Progestin Pollution in Surface Waters of a Major Southwestern European Estuary: The Douro River Estuary (Iberian Peninsula)

Abstract

The concentrations and spreading of eight synthetic and two natural progestins (PGs) were investigated in surface waters from ten sites at the Douro River Estuary. Samples were filtrated and subjected to solid-phase extraction (SPE) to isolate and concentrate the target PGs. The extracts were cleaned by silica cartridges and analyzed by LC-MS/MS. The finding of biologically relevant amounts of gonanes (22.3 ± 2.7 ng/L), progesterone derivatives (12.2 ± 0.5 ng/L), drospirenone (4.1 ± 0.8 ng/L), and natural PGs (9.4 ± 0.9 ng/L) support the possibility of these compounds acting as endocrine disruptors. Despite the absence of significant differences amongst sampling sites and seasons, the principal component analysis (PCA) and the linear discriminant analysis (LDA) approaches reveal that spring and summer have different patterns of PG distribution compared to autumn and winter. The assessment of risk coefficients (RQs) and the potential concentrations of synthetic progestins in fish blood sustains that all tested compounds pose a significant risk to local biota (RQs > 1). Additionally, three progestins-norethindrone, norethindrone acetate, and medroxyprogesterone acetate-should reach human-equivalent therapeutic levels in fish plasma. Overall, the current data show PGs' presence and potential impacts in one of the most important estuaries of the Iberian Peninsula.

Keywords: aquatic toxicology; gonanes; progesterone derivatives; progestogen; risk quotients (RQs); spirolactone derivatives.

Figure 1

Location of the sampling sites along both margins of the Douro River Estuary (S1 to S10), Portugal. Salinity levels in the studied areas varied from 1.8 ± 2.2 PSU in the inner estuary to 10.0 ± 4.1 PSU near the coastline. This image also points out anthropogenic sources that could contribute to increased concentrations of PGs (adapted from Google KML Map).

Figure 2

Concentrations (ng/L) by target PGs (A) and seasons (B). Data are expressed in boxplots with the minimum, median, maximum, average (+), and interquartile ranges Q1–Q3. In (A) dots indicate the average values recorded for each PG by season (n = 4), whereas those in (B) show the average levels of all PGs by season (n = 10). Significant differences in annual PG concentrations are indicated by different letters (A) (ANOVA, p < 0.05). No significant differences were detected among sampling sites or across seasons (B) (ANOVA, p > 0.05).

Figure 3

Levels (ng/L) of (A) gonanes, (B) progesterone derivatives, (C) drospirenone, and (D) natural PGs. Results are expressed in boxplots with the minimum, median, maximum, average (+), and interquartile range Q1–Q3. Dots represent individual values measured on different sampling occasions at each sampling site (n = 80). No significant differences were observed amongst PGs of the same group (ANOVA, p > 0.05).

Figure 4

Global amounts of PGs in each Douro River Estuary sampling site. Results are expressed in boxplots with the minimum, median, maximum, average (+), and interquartile range Q1–Q3. Dots indicate the average individual values measured on different sampling occasions (n = 80). No significant differences were found among sampling sites (ANOVA, p > 0.05).

Figure 5

(A) Principal component analysis (PCA) score plots of PC1 vs PC2 show the distribution of individual PGs by sampling occasion (n = 4) and sites (S1 to S10, n = 10). (B) Linear discriminant analysis (LDA) score plots of Axis 1 vs。 Axis 2 illustrate the similarity between individual PGs by sampling season (n = 4) and sampling sites (S1 to S10, n = 10).