the Creative Commons Attribution 4.0 License.
the Creative Commons Attribution 4.0 License.
| 30 Jul 2026
Development and first application of a new online system for continuous measurement of oxidative potential
Abstract. This study introduces a new online system for measuring oxidative potential (OP), using the dithiothreitol (DTT) assay with a direct-to-reagent approach. The instrument enables real-time OPDTT assessments with 5-minute resolution and a competitive instrumental detection limit (0.066 nmolDTT min-1). Its performance was evaluated using model compounds and under controlled experiments in the CESAM atmospheric simulation chamber. The system exhibited clear and consistent responses to major OP-active species, including quinones, copper, manganese, secondary organic aerosols (SOA), and black carbon. In chamber experiments conducted without photochemistry, the OPvDTT values remained consistently lower than during experiments involving photochemistry, despite the higher concentrations of metals and similar particulate masses. This finding underscores the critical role of photochemically produced species in driving OP. Overall, the instrument demonstrated its capacity to monitor OPvDTT at a high temporal resolution, paving the way for a deeper understanding of the influences on OP from emission sources and atmospheric processes, particularly transient ones.
- Preprint
(1054 KB) - Metadata XML
-
Supplement
(241 KB) - BibTeX
- EndNote
Status: open (until 27 Oct 2026)
- RC1: 'Comment on egusphere-2026-4293', Anonymous Referee #1, 10 Aug 2026 reply
-
RC2: 'Comment on egusphere-2026-4293', Anonymous Referee #2, 02 Oct 2026
reply
Review comments
This manuscript by Audoux et al., entitled “Development and first application of a new online system for continuous measurement of oxidative potential”, presents the development and initial evaluation of a continuous online system for measuring aerosol oxidative potential (OP) using the dithiothreitol (DTT) assay. By combining a particle-into-liquid sampler (PILS), a direct-to-reagent approach, and spectrophotometric detection, the authors developed a system capable of continuously measuring OP with a temporal resolution of 5 minutes. The instrument was characterized using several model compounds and further evaluated through experiments conducted in the CESAM atmospheric simulation chamber to investigate the influence of different aerosol components and photochemical processes on OP.
The development of online OP measurement techniques is important, particularly for capturing short-lived species, rapid changes in aerosol oxidative properties and minimizing potential artifacts associated with conventional offline sampling and analysis. The continuous measurement approach and the combination of laboratory characterization with simulation chamber experiments are strengths of this study. Overall, the manuscript is well organized and clearly written, and the results provide useful insights into the development and potential applications of online OP measurement techniques.
However, several methodological aspects require further clarification and evaluation, particularly regarding potential contributions from gas-phase species and water-insoluble particulate components to the measured OP, as well as the validity of DTT consumption measurements across varying aerosol concentrations. Addressing these issues would help strengthen the reliability of the measurements and clarify the applicability of the developed system. Therefore, I recommend that the authors address the following comments before the manuscript can be considered for publication.
Major comments:
- The potential contribution of gas-phase species to the measured OP is an important concern. The authors acknowledge this limitation in Section 3.4 and propose using HEPA filters and denuders to distinguish gas-phase and particle-phase contributions in future studies. However, given that the authors are already aware of this issue and have identified possible approaches to address it, it is unclear why such tests were not performed as part of the current instrument evaluation. Although the authors examined the responses to O3 and NO2, other gas-phase compounds, particularly oxygenated organic products formed during photochemical oxidation, may also contribute to DTT consumption. This is especially relevant to the CESAM experiments, where the enhanced OP under photochemical conditions is partly attributed to secondary organic aerosol formation. I believe that experimentally evaluating potential gas-phase contributions is important for establishing the reliability of this new online OP measurement system. Could the authors explain why these tests were not included in the current study and, preferably, provide additional measurements to assess the potential gas-phase interference?
- In Section 2.1.1, the authors state that the PILS is used to collect the PB-soluble fraction of aerosols. However, according to the description of the experimental setup, the collected sample (dissolved + particle fraction) is mixed directly with DTT, while the 0.2 μm filter is placed after the DTT and DTNB reaction loops. Does this mean that water-insoluble particles remain in the reaction mixture and may also contribute to DTT consumption? If so, the system would not exclusively measure the water-soluble fraction of aerosol OP, but could also capture contributions from water-insoluble components. This is not necessarily a limitation and may even provide a more comprehensive assessment of particle OP. However, the authors should clarify at which stage insoluble particles are removed, whether they are present during the DTT reaction, and how the measured OP should be defined and interpreted accordingly.
- The initial DTT concentration and reaction residence time are fixed in the current system, whereas the concentrations and chemical composition of sampled aerosols can vary substantially. In conventional offline DTT assays, sample loading is typically adjusted to ensure that DTT depletion remains within an appropriate linear reaction regime. In Section 3.1, the authors have already observed a deviation from linearity at higher concentrations of 9,10-PQ, indicating that the system has a limited linear operating range. This raises an important question regarding its application to continuously changing aerosol concentrations, particularly during the chamber experiments and future ambient measurements. How can the authors ensure that DTT consumption remains within the valid linear range under such conditions? Have the authors evaluated the fraction of DTT consumed during the chamber experiments, and were any measurements approaching or exceeding the linear operating range? Furthermore, since the current system determines DTT consumption at a fixed reaction time rather than from multiple reaction time points, it would be helpful to clarify how the linearity of DTT consumption over the reaction period was validated. I suggest that the authors better define the valid operating range of the instrument and explain how measurements outside this range can be identified or handled.
Minor comments:
- Lines 118-122: The authors mention that the reaction temperature was set to 37°C but the actual measured temperature was approximately 30°C. Since DTT consumption kinetics are temperature-dependent, this difference may affect the measured OP values and their comparability with conventional DTT assays performed at 37°C. Could the authors clarify how stable the reaction temperature was during the experiments, and whether the laboratory calibration and chamber measurements were performed under comparable temperature conditions? A brief discussion of the potential influence of this temperature difference on the reported OP values would be helpful. In addition, in Lines 280-289, the authors attribute the lower DTT consumption rates compared with previous studies mainly to differences in the initial DTT concentration and analytical conditions. Could the lower reaction temperature also contribute to this discrepancy?
- Lines 130-131: The authors mention that the DTT solution was maintained below 4°C using ice and a thermoregulated bottle holder. Does this setup require regular replacement of ice during operation, or can the temperature be maintained automatically? Please clarify whether the DTT solution can remain below 4°C throughout the reported 20-hour autonomous operation period without manual intervention.
- Section 2.2.3, Lines 240-243: The authors state that particle mass concentrations were calculated from DMPS measurements using an average particle density derived from ToF-ACSM data. Could the authors provide more details on how the particle density was estimated, including the assumed densities of individual chemical components? It would also be helpful to clarify why DMPS-derived mass concentrations were preferred over those obtained from ToF-ACSM combined with AE33 measurements?
Technical comments:
- Figure 1: The DTT reaction loop appears to be labelled as 6.5 mL, whereas Section 2.1.1 reports a volume of 6.3 mL. Please check and ensure consistency.
- Table 1: The particle diameter range reported for the BioSampler collection efficiency is given as 0.05-0.08 nm. Should this be 0.05-0.08 μm? Please check the units.
- Lines 277-279: “The reported concentrations correspond to the those injected into the system” contains an extra “the”.
- Lines 283-285: “Charrier and Anastasio (2012) reported a DTT consumption rates” should be corrected to “Charrier and Anastasio (2012) reported DTT consumption rates”.
- Lines 313-315: The sentence “DTT decay is the results of the reactivity of a complex mixture” contains a grammatical error. Please revise to “DTT decay results from the reactivity of a complex mixture”.
- Lines 323-326: The phrase “all datasets were reprocessed and at a unified temporal resolution of 15 minutes” is grammatically incorrect. Please removed the “and”.
- Lines 378-381: There appear to be redundant citation numbers (“39,65,66”) following the author-year citations. Please check and remove them if they are remnants of a previous reference format.
- Lines 432-435: The phrase “Under pre operating conditions” is unclear and may contain a typographical error. Please clarify the intended meaning.
- Lines 456-459: “This provide a mechanistic understanding” should be corrected to “This provides a mechanistic understanding”.
Citation: https://doi.org/10.5194/egusphere-2026-4293-RC2 -
RC3: 'Comment on egusphere-2026-4293', Anonymous Referee #3, 17 Oct 2026
reply
General comments
This manuscript presents a new online system for measuring aerosol oxidative potential via the DTT assay, using a fully continuous, direct-to-reagent configuration (PILS sampling, flow-loop reaction, and continuous absorbance detection) that provides 5-minute time resolution. The instrument is characterized with model compounds (9,10-PQ, 1,4-NQ, Cu) and evaluated in the CESAM simulation chamber under photochemical and non-photochemical conditions.
The development of continuous online OP measurement techniques is highly relevant to the scope of AMT, and the fully continuous direct-to-reagent DTT design is, to my knowledge, a first among absorbance-based systems; the combination of laboratory characterization with simulation-chamber evaluation is a sensible validation strategy. The manuscript is generally well organized and clearly written.
However, as a measurement-technique paper, the manuscript still falls short of AMT’s requirements on two counts: (i) the metrological traceability of the new measurement has not been anchored against any reference method, and (ii) several aspects of the signal quantification and uncertainty budget (fixed-residence-time kinetics, air-volume normalization, linear operating range, blank/baseline treatment) are insufficiently documented to allow reproduction and independent assessment of the reported performance.
These issues, detailed below, must be resolved before the manuscript can be considered for publication. I note that Referees #1 and #2 have already raised several related points (field applicability, attribution of chamber results, gas-phase contributions, insoluble fractions); my comments are intended to be complementary, and where they overlap I explicitly support those concerns.
Major comments
Six major issues concerning the metrological validity and documentation of the new measurement system are detailed below.
1. Non-standard DTT concentration (10 µM) and cross-comparability of the assay
The reaction-loop DTT concentration is 10 µM (20 µM stock diluted 1:1), whereas the consensus offline protocol and virtually all existing online OPDTT systems operate at 50–212 µM (the manuscript’s own Table 1). As demonstrated systematically by Lin and Yu (2019), the initial DTT concentration fundamentally reshapes the reaction kinetics and the concentration–response relationship, and lower DTT-to-analyte ratios can introduce competitive complexation artifacts (e.g., shifts in metal–ligand speciation).
I acknowledge the authors’ argument, based on Sameenoi et al. (2012), that lower DTT concentrations can improve analytical sensitivity, and I agree with Referee #1 that this caution should also be applied to any direct comparison of LODs across systems. However, the converse obligation follows: the authors must demonstrate that the 10 µM system produces chemically meaningful, interpretable OPDTT values, and they must clearly state that the resulting data are not directly comparable with the large body of ambient OPDTT data generated at 100 µM.
Specifically, the authors should either (a) modify the optical path or fluidics to allow operation at the standard 100 µM (e.g., replace the 10-cm LWCC with a shorter path length, or introduce an online post-reaction dilution before detection), or (b) retain the current configuration but provide a rigorous kinetic justification. This justification should include multi-point DTT decay curves demonstrating that pseudo-first-order behavior holds within the analytical window and across the full linear range reported in Section 3.1, together with a discussion of DTT-to-metal/quinone ratios. At present, only single-point endpoint responses at a fixed residence time are shown, which is not sufficient to establish the kinetic validity of the measurement.
2. Absence of side-by-side validation against a reference DTT assay
For any new automated analytical system, a direct cross-validation against the reference (manual) method on identical samples is the fundamental anchor of accuracy, and it is explicitly listed among AMT’s referee criteria (traceability of results). The manuscript contains calibration with model compounds, but no comparison of the automated system with a conventional manual DTT assay for the same standard solutions or generated-aerosol extracts.
Such an experiment requires no field campaign and is entirely feasible with the existing laboratory setup: identical solutions can be processed through both the prototype and a bench-scale DTT assay under matched (or deliberately mismatched, to probe the 10 µM issue) reaction conditions. The authors state that an offline intercomparison is planned as “future work”; for an instrument-development paper, this validation should be part of the present study, or at minimum a clearly designed laboratory comparison of the automated and manual assays for the three model compounds should be added. Without it, systematic artifacts introduced by the continuous-flow, fixed-residence-time configuration cannot be ruled out.
3. Quantification of the continuous-flow signal: endpoint kinetics and air-volume normalization
I support and extend the concerns of Referee #1 regarding the calculation of OPDTT:
- Endpoint signal as a rate. The system measures remaining DTT at a fixed residence time rather than a decay slope over multiple time points. The validity of converting this endpoint absorbance into a DTT consumption rate (Eq. 1) rests on the assumption that DTT consumption is (approximately) linear in time over the residence window and that the reaction has not entered the plateau regime. This assumption is not demonstrated for any of the model compounds, let alone for complex chamber mixtures; multi-point kinetic evidence (see Comment 1) would resolve this, and at minimum the authors must explain and justify it explicitly.
- Air-volume normalization (Eq. 2). The nominal residence time of the 6.3 mL reaction loop at 0.50 mL min-1 is ~12.6 min, yet Vair in Eq. (2) is computed from the 5-minute system response time, with an additional 20-min lag between sampling and detection. As written, the physical meaning of OPvDTT (nmol min-1 m-3) is therefore ambiguous. The authors should provide a clear mass-balance derivation showing which air volume corresponds to the liquid sample analyzed in each 5-minute interval, and how the residence time, response time, and lag enter the calculation. Please also state whether the OP time series in Figure 4 has been time-shifted to account for the 20-min lag before comparison with the ancillary measurements.
- Blank correction. The interpolation of analytical blanks by linear regression (Section 2.2.4) assumes linear DTT ageing between blanks; please report the typical spacing between blanks, the observed drift rate, and the fraction of valid sampling time lost to blanks and transition periods.
4. Validity of the linear operating range under chamber conditions
The authors observed deviation from linearity at the highest 9,10-PQ concentrations, confirming a limited linear dynamic range at 10 µM DTT (as also noted by Referee #2). For the chamber experiments, however, no information is given on the fraction of DTT consumed — that is, whether any measurements approached or exceeded the linear range with the evolving mixtures.
The authors should (i) report the estimated DTT depletion fraction throughout the chamber experiments, (ii) define the upper limit of the valid operating range in terms of a measurable quantity, and (iii) describe how out-of-range or non-linear measurements would be flagged during autonomous operation. This is essential for the future ambient deployment of the system, where aerosol loading cannot be pre-adjusted as in the laboratory.
5. Transparency of blank/baseline data and the Cu detection limit
In Figures 4 and S2, HEPA-blank periods are shown only as grey shaded blocks; the underlying data points and their variability are not displayed. For a continuous-measurement prototype, the absolute level, drift, and scatter of the blank are central to the credibility of the reported instrumental LOD (0.066 ± 0.016 nmol min-1) and of the signal-to-noise ratio during chamber experiments. Please display the actual blank data points (with uncertainty) in these figures, and clarify the exact mathematical procedure used for baseline subtraction.
Furthermore, the analyte-specific LOD for Cu (2.0 µM; ~2.6 µg m-3 equivalent) is nearly two orders of magnitude poorer than for 9,10-PQ. Given the known propensity of Cu may adsorb to and bleed from flow path, this raises the question of carryover/memory effects and blank-subtraction bias rather than intrinsic chemistry. The authors should present the raw (uncorrected) Cu calibration data, the absolute blank values subtracted, and a discussion of whether carryover between sequential injections was observed, including evidence that the campaign-level 0.05% HNO3 rinse fully removes adsorbed metals from the flow path.
6. Benchmarking of chemical responses against previous studies
The chemical responses of the model compounds (Figure 3) are discussed only qualitatively relative to literature values. For an instrumentation paper, a quantitative comparison of response factors (DTT consumption per µM of Cu, Mn, 1,4-NQ, 9,10-PQ) with published offline and online systems is needed — ideally as a summary table — together with a kinetic explanation of the differences. The manuscript currently attributes a ~5-fold discrepancy versus Charrier and Anastasio (2012) to “assay conditions” in a single sentence.
This is directly connected to Comments 1 and 2: without such a benchmark, the reader cannot judge whether the continuous-flow configuration introduces mass-transfer limitations or systematic underestimation.
Minor comments
- The reaction temperature was set to 37°C but measured at ~30°C. Since DTT kinetics are temperature-dependent, please report the stability of the measured temperature, confirm that calibration and chamber experiments were conducted under comparable conditions, and discuss the potential impact on the reported OPDTT values and on the discrepancy noted in Section 3.1 (also raised by Referee #2).
- Please clarify whether the ice/thermoregulated holder maintaining DTT below 4°C requires manual intervention during the 20-hour autonomous operation period.
- Figure 1 labels the reaction loop as 6.5 mL while the text reports 6.3 mL; Table 1 reports a BioSampler collection efficiency range of “0.05–0.08 nm”, which should presumably be µm. Please correct for consistency.
- Given the placement of the 0.2 µm filter after the reaction loops, the measured signal includes contributions from water-insoluble particles during the reaction; the definition and intended interpretation of “OPDTT” for this system should be stated explicitly (as also requested by Referee #2), including the gas-phase contribution acknowledged in Section 3.4.
- Given the acknowledged contributions of gas-phase species and matrix absorption (Section 3.4), statements in the abstract and conclusion such as “clear and consistent responses” to specific species should be tempered, consistent with Referee #1’s comment on component attribution.
- Please reconsider the phrase “first application” in the title in view of the laboratory/chamber-only evaluation presented.
Citation: https://doi.org/10.5194/egusphere-2026-4293-RC3
Viewed
| HTML | XML | Total | Supplement | BibTeX | EndNote | |
|---|---|---|---|---|---|---|
| 252 | 110 | 28 | 390 | 58 | 35 | 30 |
- HTML: 252
- PDF: 110
- XML: 28
- Total: 390
- Supplement: 58
- BibTeX: 35
- EndNote: 30
Viewed (geographical distribution)
| Country | # | Views | % |
|---|
| Total: | 0 |
| HTML: | 0 |
| PDF: | 0 |
| XML: | 0 |
- 1
This manuscript describes a new online DTT system based on a continuous-flow, direct-to-reagent design. Aerosols collected by the PILS are mixed with DTT shortly after collection, followed by reaction in a flow loop and continuous absorbance measurement. The system was characterized using several model compounds and then tested in the CESAM chamber under photochemical and non-photochemical conditions. I found the continuous-flow design interesting, particularly because it shortens the time between aerosol collection and reaction with DTT. However, several points regarding the method validation and interpretation of the chamber results need further clarification.
Major comments
I do not necessarily think that a new field campaign is required for this manuscript, but the authors should discuss this limitation more clearly. In particular, I would like to better understand whether the current sensitivity, baseline stability, reagent consumption, and potential matrix effects are expected to allow reliable measurements at typical ambient OP levels. At present, the manuscript sometimes reads as if field applicability has already been demonstrated, whereas the current study is more appropriately viewed as an initial laboratory/chamber evaluation.
Also, the present assay uses a substantially lower DTT concentration than most previous online DTT systems. Since the authors themselves note that lower DTT concentration can increase analytical sensitivity, direct comparison of absolute LOD values among systems should be made cautiously. I would be interested to see a clearer discussion of what the reported LOD means for actual ambient measurements.
For this reason, I do not think the current data clearly demonstrate independent responses to Mn, SOA, or BC. In particular, the “BC” was generated from wood-pellet combustion and therefore represents a complex combustion aerosol, and the ACSM measurement of OA should not automatically be interpreted as SOA. Statements such as “clear and consistent responses,” “main driver,” or “directly attribute” should therefore be reconsidered. The results are convincing as time-resolved measurements of the integrated OP of evolving chamber mixtures, but component-specific attribution is less certain.
I am also unclear about the air-volume normalization in Eq. (2). The 6.3 mL reaction loop and the combined liquid flow of 0.50 mL min-1 correspond to a nominal residence time of about 12.6 min, whereas the air volume is calculated using the 5 min system response time. Please provide a clearer explanation or mass-balance derivation showing why the 5 min response time is the appropriate quantity for the calculation.
Minor comments