at Danish workplaces and reported median

values of time-weighted average exposure to Cr(VI) in air (Bonde

and

Christensen, 1991).

For TIG welders in stainless steel, Cr(VI) was 3

g/m

and for mild steel welding, 1

g/m

was reported. The corre-

sponding median Cr(VI) air concentration for welders in the current

study was 0.2

g/m

, (95% CI: 0.02; 28.96). Thus, the occupational

exposure levels to Cr(VI) for welders seem to be lower as compared with

the two more than 30-year-old Danish studies on welders. When

comparing to contemporary studies, the Swedish SafeChrom study, re-

ported similar levels as in the present study: GM for welders as 0.17

g/m

, (95% CI: 0.08–0.37) and the median 0.1

g/m

, (P5-95:

0.02–14.73) (Jiang

et al., 2024).

In the large HBM4EU study, GM for Cr

(VI) measured outside any respiratory protective equipment (RPE) was

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0.5

g/m

and median air concentration was also 0.5

g/m

In the current study, Cr(VI) exposure levels for the work task process

operation, representing bath plating was GM: 0.86

g/m

, (95% CI:

0.47; 1.60). In SafeChrom, process operation (representing bath plating)

had GM of 0.19

g/m

Cr(VI) (95% CI: 0.12–0.31), while HBM4EU re-

ported a GM of 0.3

g Cr(VI)/m

outside RPE. Thus, in comparison with

the Swedish and the pan-European study, the Danish Cr(VI) levels for

bath plating were relatively high, being 3–5 fold higher as compared to

the other contemporary European studies.

4.2. Exposure biomarkers

For all biomarkers of Cr(VI) exposure, bath platers had highly

increased levels compared to controls, whereas welders represented in

the category manufacturing and processing of metal products did not

differ from the controls for any of the biomarkers of exposure.

4.2.1. Urinary Cr

Kristiansen et al. (1997)

reported a reference interval for urinary Cr

in the general population in Denmark to be a mean

standard deviation

of 5.2 nmol/L

4.1 nmol/L corresponding to 0.27

g/L

0.21

g/L

(Kristiansen

et al., 1997).

Median values were 4.2 nmol/L corresponding

to 0.22

g/L with a 95% CI of 0.20–1.35

g/L. In the present study, the

within company controls had a mean urinary Cr of 0.78

g/L and a

median of 0.4

g/L Cr. These values are higher than the reference values

and higher than the outwith company controls in the Swedish Safe-

Chrom study, where median post-shift urinary Cr values of 0.11

g/L

were reported (Jiang

et al., 2024).

This probably reflects that the within

company controls in the current study were exposed to Cr to some

extent. This is in agreement with the findings from the large European

HBM4EU study (Santonen

et al., 2022; Viegas et al., 2022),

who re-

ported higher urinary Cr for within company controls than for outwith

company controls. HBM4EU reported urinary Cr for within company

controls with median value of 0.3

g Cr/g Crea, which is similar to the

median urinary Cr for within company controls in the present study of

0.31

g Cr/g Crea. In comparison, the HBM4EU outwith company

controls had median urinary Cr of 0.1

g Cr/g Crea, which is in line with

the SafeChrom study outwith controls (median urinary Cr: 0.10

g Cr/g

Crea). Taken together, this suggests that the within company controls in

the present study have increased urinary Cr as compared to Danish

reference values and as compared to outwith company controls from

other studies.

Occupational exposure to Cr(VI) via Cr in urine has been assessed in

the two previously mentioned Danish studies of welders. The reference

persons were metalworkers and electricians. In

Bonde and Christensen

(1991),

MMA/stainless steel welders had median urinary Cr of 0.635

Cr/g Crea, TIG/stainless steel welders a median urinary Cr of 0.95

Cr/g Crea and mild-steel welders a urinary Cr of 0.60

g Cr/g Crea

(Bonde

and Christensen, 1991).

Metalworkers and electricians had

median urinary Cr levels of 0.34 and 0.32

g Cr/g Crea, respectively

(Bonde

and Christensen, 1991).

In the study by

Knudsen et al. (1992),

the average post-shift mean Cr concentrations in urine from exposed

stainless steel welders varied from 2.38 to 7.85

mol/mol creatinine or

0.78–2.16

g Cr/g Crea. The reference group had a mean urinary Cr of

0.37

g Cr/g Crea. In comparison, within company controls in

the present study have very similar median urinary Cr (0.31

g Cr/g

Crea) as the reference persons in the 30-year-old studies by Bonde

(Bonde

and Christensen, 1991).

The mean urinary Cr for within com-

pany controls in the present (0.62

g Cr/g Crea) is higher than the mean

urinary Cr of 0.37

g Cr/g Crea in

Knudsen et al. (1992).

The welders in

the current study had a median urinary Cr level of 0.65

g Cr/g Crea,

which is similar to the urinary Cr levels reported by

Bonde (1991)

for

MMA and mild steel welders. The welders in the current study had a

mean urinary Cr level of 0.87

g Cr/g Crea, which is similar to the

urinary mean Cr for MIG welders but lower than the urinary mean Cr

values for MMA

TIG and TIG welders in Knudsen et al. The urinary Cr

levels in welders in the current study was similar to the levels reported

by HBM4EU (post-shift urinary Cr: median value 0.7

g Cr/g Crea and

mean value: 1.1

g Cr/g Crea). Conversely, urinary Cr for welders was

lower in the Swedish SafeChrom study (median urinary Cr: 0.41

g Cr/g

Crea). However, the type of welders in the present study and the

Swedish study differed as the Swedish study included TIG welding and

the values may therefore not be directly comparable. In the bath plating

companies, Cr(VI) exposed workers had median urinary Cr value (4.68

g/L) that exceeds the French BOELs of 2.5

g/L. Similarly, workers

with the work task ‘process operation’ (i.e. performing bath plating) had

a median urinary Cr value of 4.70

g/g Crea. In comparison, bath platers

in HBM4EU had median urinary Cr levels of 1.1

g Cr/g Crea (Viegas

et al., 2022),

and in SafeChrom, workers at bath plating companies had

urinary Cr levels of 0.56

g Cr/g Crea (Jiang

et al., 2024).

Thus, the

Danish bath plates have much higher urinary Cr levels as compared to

the Swedish and the European studies.

4.2.2. Cr in RBC

In the current study, workers with occupational exposure to Cr(VI)

had significantly higher Cr levels in RBCs as compared to within com-

pany controls. When subdividing by company type, only workers within

bath plating had significantly increased RBC levels of Cr (VI).

In the Swedish SafeChrom study, workers at bath-plating companies

had median RBC levels of Cr of 0.83

g/L. Bath platers in HBM4EU

(Ndaw

et al., 2022)

had higher RBC levels of Cr (median value 4.34

g/L) as compared to 1.33

g/L in the current study for both Cr(VI)

exposed workers at bath plating companies and workers with the work

task ‘process operation’.

4.3. Interviews, questionnaire and observations of OSH risk prevention

Triangulation regarding OSH risk prevention through the use of in-

terviews, a questionnaire and systematic safety observations revealed

consistent results in terms of use of PPE and technical assistive devices

such as local exhaust ventilation systems, and the use of plastic balls and

sheet or cardboard coverings to reduce aerosol formation in bath

plating. The relatively high OSH compliance rate reflected by the sys-

tematic safety observation safety indexes may be due to heightened

attention given to the various data collectors by the companies on the

day of data collection, as data were mainly collected on announced visits

by the research team on the same day for both biomonitoring, air

sampling, interviews, questionnaires and safety observations.

A recent systematic review on occupational exposure to Cr(VI) pro-

vides evidence of the use of OSH risk prevention measures such as

technical solutions, PPE, job-rotation and limiting lengths of shift-work

(Verdonck

et al., 2021).

A study in nine European countries (HBM4EU

project) concluded that use of PPE in bath plating and welding showed

lower urinary Cr (Viegas

et al., 2022).

In addition, results from the

recent SafeChrom study in Sweden with visual observations in 113 Cr

(VI) exposed workers showed lower levels of both compliance to use of

local exhaustion ventilation and proper use of respiratory protective

equipment (Jiang

et al., 2024).

The authors of the study from Sweden

concluded that local exhaustion ventilation had a greater preventive

effect than respiratory protective equipment, and that the results suggest

that the use of respiratory protective equipment is inadequate due to

factors such as other exposure routes, irregular use or premature

removal of respiratory protective equipment during exposure, as well as

lack of fit test presumably leading to leaking from the mask when worn

(Jiang

et al., 2024).

Qualitative and quantitative fit testing can

contribute to more effective use of respiratory protective equipment

(HSE,

2019),

however results of the current study reinforce the need to

focus on applying the upper levels of the hierarchy of controls with

eliminating or substituting Cr, and through the use of more effective

technical solutions (e.g. automation) in reducing occupational exposure

to Cr(VI), as supplements to the use of organizational measures (e.g. job

rotation), and worker related measures (e.g. instruction, training and

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use of personal protective equipment).

4.4. Cr(VI) exposure in welders

In the current study, welders were statistically significantly exposed

to Cr(VI) when assessed by air measurement but not by measurement of

the actual exposure in blood or urine. However, the reason for this may

primarily be the modest statistical power caused by the limited number

of welders (n

7–9) and the use of within company controls who have

low Cr(VI) exposure. Comparing our findings to the data from the 30-

year-old Danish studies of welders, may suggest that the Cr(VI) expo-

sure of welders may have decreased over time as Cr(VI) in air was lower

in the current study as compared to (Bonde

and Christensen, 1991;

Knudsen et al., 1992).

However, the urinary Cr levels for welders in the

current study were not substantially different from the reported levels in

older Danish studies, and quite similar to the levels reported in the

HBM4EU study. Notably, lower urinary Cr level were reported in the

Swedish SafeChrom study. Despite the lower urinary Cr levels, urinary

Cr was correlated positively with welding in linear regression analyses

in the SafeChrom study (Jiang

et al., 2024).

This, in turn, suggests that

for the Swedish welders, the increased urinary Cr levels were likely

caused by occupational exposure.

4.5. Cr(VI) exposure in bath platers

We consistently found indications that workers with possible Cr(VI)

exposure in path plating companies and workers with the work task

‘process operation’ (i.e. bath plating) had increased Cr(VI) exposure in

terms of Cr(VI) in air, urinary Cr and Cr levels in red blood cells. Urinary

Cr exceeded the French BOELs of 2.5

g/L, which corresponds to an OEL

of 1

g/m

for bath plating, even though only a few air measurements

exceeded the Danish OEL assuming that the air measurement are

representative for an 8-h working day. This may indirectly suggest that

other routes of exposure may contribute to Cr(VI) exposure in addition

to inhalation. Both the interviews, questionnaires and systematic safety

observations suggested high OSH compliance - at least on the day of

observation. However, the biomarkers of exposure reflecting recent

exposure (urinary Cr) or exposure during the last 4 months (Cr in RBC)

may point to variation in OSH compliance in general. Similar high Cr

exposure of bath platers was found in HBM4EU (Viegas

et al., 2022),

who suggested that oral exposure by hand to mouth contact may

contribute to Cr(VI) exposure. In addition, in the HBM4EU study, it was

shown that automated bath plating significantly reduced urinary Cr

(Viegas

et al., 2022).

4.6. Micronuclei

Genotoxicity was assessed in terms of MNRET in the current study.

We found that Cr(VI) exposed workers had increased levels of MNRET as

compared to within company controls. Specifically, workers in bath

plating companies and workers with the work task ‘process operation’

had increased levels of MNRET as compared to within company controls

(median micronuclei frequencies 1.89, 1.89 and 1.06, respectively.).

Taken together with the increased levels of biomarkers of Cr(VI) expo-

sure, this suggests that the increased MNRET levels may be attributed to

occupational exposure.

In the HBM4EU chromate study, several biomarkers of genotoxicity

were studied including MNRET. In general, levels of biomarkers of

genotoxicity were increased in Cr(VI) exposed workers as compared to

outwith company controls but not as compared to within company

controls (Tavares

et al., 2022).

Specifically, MNRET levels were

increased for all Cr(VI) exposed as compared to outwith company con-

trols, and MNRET for welders was increased as compared to both out-

with and within company controls. Of note, MNRET was assessed for

four countries (Belgium, Finland, Poland and Portugal) and outwith

company controls were only available for Portugal and Finland.

Knudsen et al. (1992)

also assessed biomarkers of genotoxicity and

reported increased levels of chromosomal aberrations in stainless steel

welders as compared to within company controls (Knudsen

et al., 1992).

Our results agree with those previous studies reporting chromosome

damage. A major genotoxic effect of Cr(VI) that contributes to carci-

nogenesis is the formation of DNA adducts, which can lead to DNA

damage (Alur

et al., 2024).

Intracellular reduction of Cr(VI) results in

the generation of Cr(III), which forms several types of Cr-DNA adducts.

If not properly repaired, Cr-DNA adducts may generate mutations and

DNA double-stranded breaks (Krawic

and Zhitkovich, 2023),

which can

give rise to micronuclei and chromosome aberrations.

Whereas increased levels of micronuclei in peripheral blood lym-

phocytes have shown to be predictive of cancer risk in prospective

studies (Bolognesi

et al., 2021; Bonassi et al., 2007; Dhillon et al., 2021),

analysis of MNRET is a recent tool in human biomonitoring studies

(Costa

et al., 2011)

and has not yet been assessed for predictivity of

cancer risk. Nevertheless, MNRET is a promising mutagenicity

biomarker, especially as indicator of potential leukemogenic agents

(Albertini

and Kaden, 2020).

4.7. Study strengths and limitations

This study on occupational exposure to Cr(VI) is to our knowledge

the first in Denmark in 30 years. We used the same study protocol and

questionnaires as the Swedish SafeChrom study and the large HBM4EU

study with some modifications. Cr exposure in air was quantified using

accredited methods that were aligned with the Swedish SafeChrom

study. Thus, air measurements were performed according to the US EPA

and Cr(VI) content in inhalable dust was quantified by LC-ICP-MS

technique according to US EPA. For Cr in urine and blood, the work

was based on the analysis protocol from

Jiang et al., 2024)

with an

adapted volume of urine sampled (250

L), a similar dilution factor and

the same alkaline solution for the dilution (Jiang

et al., 2024).

For ma-

trix matching of the calibration standards, SERNORM Whole blood L-1,

Lot: 2011920 and urine GEQUAS 64/2019 8 A were prepared in the

same way as the samples and spiked with Cr. Inductively coupled plasma

mass spectrometry (ICP-MS), as applied in this study, was one of the two

methods that was evaluated as appropriate for the determination of

chromium in urine and whole blood regarding occupational exposure

levels by the HBM4EU quality assurance program (Nübler

et al., 2022).

The LOQs in this study (0.16

g/L for urine and 0.60

g/L for blood)

were below or similar to the mean LOQs reported in the HBM4EU

inter-laboratory comparison (0.27–0.42

g/L for urine and 0.42–0.95

g/L for whole blood, depending on test round). The selected reference

materials in this study had concentrations close to the low-level control

materials used in the inter-laboratory comparison and satisfying preci-

sion and trueness were obtained. Thus, the current study should be

comparable to the SafeChrom and HBM4EU studies (Jiang

et al., 2024;

Santonen et al., 2022).

Furthermore, the present study describes NACE

and ISCO-codes for the study population, which adds to transparency,

and facilitates replication and pooling of the data.

The current study has a number of limitations. The most important

limitation is the limited number of participants mainly caused by Covid-

19 lock-downs. Challenges in the recruiting of companies and employees

were also highlighted by the HBM4EU study (Galea

et al., 2021).

Furthermore, we note that the companies who agreed to participate

likely represent companies who have special interest in occupational

health and safety, and thus, may not be representative for the entire

field, potentially leading to underestimation of the true exposure and

accompanying risks. In addition, we only included employees who were

able to read and write in Danish, again potentially introducing bias in

the representativeness of the study participants. Another limitation is

that Cr(VI) was only measured once for each study participant. We only

collected one urine sample per day instead of morning and end-of-shift

samples for logistic reasons. Both the HBM4EU study and SafeChrom

convincingly showed increased urinary Cr levels in Cr(VI)-exposed

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workers could be attributed to occupational exposure (Jiang

et al., 2024;

Santonen et al., 2022).

The Cr(VI)-exposed group were all male, whereas

there were 4 women among the controls. The study group also included

present smokers, even though smoking is suggested to contribute to Cr in

blood and urine as well as MNRET (Offer

et al., 2005).

Of note, smoking

status did not influence urinary Cr in a study of reference levels in Danes

(Kristiansen

et al., 1997).

Due to the limited statistical power, we did not

adjust the analyses for age, sex and smoking status. Of note, these ad-

justments did not influence the results in the SafeChrom study (Jiang

et al., 2024).

As the air measurements for most of the participants were

done on the same day as the biological sample collection they may differ

from the exposure levels present 3 days prior to day of measurement,

which is the time when the RETMN were formed. Furthermore, three of

the air measurements at one of the bath plating companies were per-

formed five months after the biological sampling. However, the

day-to-day variation in the exposure from the baths is assumed to be

small, as the main source of Cr(VI) exposure is the baths. Another lim-

itation is that the Cr(VI) air exposure levels were only measured for 3 h

thus not reflecting an 8 h working day. If the worker change work task

during the day this may influence the average daily exposure level.

5. Conclusion

The SAM-Krom study shows that bath platers are highly exposed to

Cr(VI) as suggested by relatively high urinary Cr levels, Cr levels in RBC

and increased levels of micronuclei. The urinary Cr levels were high as

compared to the BOEL corresponding to 1

g/m

, thus indirectly sug-

gesting that additional exposure routes contribute to the exposure. For

welders, no statistically significant increases as compared to within

company controls were observed, however, the observed urinary Cr

levels were similar to the levels observed in HBM4EU and higher than

the levels observed for welders in SafeChrom. On the announced visits, a

consistent high degree of compliance to OSH risk prevention measures

was seen in bath plating for both technical devices (ventilation, plastic

balls, coverings) and use of personal protective equipment, and to a

lesser degree of compliance to OSH risk prevention measures in welding

with stainless steel. However, the biomarkers of exposure reflecting

recent exposure (urinary Cr) or exposure during the last 4 months (Cr in

RBC) may point to variation in OSH compliance in general. This, in turn,

may imply that a more consistent focus on OSH risk prevention mea-

sures, according to the upper levels of the hierarchy of controls (e.g.

substituting Cr, automation), would reduce occupational exposure to Cr

(VI).

CRediT authorship contribution statement

Anne Thoustrup Saber:

Writing

–

review

editing, Writing

–

original draft, Project administration, Methodology, Investigation,

Funding acquisition, Formal analysis, Data curation, Conceptualization.

Marcus Levin:

Writing

–

review

editing, Investigation, Formal anal-

ysis, Data curation.

Pete Kines:

Writing

–

review

editing, Methodol-

ogy, Investigation, Funding acquisition, Formal analysis, Data curation,

Conceptualization.

Kukka Aimonen:

Writing

–

review

editing, Re-

sources, Investigation.

Lucas Givelet:

Writing

–

review

editing,

Investigation, Formal analysis, Data curation.

Christina Andersen:

Writing

–

review

editing, Investigation, Formal analysis, Data cura-

tion.

Anja Julie Huusom:

Writing

–

review

editing, Investigation.

Tanja Carøe:

Writing

–

review

editing, Investigation.

Niels Erik

Ebbehøj:

Writing

–

review

editing, Methodology, Conceptualization.

Frans Møller Christensen:

Writing

–

review

editing, Project admin-

istration, Methodology, Investigation, Funding acquisition, Conceptu-

alization.

Zheshun Jiang:

Writing

–

review

editing.

Thomas Lundh:

Writing

–

review

editing.

Håkan Tinnerberg:

Writing

–

review

editing.

Maria Albin:

Writing

–

review

editing.

Malin Engfeldt:

Writing

–

review

editing.

Karin Broberg:

Writing

–

review

editing,

Funding acquisition.

Julia Catalan:

Writing

–

review

editing,

Methodology, Funding acquisition.

Katrin Loeschner:

Writing

–

review

editing, Methodology, Investigation, Formal analysis, Data curation.

Karsten Fuglsang:

Writing

–

review

editing, Methodology, Investi-

gation, Funding acquisition, Conceptualization.

Ulla Vogel:

Writing

–

review

editing, Writing

–

original draft, Validation, Supervision, Re-

sources, Methodology, Funding acquisition, Conceptualization.

Acknowledgements

The technical assistance from Ismo Koponen, Anne Abildtrup, Noor

Irman and Ulla Tegner is gratefully acknowledged. A special thanks goes

to the participating companies and vocational school. We are also

grateful to the study participants for their time and willingness put into

this study. We established a reference group which includes stake-

holders from e.g. the social partners, trade unions and The Danish

Working Environment Authority. We thank the reference group for their

support and fruitful discussions. We also wish to thank the SafeChrom

team in Sweden for fruitful discussions and knowledge sharing.

This work was supported by the Danish Working Environment

Research Fund (SAM-Krom, grant numbers 2019 5100 337, 2020 5100

706), Forskningsrådet for halsa, arbeitsliv och v

rd (Forte, grant

a a

number 202000208) and Afa Forsakring (grant number 200279).

Appendix A. Supplementary data

Supplementary data to this article can be found online at

https://doi.

org/10.1016/j.ijheh.2024.114444.

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