Boric acid

Epidemiological data

Administrative data

Endpoint:

epidemiological data

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2008-2017

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference

Reference Type:

publication

Title:

Boron-exposed male workers in Turkey: no change in sperm Y:X chromosome ratio and in offspring’s sex ratio

Author:

Duydu, Y., Basaran, N., Özgür Yalcin, C., Üstündag, A., Aydin, S., Gül Anlar, H., Bacanli, M., Aydos, K., Somer Atabekoglu, C., Golka, K., Ickstadt, K., Schwerdtle, T., Werner, M., Bolt, H.M.

Year:

2019

Bibliographic source:

Arch Toxicol. 93(3):743-751

Materials and methods

Study type:

cohort study (retrospective)

Endpoint addressed:

basic toxicokinetics

toxicity to reproduction / fertility

Test guideline

Qualifier:

no guideline required

Principles of method if other than guideline:

304 male workers in Bandirma and Bigadic (Turkey) with different degrees of occupational and environmental exposure to boron were investigated. Boron was quantified in blood, urine and semen. Investigation addressed the association between boron exposure and the Y:X sperm ratio in semen samples of male workers, including extreme occupational and environmental boron exposure conditions and the sex ratio at birth.

GLP compliance:

no

Test material

Test material information

Constituent 1

Reference substance name:

Boron

Cas Number:

7440-42-8

Molecular formula:

B

IUPAC Name:

Boron

Method

Type of population:

occupational

other: and non-occupational exposure

Ethical approval:

confirmed and informed consent free of coercion received

Remarks:

Sampling was approved by the Ethics Committees of the Hacettepe University School of Medicine, Ankara and Ankara University School of Medicine

Details on study design:

HYPOTHESIS TESTED (if cohort or case control study): Clarification of the reported boron-associated effect on the Y:X sperm ratio in semen samples of male workers

METHOD OF DATA COLLECTION
- Type: Questionnaire, other: sampling
- Details: Demographic information, such as age, the number of fathered children, duration of employment, pesticide application, smoking habits and alcohol consumption had been gathered within the former “Boron Projects I and II”. Peripheral blood, urine and semen were obtained after completing the questionnaire survey. Semen was sampled in accordance with WHO criteria (All participants gave their informed consent prior to participation).

STUDY PERIOD: 2008-2017 The semen samples for this study were sampled within the previous projects “Boron Project I” (2008–2010) and the “Boron Project II” (2014–2017)

SETTING: Bandirma and Bigadic (Turkey)

STUDY POPULATION
- Total population (Total no. of persons in cohort from which the subjects were drawn): (2010) Bandirma, one of the districts of Balikesir, is located at the south coast of the Marmara Sea. Semen was sampled from workers employed in the “Bandirma Boric Acid Production Plant” and from neighboring facilities (n=204). (2015) Bigadic, also a district of Balikesir, is located about 130 km south of Bandirma, at an altitude of 165 m above sea level. This area has the largest of boron deposits in Turkey. The deposits are specifically located around the small villages of Osmanca and Iskele. Because of the geology of the surroundings, boron concentrations in the drinking water of Iskele were much higher (12.2 mg B/L) than limits recommended in drinking water guidelines of the EU (1 mg B/L) and WHO (2.4 mg B/L). The “Bigadic Boron Works” are located in the village Osmanca, being the largest employer in the region. Within this facility colemanite and ulexite ores are mined in 4 open pits. Semen and blood samples of 76 workers could be analyzed within the “Boron Project II”. The study persons worked in the “Bigadic Boron Works” and resided in Iskele. The workers were both environmentally (by drinking water) and occupationally exposed to boron. (2016) Due to the local conditions boron exposure in Bandirma was possible only for the workers employed in the boric acid production and packaging units located at the “Bandirma Boric Acid Production Plant”. Environmental boron exposure by drinking water was negligible for the workers residing in Bandirma, due to very low boron concentrations in the municipal water. Semen and blood samples of 65 workers were used as low-exposed controls within the “Boron Project II”. These workers worked in different industrial facilities of the Bandirma production zone, but not in the boric acid production and packaging units.
- Selection criteria: occupationally and not occupationally exposed
- Total number of subjects participating in study: 345
- Sex/age/race: male/ age: 26-48 control group, 23-49 low exposure group, 27-48 medium exposure group, 22-53 high exposure group, 23-50 extreme exposure group/ no data for race
- Total number of subjects at end of study: 304 (n=163 from 2008-2010, n=141 from 2014-2017)
- Other: this study could be based on the assumption that individuals were chronically exposed to their area-specific level of boron. Based on this assumption the workers were classified into “control”, “low”, “medium”, “high” and “extreme” exposure groups, along with their measured blood boron concentrations: Workers with blood boron concentrations lower than 50 and lower than 100 ng B/g blood were assigned to the “control group” (n = 38) and “low exposure group” (n = 60), respectively. Fifty workers with blood boron between 100 and lower than 150 ng B/g blood were classified as “medium exposure group”. The lower limits of blood boron concentrations for the “high exposure group” (n = 87) and the “extreme exposure group” (n = 69) were 150 and 400 ng B/g blood, respectively

COMPARISON POPULATION
- Type: blood boron concentration below 50 ng B/g blood

OTHER DESCRIPTIVE INFORMATION ABOUT STUDY:

Details on exposure:

TYPE OF EXPOSURE: occupational and non-occupational

TYPE OF EXPOSURE MEASUREMENT: Area air sampling / Personal sampling / Exposure pads / Biomonitoring (urine) / Biomonitoring blood / other: Daily boron exposure (DBE) of the participating workers was determined by summing the boron intakes in daily consumed food, drinking water and inhaled dust. The DBE via food and drinking water was determined by taking water and food samples (double plate method) from their workplace
and home. The daily drinking water consumption of participating workers was assumed to be 2 L/day. Dust/air samples were sampled by using IOM samplers and personal air sampling pumps (SKC, AirCheck 2000).

EXPOSURE LEVELS: Daily boron exposure (DBE) of the participating workers was determined by summing the boron intakes in daily consumed food, drinking water and inhaled dust; Low, medium, high and extreme exposure groups were classified by measured blood boron concentration

EXPOSURE PERIOD: continuously

DESCRIPTION / DELINEATION OF EXPOSURE GROUPS / CATEGORIES:Major differences between historical and current boron concentrations in drinking water sources of the sampling areas were not apparent. Therefore,
this study could be based on the assumption that individuals were chronically exposed to their area-specific level of boron. Based on this assumption the workers were classified into “control”, “low”, “medium”, “high” and “extreme” exposure groups, along with their measured blood boron concentrations: Workers with blood boron concentrations lower than 50 and lower than 100 ng B/g blood were assigned to the “control group” (n = 38) and “low exposure group” (n = 60), respectively. Fifty workers with blood boron between 100 and lower than 150 ng B/g blood were classified as “medium exposure group”. The lower limits of blood boron concentrations for the “high exposure group” (n = 87) and the “extreme exposure group” (n = 69) were 150 and 400 ng B/g blood, respectively.

Statistical methods:

Box plots, Pearson’s correlation coefficient and linear regression display the empirical distribution and possible linear dependencies. In order to investigate the global hypothesis that the means of the five groups are equal, a Kruskal–Wallis test was used for all variables in Table 1. If the resulting p value was significant (p < 0.05), i.e., if the hypothesis of equal group means was rejected, the two-sided Wilcoxon–Mann–Whitney test as a post hoc test was applied to find distinct differences between each of the pairs of groups. The corresponding p values were adjusted with the Bonfferoni–Holm method for each variable in order to account for the problem of multiple testing. These nonparametric tests were deliberately employed as some of the variables showed outliers. All statistical tests were performed with R, version 3.4. (R Core Team 2017). The local as well as multiple significance levels of the tests were set at 0.05. For theoretical details of the nonparametric Kruskal–Wallis as well as Wilcoxon–Mann–Whitney test.

Results and discussion

Results:

The study population (n = 304) consisted of the control (n = 38), low exposure (n = 60), medium exposure (n = 50), high exposure (n = 87) and extreme exposure (n = 69) groups. The mean DBE, semen boron and blood boron concentrations of the “extreme” exposure group were 44.91 ± 18.32 mg B/day, 1643.23 ± 965.44 ng B/g semen and 553.83 ± 149.52 ng B/g blood, respectively.A signifiant association (p < 0.05) between blood and urine boron confirms both parameters as reliable boron exposure biomarkers. By contrast, a lack of association between blood boron and semen boron concentrations supports the earlier findings that boron concentrations in semen is not a reliable biomarker of boron exposure. When we compared the mean Y:X sperm ratio and the percentage of Y-bearing sperm in ejaculates of workers, statistically significant differences were not seen (p > 0.05). With all related data, the correlations between blood/semen boron and Y:X sperm ratio were statistically not significant (p > 0.05), which proves an absence of a dose-dependent decrease in Y:X sperm ratios.

Confounding factors:

A linear regression analysis of the pooled data showed no statistically significant associations between Y:X sperm ratio and demographic parameters (age, duration of employment), exposure biomarkers (boron concentrations in blood/urine/ semen), DBE, sperm concentrations, cofounders (alcohol consumption and smoking). However, there was an association between reported pesticide application and Y:X sperm ratio, which reached a statistically significant level (p = 0.02)

Any other information on results incl. tables

Table 1: Exposure biomarkers, Y:X sperm ratio and other related parameters of male workers

Parameters	Control (C),n= 38 < 50 ng B/g blood	Low (L) exposure,n=60 50 to <100 ng B/g blood	Medium (M) exposure,n= 50 100 to <150 ng B/g blood	(H) High exposure, n=87 150 to <400 ng B/g blood	(E) Extreme exposure,n= 69 > 400 ng B/g blood	p value
Age	42.89 ± 5.32 (26-48)	41.50 ± 6.05 (23-49)	40.22 ± 6.09 (27-48)	37.26 ± 7.46 (22-53)	36.61 ± 6.68 (23-50)	<0.05^a
Duration of employment, year	18^0 ± 6A9 (2-26)	15.79 ± 7.47 (0.17-23)	15.74 ± 7.51 (1-25)	9U5 ± 6A2 (0,5-23)	6.65 ± 4.84 (1-26)	<0.05^b
Blood B, ng B/g blood	30.00 ± 10.12 (16.23-49.23)	76.00 ± 15.22 (50.17-99.91)	122.88 ± 15.34 (101.28-149.84)	247.37 ± 71.32 (150.99-391.92)	553.83 ± 149.52 (401.62-1099.93)	<0.05^c
Semen B, ng B/g semen	1077.11 ± 1845.34 (52-8597)	1598.46 ± 2027.85 (111-8615)	1526.93 ± 1265.36 (189-4897)	1259.65 ± 1446.11 (100-10,542)	1643.23 ± 965.44 (188-8086)	<0.05^d
Urine B, mg B/g creat	6.8 ± 1.32 (0.785.56)	4.97 ± 2.28 (1.09-13.54)	6.35 ± 2.48 (1.79-12.67)	7.33 ± 4.72 (1.13-32.68)	14.38 ± 7.76 (1.06-29.79)	<0.05^e
DBE, mg B/day	4.57 ± 1.69 (0.20-7.54)	8.32 ± 5.71 (2.56-35.61)	14.81 ± 9.99 (2.56-47.18)	23.50 ± 13.94 (3.32-55.10)	44.91 ± 18.32 (7.95-106.79)	<0.05^c
Sperm conc., M/mL	73.04 ± 41.47 (2.00-166.30)	69.22 ± 47.93 (5.20-224.20)	69.02 ± 56.52 (5.90-277.20)	82.97 ± 60.10 (2.00-292.30)	88.05 ± 59.73 (14.70-259.00)	>0.05
Y:X sperm ratio	0.98 ± 0.03 (0.85-1.02)	0.99 ± 0.02 (0.89-1.04)	0.99 ± 0.02 (0.94-1.09)	0.99 ± 0.02 (0.86-1.03)	0.99 ± 0.02 (0.95-1.06)	>0.05
Y-bearing sperm, %	49.57 ± 0.88 (46.04-50.48)	49.74 ± 0.62 (47.17-50.93)	49.65 ± 0.61 (48.37-52.26)	49.71 ± 0.6 (46.2350.71)	49.74 ± 0.56 (48.72-51.48)	>0.05 >0.05
Children; mean ± SD	1.91 ± 0.45	1.88 ± 0.56	1.79 ± 0.62	1.82 ± 0.70	1.70 ± 0.61
Total	67	111	85	151	112
Boys; mean ± SD	0.95 ± 0.80	0.85 ± 0.68	0.90 ± 0.71	0.97 ± 0.84	0.87 ± 0.66	>0.05
Total and %	36; 53.73	51; 45.95	45; 52.94	84; 55.63	60; 53.57
Girls; mean ± SD	0.82 ± 0.80	± 0.76	0.80 ± 0.73	0.77 ± 0.90	0.75 ± 0.65	>0.05
Total and %	31; 46.27	60; 54.05	40; 47.06	67; 44.37	52; 46.43
Sex ratio at birth, boys/girls	1.16	0.85	1.13	1.25	1.15	>0.05

Mean ± SD (min–max), DBE daily boron exposure, M million. Kruskal–Wallis test for global comparison. Wilcoxon–Mann–Whitney test with

Bonferroni–Holm correction as post hoc test

^a C–H, C–E, L–H, L–E, M–E

^b C–H, C–E, L–H, L–E, M–H, M–E, H–E

^c All pairwise

^d C–M, C–H, C–E, L–E, H–E

^e C–L, C–M, C–H, C–E, L–M, L–H, L–E, M–E, H–E

Table 2: Linear regression analysis for the Y:X sperm ratio

	Total study group
	Estimate	Std. error	t	p value
Intercept,(β0)	9.91e-01	1.278e-02	77.59	<2e-16
Age, (β1)	-1.55e-04	3.59e-04	- 0.431	0.67
Duration of employment, (β2)	1.11e-04	3.42e-04	0.326	0.745
Blood boron conc., (β3)	1.53e-06	1.95e-05	0.08	0.94
Urine boron conc., (β4)	-5.65e-04	4.15e-04	- 1.36	0.18
Semen boron conc., (β5)	3.63e-07	1.10e-06	0.33	0.74
DBE (β6)	1.73e-04	1.67e-04	1.04	0.30
Sperm conc., (β7)	5.46e-05	3.31e-05	1.65	0.10
Pesticide application (β8)	-1.03e-02	4.25e-03	- 2.436	0.02*
Alcohol consumption (β9)	4.13e-03	3.83e-03	1.08	0.28
Smoking (β10)	- 2.10e-03	3.68e-03	- 0.57	0.56

Applicant's summary and conclusion

Conclusions:

In essence, a statistically significant association between high levels of boron exposure and a lower sperm Y:X ratio (lower percentage of male offspring), as suggested by Robins et al. (2008), is not in agreement with the results of the present study, which clearly support the lack of boron-mediated effects on the proportion of Y- to X-bearing sperm and on the sex ratio at birth after paternal exposure.

Executive summary:

The present study was conducted to investigates the association between boron exposure and the Y:X sperm ratio in semen samples of male workers, including extreme occupational and environmental boron exposure conditions and the sex ratio at birth. 304 male workers in Bandirma and Bigadic (Turkey) with different degrees of occupational and environmental exposure to boron were investigated. Boron was quantified in blood, urine and semen, and the persons were allocated to exposure groups along B blood levels. In the highest (“extreme”) exposure group (n = 69), calculated mean daily boron exposures, semen boron and blood boron concentrations were 44.91 ± 18.32 mg B/ day, 1643.23 ± 965.44 ng B/g semen and 553.83 ± 149.52 ng B/g blood, respectively. Overall, an association between boron exposure and Y:X sperm ratios in semen was not statistically significant (p > 0.05). Also, the mean Y:X sperm ratios in semen samples of workers allocated to the different exposure groups were statistically not different in pairwise comparisons (p > 0.05). Additionally, a boron-associated shift in sex ratio at birth towards female offspring was not visible. In essence, the present results do not support an association between boron exposure and decreased Y:X sperm ratio in males, even under extreme boron exposure conditions