High-resolution temporal monitoring of rare earth elements in acidic drainages from an abandoned sulphide mine (iberian pyrite belt, Spain).

Rare earth elements (REE) are strategic elements due to their economic importance. However, the studies dedicated to the distribution and behaviour of REE in aquatic systems have been scarce until a few decades ago. This work studies the seasonal variations of REE concentrations in acid mine drainage (AMD) affected water courses and the factors controlling their mobility under different hydrological conditions. To address this issue, a high-resolution sampling was performed for two years in selected sampling sites. REE concentrations were very high (median values of 2.7-3.4 mg/L, maximum of 7.0 mg/L). These values are several orders of magnitude higher than those found in natural waters, highlighting the importance of AMD processes on the release of REE to the hydrosphere. No good correlations were found between pH and REE concentration, while REE correlated positively (r Spearman coefficient of 0.78-0.94) with EC and negatively (r -0.88 to -0.90) with discharge in AMD-affected streams. A conservative behaviour of REE was observed due to the strongly acidic conditions observed in the study area. The waters also showed an enrichment in MREEs over LREEs and HREEs (mean values of GdN/LaN>1.8 and YbN/GdN < 0.7), typical of AMD waters. An asymmetry in the content of LREE and HREE was observed in AMD samples studied, which could be explained by the preferential dissolution of LREE or HREE-enriched minerals within each waste heaps. Multivariate analysis suggests the influence of Mn-rich minerals existent in the study area as a potential source of LREE.

Application of high-resolution techniques in the assessment of the mobility of Cr, Mo, and W at the sediment-water interface of Nansi Lake, China.

There are few studies on the simultaneous behavior of chromium (Cr), molybdenum (Mo), and tungsten (W) belonging to group VIB of the periodic table. Herein, based on high-resolution dialysis (HR-Peeper) and diffusive gradients in thin films (DGT) technology, the vertical distribution characteristics of DGT-labile and soluble Cr, Mo, and W in two lakes of Nansi Lake (Weishan Lake and Dushan Lake) were analyzed. In addition, the net diffusion fluxes and R-value (CDGT/Csol) were used to evaluate the mobility and release risk of metals at the sediment-water interface. The results showed that the DGT-labile concentrations of the three metal elements (Cr, Mo, and W) in Weishan Lake were higher than those in Dushan Lake, both in overlying water and sediment. This is mainly due to the dredging of the Dushan Lake area, which can permanently remove the polluted sediment in the lake. Meanwhile, the exogenous input is relatively high near the tourist area of Weishan Island. The net diffusion fluxes indicate that the W has a potential release risk of diffusion to the overlying water in Dushan Lake. The release of Cr, Mo, and W is thought to be related to the reductive dissolution of Fe/Mn (hydr)oxides based on Pearson correlation coefficients. The R-values of Cr and W indicate that Cr and W belong to the partial continuity case. The R-value of Mo was lower than the minimum value, meaning that Mo belongs to the single diffusion type and it is difficult for Mo sediments to supply pore water.

Cyanobacterial blooms increase the release of vanadium through iron reduction and dissolved organic matter complexation in the sediment of eutrophic lakes.

Vanadium (V), a hazardous environmental contaminant, can be highly toxic to aquatic or even human life. Nonetheless, knowledge of its redox geochemistry and mobility in sediments, especially those of eutrophic lakes, remains limited. In this study, we combined in situ high-resolution sampling and laboratory simulation experiments for monitoring soluble and labile V to reveal the mobilization mechanism of V in the sediment of Lake Taihu. The results showed that the concentration of soluble V (1.18-5.22 µg L-1) exceeded the long-term ecotoxicology limitation proposed by the government of the Netherlands. The highest value appeared in summer (July to September), with an average concentration of 3.87 µg L-1, which exceeded the short-term exposure limit. The remobilization of V in summer was caused by the combined effect of the reduction of Fe(hydr)oxides and dissolved organic matter (DOM) complexation, which accelerated the release of associated Fe-bound V and increased the solubility of DOM-V. Additionally, V showed high mobility in winter, owing to the species of V(Ⅲ)/V(Ⅳ) being oxidized to V(Ⅴ) with higher solubility. It is noteworthy that the elevated remobilization of V in sediments increases the risk of V release from sediments, which poses the threat of water V pollution in Lake Taihu.

Spatiotemporal characterization of vanadium at the sediment-water interface of a multi-ecological lake.

As an emerging environmentally harmful metal, vanadium (V) deserves significant research attention due to its hazardous concentrations in aquatic environments. However, the research on the characterization of V in sediment-water interface (SWI) remains limited. In this study, seasonal sampling was conducted in algal- and macrophyte-dominated zones via the method of in situ high-resolution diffusive gradients in thin films (DGT). The concentration of dissolved V in water in algal-dominated regions (12 sites) exceeded the long-term ecotoxicology limit of 1.2 µg⋅L-1. Seasonal variations of chemical speciation of V were observed in three ecological sites. DGT-labile V at the SWI exhibited two basic patterns associated with eutrophic status, one showing sharply decreasing gradients in the vicinity of the SWI and the other showing the absence of diffusion gradient. Positive correlations were observed between the water-dissolved V and the DGT-labile V, indicating DGT-labile V is a sensitive indicator for the release of V from sediment into water. Moreover, the mobility of V was influenced by the reduction of Fe(hydr)oxides and complexation with organic matter, in particular, during periods of algal blooms. It is suggested that V contamination at the SWI of algal-dominated zones deserves additional attention.

Determining effective diffusion coefficients of chlorohydrocarbons in natural clays: Unique results from highly resolved controlled release field experiments.

This study aims to precisely determine the effective diffusion coefficients of chlorohydrocarbons in low permeable units under in-situ field conditions. To this end, two controlled release field experiments using TCE and PCE as dense non-aqueous phase liquids (DNAPLs) were conducted in two natural clayey deposits. Several months to years after the controlled DNAPL release, highly resolved concentration profiles were determined for the chlorohydrocarbons that had diffused into the clayey deposits. Effective diffusion coefficients for TCE and PCE were then determined by calibrating a 3D numerical and 1D analytical model, respectively, to the measured high-resolution concentration profiles. The simulations revealed that the effective diffusion coefficients vary by as much as a factor of four within the same low permeability unit being consistent with observed small-scale heterogeneities. The determined chlorohydrocarbon effective diffusion coefficients were further used to determine the equivalent thickness of DNAPL that would completely dissolve in an idealized, parallel-plate fracture by diffusion transport into clayey deposits for the time periods of the controlled release field experiments. The equivalent TCE and PCE DNAPL film thicknesses ranged between 36 and 581 µm, respectively, comparable and exceeding fracture apertures measured in naturally fractured clay rich deposits. Hence, films of DNAPL initially contained within fractures in clay-rich deposits can completely dissolve away within a few months to a few years due to diffusion. This stored contaminant mass poses a risk to adjacent aquifers if it is re-released due to diffusion out of the matrix after source depletion or remediation.

Temporal evolution of acid mine drainage (AMD) leachates from the abandoned tharsis mine (Iberian Pyrite Belt, Spain).

Acid mine drainage (AMD) due to the mining of sulfide deposits is one of the most important causes of water pollution worldwide. Remediation measures, especially in historical abandoned mines, require a deep knowledge of the geochemical characteristics of AMD effluents and metal fluxes, considering their high spatial and temporal evolution, and the existence of point and diffuse sources with a different response to rainfall events. This study investigates the temporal variations and hydrogeochemical processes affecting the composition of main AMD sources from the Tharsis mines (SW Spain), one of most important historical metal mining districts in the world. To address this, a fortnightly-monthly sampling was performed during two years in the main AMD sources and streams within the mine site covering different hydrological conditions. A seasonal pattern was observed linked to hydrological variations; higher pollutant concentrations were observed during the dry season (maximum values of 4,6 g/L of Al, 11,8 g/L of Fe, and 67 g/L of sulfate) and lower ones were observed during the rainy periods. Stream samples exhibited a negative correlation between electrical conductivity (EC) and flow, while positive values were observed in AMD sources, where groundwater fluxes were predominant. High flow also seems to be the main driver of Pb fluxes from AMD sources, as the concentration of Pb in waters increased notably during these events. The precipitation of secondary Fe minerals may limit the mobility of As and V, being retained in the proximity of mine sites. The concentration of Zn in waters seems to be controlled by the original grade in the metal deposit from which the waste is generated, together with the age of these wastes. The pollutant load delivered by the Tharsis mines to the surrounding water courses is very high; e.g., mean of 733 ton/yr of Al or 2757 ton/yr of Fe, deteriorating the streams and reservoirs downstream.

High cadmium pollution from sediments in a eutrophic lake caused by dissolved organic matter complexation and reduction of manganese oxide.

Eutrophication and metal pollution are global environmental problems. The risk of metal pollution is high in the eutrophic lakes because of high mobility of metal in sediments. However, the mechanism of cadmium (Cd) mobility in sediments is still unclear. Here we study the mobilization of Cd in sediments from the eutrophic Lake Taihu via monthly field monitoring of mobile Cd using diffusive gradient in thin films (DGT) and high resolution dialysis (HR-Peeper) techniques. We found a high mobility of Cd in sediments in February and March, resulting from reductive dissolution of Mn oxide mediation by high microbial activities, as shown by the similarities in distribution patterns of DGT-labile Cd and Mn. A two orders of magnitude increase in dissolved Cd concentrations (about 28 µg L-1) was observed in May and June, with dissolved Cd concentrations in overlying water about 110 times higher than the criteria continuous concentration set by Environmental Protection Agency. Hourly changes were found to coincide and correlate between dissolved Cd and dissolved organic matter (DOM) under simulated anaerobic conditions, strongly suggesting that the sudden outbreak of Cd pollution observed in the field resulted from the complexation of DOM with Cd in sediments. This was further supported by the NICA-Donnan model that more than 71% of dissolved Cd in the pore water in May and June was present as Cd-DOM complexes. Three components of DOM including humic-, tryptophan-, and tyrosine-like components in the sediments in June was identified using the fluorescence excitation emission matrix-parallel factor analysis. We found that Cd was stable complexed with tyrosine-like component. The Fourier transform infrared and two-dimensional correlation spectroscopy further revealed that Cd was bound to phenolic OH, alkene CC, alcoholic CO, aromatic CH, and alkene CH groups. Our study effectively promotes the understanding of Cd mobilization in sediments and highlights the risk of sudden Cd pollution events in the eutrophic lakes.

A selective comprehensive reversed-phase×reversed-phase 2D-liquid chromatography approach with multiple complementary detectors as advanced generic method for the quality control of synthetic and therapeutic peptides.

There is a huge, still increasing market for synthetic and therapeutic peptides. Their quality control is commonly based on a generic reversed-phase liquid chromatography (RPLC) method with C18 stationary phase and acetonitrile gradient with 0.1% trifluoroacetic acid in the mobile phase. It performs exceptionally well for a wide variety of impurities, yet structurally closely related impurities with similar sequences, not resolved in preparative RPLC, may easily coelute in the corresponding QC run as well. To address this problem an advanced generic 2D-LC impurity profiling method was developed in this work. It employs a selective comprehensive (high resolution sampling) RP×RP 2D-LC separation using a 100×2.1 mm ID column with the common acidic generic gradient in the first dimension, while RPLC under basic pH on a short 30×3 mm ID column is used in the second dimension. Recording data with a UV detector at 215 nm after 1D separation provides the common generic 1D chromatogram. However, after the 2D separation a flow splitter enabled recording of the signals of complementary detectors comprising a diode array detector (DAD) in-line with a charged aerosol detector (CAD) and a quadrupole-time-of-flight (QTOF) mass spectrometer (MS) with an electrospray ionization (ESI) source. Generic conditions of this 2D-LC method have been established through optimization of 2D stationary and mobile phase considering different pH values and buffer concentrations. The orthogonal separation principle has been documented by a number of therapeutic peptides including Exenatide, Octreotide, Cyclosporine A and Oxytocin as well as some other proprietary synthetic peptides. The information density can be further enhanced by using the QTOF-MS detector by data independent acquisition with SWATH. Through this sequential window acquisition of all theoretical fragment ion mass spectra it became possible to collect MS/MS data comprehensively in the high-resolution sampling window, thus enabling the extraction of 2D-EICs from fragment ions and the generation of 2D-contour plots of all product ions. Using Oxytocin as an example for an important therapeutic peptide, the ability of this advanced generic sRP-UV×RP-DAD-CAD-ESI-QTOF-MS/MS method with SWATH for peptide quality control is discussed.

Application of DGT/DIFS combined with BCR to assess the mobility and release risk of heavy metals in the sediments of Nansi Lake, China.

The heavy metal contamination of the aquatic ecosystem is still prevalent even after reduction of the external anthropogenic inputs of the metals. The release of labile heavy metals from the sediments into the water is a potential risk, responsible for the contamination of the aquatic system. Herein, samples of sedimentary column cores were collected in Nansi Lake, and the distribution profiles of the labile and soluble metals (Cd, Cu, Ni, Pb, and Zn) were obtained by the diffusive gradient in thin films (DGT) and the high-resolution dialysis (HR-peeper) technique. Furthermore, the mobility, bioavailability and release risk of the heavy metals were assessed using the results of geochemical sequential extraction, DGT as well as the DGT-induced fluxes in sediments (DIFS) model. The results showed that the profile characteristics of the DGT-labile and soluble heavy metals showed irregular distribution in the sediment cores and Cd, Pb, Zn had an obvious positive correlation with Fe/Mn (p < 0.05). Ni, Cu, and Zn existed primarily in the residual fraction (accounting for 58-76%), while Cd and Pb existed in the reducible fraction (accounting for 50-67%). The Cd and Ni (0.027-0.185) had higher mobility coefficients compared with Pb, Cu, and Zn (0-0.011), and positive diffusive fluxes also proved that Cd and Ni were easy to be released from the sediments. In addition, the R values of five metals (0.18-0.85) ranged between Rdiff to 0.95, indicating that all the metals had partially sustained case from the sediments solid phase. Based on the DIFS model, the five metals had weak mobility from the sediment to pore water, but the release risks in the Nansi Lake should also be of concern, especially for the highly mobile Cd and Ni in the Dushan Lake.

Evaluating the mobility and labile of As and Sb using diffusive gradients in thin-films (DGT) in the sediments of Nansi Lake, China.

Arsenic (As) and antimony (Sb) contamination in the aquatic environment have received significant attention recently due to the potential risks they pose. However, there have been few studies about the simultaneous behaviors of As and Sb, resulting in a poor understanding of their occurrence at the sediment-water interface (SWI), especially at the millimeter scale. In this study, soluble and labile concentrations of As and Sb were investigated using high-resolution dialysis (HR-Peeper) and diffusive gradients in thin films technique (DGT) in Nansi Lake, China, respectively. Results showed mean soluble concentrations of As and Sb were 5.00 µg/L and 2.05 µg/L, respectively. DGT-labile concentrations of As and Sb ranged from 0 to 0.80 µg/L and from 0.50 to 0.67 µg/L, respectively. In the vertical profile, different tends for DGT-labile concentration As and Sb were observed. The reductive dissolution of Fe/Mn (hydr)oxides was considered as a crucial driver for As release and mobility, which was supported by its significant correlation (r = 0.348, p < .05) with Fe. While DGT-labile Sb concentration was negatively correlated with DGT-labile Fe (r = -0.24, p < .05) and Mn (r = -0.324, p < .05), this may be attributed to the absorption of the Sb(III) by the green rusts in sub-oxic and mildly alkaline environments. The significant differences between DGT-labile concentration and community Bureau of Reference (BCR) sequential extraction were shown using a linear regression relationship, indicating that BCR chemical fractions cannot reflect the mobility of As and Sb in the sediment. Furthermore, the net diffusive fluxes of As and Sb based on DGT-labile concentration were 0.24 and - 0.56 µg∙m-2∙day-1, respectively. There was a potential risk of toxicity to the overlying water from As.

A new method to overall immobilization of phosphorus in sediments through combined application of capping and oxidizing agents.

A new method has been developed for improving the overall immobilization efficiency of phosphorus (P) in sediment. A capping agent (lanthanum modified bentonite, LMB) was sprinkled on the sediment surface to prevent the release of P in the top sediment layer. Meanwhile, an oxidizing agent (calcium nitrate, CN) was injected into the sediment layer (~5 cm) to immobilize labile P in deep sediment layers. High-resolution sampling techniques, including diffusive gradients in thin films (DGT) and high-resolution dialysis (HR-Peeper) were employed to investigate the fine-scale changes of labile and/or soluble nitrogen, P, sulfide and iron in sediments, respectively. The results showed that the combined application of LMB and CN had significant advantages over the individual treatments. The average concentrations of soluble reactive phosphorus (SRP) (0.01 mg/L) in the overlying water after a 68-day incubation were only 10%, 21% and 4% for the CK, LMB and CN treatments, respectively. Furthermore, the immobilization effect caused by the combined treatment reached from the sediment-water interface to a depth of 60 mm in the sediment, and the effective depth was much >20 mm caused by LMB treatment. The concentrations of SRP in the sediment profile were also lower than those of the other treatments. The results of this work indicate that the combined application of capping and oxidizing agents is a promising method to control water eutrophication by preventing the release of P from both the top and deep sediment layers.

[Simultaneous determination of chlorogenic acid and cynaroside contents in Lonicerae Japonica Flos by high resolution sampling two-dimensional liquid chromatography].

An innovative analytical method based on high resolution sampling two-dimensional liquid chromatography (HiRes 2D-LC) was established for determination of chlorogenic acid and cynaroside in Lonicerae Japonica Flos. A C18 column was used in the first dimension (1D)-LC separation with acetonitrile and 0.4% (v/v) phosphoric acid aqueous solution as mobile phases. Five heart cuts of chlorogenic acid and four heart cuts of cynaroside were stored in 2D-LC interface, which was a 5-position-10-port valve equipped with two multiple heart-cutting valves. The stored cuts were sequentially separated in the second dimension (2D)-LC. The 2D separation was carried out on an SB-Phenyl column with acetonitrile and 0.5% (v/v) acetic acid aqueous solution as mobile phases. The results showed that chlorogenic acid peaks in the 1D were well separated, whereas cynaroside peaks in the 1D were co-eluted with interferences. The above two targets were accurately quantified through a high resolution sampling mode based on continuous slice cuts of the whole target peaks. The method had good linearity, recovery and repeatability. The HiRes 2D-LC system could be used to improve separation and quantification of (un)targets in traditional Chinese medicine samples.

Seasonal antimony pollution caused by high mobility of antimony in sediments: In situ evidence and mechanical interpretation.

Antimony (Sb) mobilization in sediments and its impact on water quality remained to be studied. In this study, high-resolution dialysis (HR-Peeper) and diffusive gradients in thin films (DGT) technique were used to measure soluble Sb and labile Sb in sediment-overlying water profiles for a full year in a eutrophic region of Lake Taihu. Results showed that the highest mean concentrations of soluble Sb in overlying water (11.27 and 6.99 µg/L) appeared in December 2016 and January 2017, due to oxidation of Sb(III) to Sb(V) by Mn and Fe oxides, all of which exceeded the surface or drinking water limits set by China, United States and European Union. From April to November 2016, the concentrations of soluble Sb remained low with small monthly fluctuations and mean values ranging from 1.79 to 2.93 µg/L. This was attributed to the predominance of insoluble Sb(III) in sediments under anoxic conditions. The concentration of soluble Sb was slightly higher in summer than in autumn, due to the complexation of Sb(III) with DOM, as shown under anaerobic incubation. The mobility of inorganic Sb in sediments was mostly determined by the transition between Sb(III) and Sb(V), with Sb pollution in bottom water during winter being of concern.

Seasonal changes of lead mobility in sediments in algae- and macrophyte-dominated zones of the lake.

This study examined lead (Pb) pollution in algae- and macrophytes-dominated sediments, using diffusive gradient in thin films (DGT) and dialysis (HR-Peeper) techniques. Lead pollution varied by season in the two different ecotype sediments. In the algae-dominated zone, the highest concentrations of DGT-labile Pb and dissolved Pb occurred in April and July, respectively. The reductive dissolution of Fe/Mn oxides was identified as an important driver for Pb releases in April and July. This was supported by the decrease of the reducible fraction of Pb in sediments during those sampling periods. Furthermore, dissolved organic matter (DOM) complexation with Pb in sediments also significantly increased the dissolved Pb concentrations in July. The Pb-DOM complexes accounted for 95% of the total chemical species of Pb in pore water, calculated by Visual MINTEQ 3.1 model. Low concentrations of labile and dissolved Pb were observed in October and January; these resulted from the formation of Pb-sulfide precipitates and adsorption by Fe/Mn oxides. It was supported by the high rate of Pb(HS)2 precipitation (saturation index > 0), at 36%, in October samples and the high reducible fraction of Pb in sediments in January samples. In the macrophytes-dominated region, there was a decrease of labile and dissolved Pb concentrations in April and July. It is likely because of the uptake of Pb by submerged macrophyte roots and the Fe/Mn plaques in the root surface. High concentrations of labile and dissolved Pb were observed in October and January, likely resulting from the DOM complexation with Pb in sediments. This was supported by the fact that the Pb-DOM complexes accounted for 90% and 87% of the total chemical species of Pb in October and January, respectively.

Determination of in situ biodegradation rates via a novel high resolution isotopic approach in contaminated sediments.

A key challenge in conceptual models for contaminated sites is identification of the multiplicity of processes controlling contaminant concentrations and distribution as well as quantification of the rates at which such processes occur. Conventional protocol for calculating biodegradation rates can lead to overestimation by attributing concentration decreases to degradation alone. This study reports a novel approach of assessing in situ biodegradation rates of monochlorobenzene (MCB) and benzene in contaminated sediments. Passive diffusion samplers allowing cm-scale vertical resolution across the sediment-water interface were coupled with measurements of concentrations and stable carbon isotope signatures to identify zones of active biodegradation of both compounds. Large isotopic enrichment trends in 13C were observed for MCB (1.9-5.7‰), with correlated isotopic depletion in 13C for benzene (1.0-7.0‰), consistent with expected isotope signatures for substrate and daughter product produced by in situ biodegradation. Importantly in the uppermost sediments, benzene too showed a pronounced 13C enrichment trend of up to 2.2‰, providing definitive evidence for simultaneous degradation as well as production of benzene. The hydrogeological concept of representative elementary volume was applied to CSIA data for the first time and identified a critical zone of 10-15 cm with highest biodegradation potential in the sediments. Using both stable isotope-derived rate calculations and numerical modeling, we show that MCB degraded at a slower rate (0.1-1.4 yr-1 and 0.2-3.2 yr-1, respectively) than benzene (3.3-84.0 yr-1) within the most biologically active zone of the sediment, contributing to detoxification.

Internal phosphorus loading from sediments causes seasonal nitrogen limitation for harmful algal blooms.

It is proposed that the internal loading of phosphorus (P) from sediments plays an important role in seasonal nitrogen (N) limitation for harmful algal blooms (HABs), although there is a lack of experimental evidence. In this study, an eutrophic bay from the large and shallow Lake Taihu was studied for investigating the contribution of internal P to N limitation over one-year field sampling (February 2016 to January 2017). A prebloom-bloom period was identified from February to August according to the increase in Chla concentration in the water column, during which the ratio of total N to total P (TN/TP) exponentially decreased with month from 43.4 to 7.4. High-resolution dialysis (HR-Peeper) and diffusive gradients in thin films (DGT) analysis showed large variations in the vertical distribution of mobile P (SRP and DGT-labile P) in sediments, resulting in the SRP diffusion flux at the sediment-water interface ranging from -0.01 to 6.76mg/m2/d (minus sign denotes downward flux). Significant and linear correlations existed between SRP and soluble Fe(II) concentrations in pore water, reflecting that the spatial-temporal variation in mobile P was controlled by microbe-mediated Fe redox cycling. Mass estimation showed that the cumulative flux of SRP from sediments accounted for 54% of the increase in TP observed in the water column during the prebloom-bloom period. These findings are supported by the significantly negative correlation (p<0.01) observed between sediment SRP flux and water column TN/TP during the same period. Overall, these results provide solid evidence for the major role of internal P loading in causing N limitation during the prebloom-bloom period.

High-resolution two-dimensional liquid chromatography analysis of key linker drug intermediate used in antibody drug conjugates.

In this manuscript, the application of high-resolution sampling (HRS) two-dimensional liquid chromatography (2D-LC) in the detailed analysis of key linker drug intermediate is presented. Using HRS, selected regions of the primary column eluent were transferred to a secondary column with fidelity enabling qualitative and quantitative analysis of linker drugs. The primary column purity of linker drug intermediate ranged from 88.9% to 94.5% and the secondary column purity ranged from 99.6% to 99.9%, showing lot-to-lot variability, significant differences between the three lots, and substantiating the synthetic and analytical challenges of ADCs. Over 15 impurities co-eluting with the linker drug intermediate in the primary dimension were resolved in the secondary dimension. The concentrations of most of these impurities were over three orders of magnitude lower than the linker drug. Effective peak focusing and high-speed secondary column analysis resulted in sharp peaks in the secondary dimension, improving the signal-to-noise ratios. The sensitivity of 2D-LC separation was over five fold better than conventional HPLC separation. The limit of quantitation (LOQ) was less than 0.01%. Many peaks originating from primary dimension were resolved into multiple components in the complementary secondary dimension, demonstrating the complexity of these samples. The 2D-LC was highly reproducible, showing good precision between runs with%RSD of peak areas less than 0.1 for the main component. The absolute difference in the peak areas of impurities less than 0.1% were within ±0.01% and for impurities in the range of 0.1%-0.3%, the absolute difference were ±0.02%, which are comparable to 1D-LC. The overall purity of the linker drug intermediate was determined from the product of primary and secondary column purity (HPLC Purity=%peak area of main component in the primary dimension×%peak area of main component in the secondary dimension). Additionally, the 2D-LC separation enables the determination of potential impurities that could impact the downstream process, like ADCs stability, efficacy and patient safety. Peak capacity of this magnitude, sensitivity and reproducibility of 2D-LC for resolving structurally similar impurities co-eluting with the main component has not been demonstrated to date. This application clearly demonstrates the power of 2D-LC in detailed analysis of structurally similar, co-eluting impurities from key linker drug intermediate used in ADCs that is impossible to achieve by conventional 1D-LC.

Unravelling organic matter and nutrient biogeochemistry in groundwater-fed rivers under baseflow conditions: Uncertainty in in situ high-frequency analysis.

In agricultural catchments, diffuse nutrient fluxes (mainly nitrogen N and phosphorus P), are observed to pollute receiving waters and cause eutrophication. Organic matter (OM) is important in mediating biogeochemical processes in freshwaters. Time series of the variation in nutrient and OM loads give insights into flux processes and their impact on biogeochemistry but are costly to maintain and challenging to analyse for elements that are highly reactive in the environment. We evaluated the capacity of the automated monitoring to capture typically low baseflow concentrations of the reactive forms of nutrients and OM: total reactive phosphorus (TRP), nitrate nitrogen (NO3-N) and tryptophan-like fluorescence (TLF). We compared the performance of in situ monitoring (wet chemistry analyser, UV-vis and fluorescence sensors) and automated grab sampling without instantaneous analysis using autosamplers. We found that automatic grab sampling shows storage transformations for TRP and TLF and do not reproduce the diurnal concentration pattern captured by the in situ analysers. The in situ TRP and fluorescence analysers respond to temperature variation and the relationship is concentration-dependent. Accurate detection of low P concentrations is particularly challenging due to large errors associated with both the in situ and autosampler measurements. Aquatic systems can be very sensitive to even low concentrations of P typical of baseflow conditions. Understanding transformations and measurement variability in reactive forms of nutrients and OM associated with in situ analysis is of great importance for understanding in-stream biogeochemical functioning and establishing robust monitoring protocols.