Posts Tagged 'methods'

Technical note: interpreting pH changes

The number and quality of ocean pH measurements have increased substantially over the past few decades such that trends, variability, and spatial patterns of change are now being evaluated. However, comparing pH changes across domains with different initial pH values can be misleading because a pH change reflects a relative change in the hydrogen ion concentration ([H+], expressed in mol kg−1) rather than an absolute change in [H+]. We recommend that [H+] be used in addition to pH when describing such changes and provide three examples illustrating why.

Continue reading ‘Technical note: interpreting pH changes’

Evaluation of actin as a reference for quantitative gene expression studies in Emiliania huxleyi (Prymnesiophyceae) under ocean acidification conditions

Gene expression studies of marine phytoplankton under ocean acidification conditions are frequently based on relative measurements, with actin commonly used as a reference gene. Evidence from other organisms suggests that actin gene expression may be regulated by environmental conditions, compromising the role of actin as a reference gene. In this work the reliability of actin as a reference gene for ocean acidification experimental conditions (high CO2 vs low CO2) in two different metabolic states (acclimated metabolism vs perturbed metabolism) for the coccolithophore Emiliania huxleyi was tested. The transcriptional response of the actin (act) is compared with the expression of specific target genes associated with inorganic carbon uptake (α-carbonic anhydrase: αca1) and assimilation (RuBisCO: rbcL), which was regulated under the experimental conditions. Our results showed act expression instability in experimental conditions, evidencing that act is not a reliable reference gene for studies assessing the effect of ocean acidification on Emiliania huxleyi. Furthermore, when the act-based normalization was quantitatively tested, rbcL and αca1 expression were compromised, leading us to conclude that absolute gene expression quantification should be considered as a potentially reliable alternative for studying gene expression under ocean acidification conditions

Continue reading ‘Evaluation of actin as a reference for quantitative gene expression studies in Emiliania huxleyi (Prymnesiophyceae) under ocean acidification conditions’

Spectrophotometric determination of the bicarbonate dissociation constant in seawater


  • K1K2 values were obtained via spectrophotometric pH measurements.
  • New method improved precision in CO2 system dissociation constants.
  • New K2 parameterization improves internal consistency of CO2 system calculations.


The aqueous carbon dioxide (CO2) system stoichiometric dissociation constants K1 and K2 express the relative concentrations of CO2, HCO3 (bicarbonate), and CO32− (carbonate) in terms of pH. These constants are critical in the study of seawater and the oceans because any mathematical expression that relates the four major CO2 system parameters (pH, here expressed on the total hydrogen ion concentration scale, pHT; total dissolved inorganic carbon, CT; total alkalinity, AT; and CO2 fugacity, fCO2) requires the use of K1 and K2. Uncertainties associated with current characterizations of pK1 and pK2 (where pK = −log K), on the order of 0.01 and 0.02, limit the accuracy of marine CO2 system calculations. This work reports the results of a spectrophotometric method to experimentally determine the product K1K2 over environmentally relevant ranges of temperature (288.15 ≤ T ≤ 308.15 K) and salinity (19.6 ≤ Sp ≤ 41) where Sp denotes the practical salinity scale. Using previously published parameterizations of K1, values of pK2 could then be calculated from the new K1K2 values. The resulting set of pK2 values was fitted as a function of Sp and T to obtain a new pK2 parameterization (denoted as SWpK2) calculated with the K1 of Waters and Millero (2013) as revised by Waters et al. (2014)SWpK2 = 116.8067 – 3655.02 T−1 – 16.45817 ln T + 0.04523 Sp – 0.615 Sp0.5 – 0.0002799 Sp2 + 4.969 (Sp/T)

The average root mean square deviation between the equation and the observed data is 0.003. Residuals of this pK2 fitting function (i.e., measured pK2 minus parameterized pK2) are substantially smaller than the residuals obtained in previous works. Similarly, the total standard uncertainty in pK2 is reduced from 0.015 (previous characterizations) to 0.010 (this work). Internal consistency assessments (comparisons of measured versus calculated values of ATCT, pHT, and fCO2) were used to evaluate the computational utility of the new K2 parameterization. Assessments from both laboratory and shipboard data indicate that the internal consistency of CO2 system calculations is improved using the K2 parameterization of this work. This new K2 parameterization provides the most precise, and potentially the most accurate, bicarbonate dissociation constant characterization presently available for open ocean conditions.

Continue reading ‘Spectrophotometric determination of the bicarbonate dissociation constant in seawater’

Synoptic assessment of coastal total alkalinity through community science

Comprehensive sampling of the carbonate system in estuaries and coastal waters can be difficult and expensive because of the complex and heterogeneous nature of near-shore environments. We show that sample collection by community science programs is a viable strategy for expanding estuarine carbonate system monitoring and prioritizing regions for more targeted assessment. ‘Shell Day’ was a single-day regional water monitoring event coordinating coastal carbonate chemistry observations by 59 community science programs and seven research institutions in the northeastern United States, in which 410 total alkalinity (TA) samples from 86 stations were collected. Field replicates collected at both low and high tides had a mean standard deviation between replicates of 3.6 ± 0.3 µmol kg−1 (σmean ± SE, n = 145) or 0.20 ± 0.02%. This level of precision demonstrates that with adequate protocols for sample collection, handling, storage, and analysis, community science programs are able to collect TA samples leading to high-quality analyses and data. Despite correlations between salinity, temperature, and TA observed at multiple spatial scales, empirical predictions of TA had relatively high root mean square error >48 µmol kg−1. Additionally, ten stations displayed tidal variability in TA that was not likely driven by low TA freshwater inputs. As such, TA cannot be predicted accurately from salinity using a single relationship across the northeastern US region, though predictions may be viable at more localized scales where consistent freshwater and seawater endmembers can be defined. There was a high degree of geographic heterogeneity in both mean and tidal variability in TA, and this single-day snapshot sampling identified three patterns driving variation in TA, with certain locations exhibiting increased risk of acidification. The success of Shell Day implies that similar community science based events could be conducted in other regions to not only expand understanding of the coastal carbonate system, but also provide a way to inventory monitoring assets, build partnerships with stakeholders, and expand education and outreach to a broader constituency.

Continue reading ‘Synoptic assessment of coastal total alkalinity through community science’

A system for the determination of surface water pCO2 in a highly variable environment, exemplified in the southern Baltic Sea


  • The system for pCO2 measurements, data storage and ship-to-shore transmission is presented.
  • In the open Baltic Sea waters the pCO2 measurements obtained an accuracy of ±1.3 µatm met the state-of-the-art requirements (±2.0 µatm).
  • We discuss redefining requirements for quality control and assurance for pCO2 measurements in the coastal zone.


Measurement of pCO2 in highly dynamic coastal zones such as the southern Baltic Sea presents many challenges. In this study, we designed a system to measure pCO2 and then validated it in a series of laboratory and seagoing tests. The fast response time of the system was shown to provide a better resolution of CO2 system gradients. In the open waters of the Baltic Sea, the accuracy of the pCO2 measurements (±1.3 µatm) met the requirements of the ICOS (±2.0 µatm). In the coastal zone, there was less consistency between pCO2, DIC and pH measurements, suggesting the need to redefine the quality assurance and control requirements for the measurement of pCO2 in dynamic regions.

Continue reading ‘A system for the determination of surface water pCO2 in a highly variable environment, exemplified in the southern Baltic Sea’

Assimilating synthetic Biogeochemical-Argo and ocean colour observations into a global ocean model to inform observing system design

A set of observing system simulation experiments was performed. This assessed the impact on global ocean biogeochemical reanalyses of assimilating chlorophyll from remotely sensed ocean colour and in situ observations of chlorophyll, nitrate, oxygen, and pH from a proposed array of Biogeochemical-Argo (BGC-Argo) floats. Two potential BGC-Argo array distributions were tested: one for which biogeochemical sensors are placed on all current Argo floats and one for which biogeochemical sensors are placed on a quarter of current Argo floats. Assimilating BGC-Argo data greatly improved model results throughout the water column. This included surface partial pressure of carbon dioxide (pCO2), which is an important output of reanalyses. In terms of surface chlorophyll, assimilating ocean colour effectively constrained the model, with BGC-Argo providing no added benefit at the global scale. The vertical distribution of chlorophyll was improved by assimilating BGC-Argo data. Both BGC-Argo array distributions gave benefits, with greater improvements seen with more observations. From the point of view of ocean reanalysis, it is recommended to proceed with development of BGC-Argo as a priority. The proposed array of 1000 floats will lead to clear improvements in reanalyses, with a larger array likely to bring further benefits. The ocean colour satellite observing system should also be maintained, as ocean colour and BGC-Argo will provide complementary benefits.

Continue reading ‘Assimilating synthetic Biogeochemical-Argo and ocean colour observations into a global ocean model to inform observing system design’

Seasonality and biological forcing modify the diel frequency of nearshore pH extremes in a subarctic Alaskan estuary

Acidification in nearshore waters is influenced by a multitude of drivers that shape the dynamics of pH and carbonate chemistry variability on diurnal, seasonal, and yearly time scales. Monitoring efforts aimed at characterizing high temporal variability are lacking in many nearshore systems, particularly in high‐latitude regions such as Alaska. To rectify this, a nearshore acidification sensor array was established in the Fall of 2017 within Kachemak Bay, Alaska. Presented here are the results from the first year of these deployments, and the first record of a year‐long high‐frequency pH time series for nearshore Alaska. SeaFET™ pH and O2 sensors deployed in Jakolof Bay and Bear Cove reveal a seasonally dynamic system in which nearshore waters in these two enclosed bays transition to being predominantly net autotrophic systems for a period of 60‐plus days. High rates and durations of primary production in late spring and early summer create high pH conditions and extreme variability. Observed pH values in Jakolof Bay and Bear Cove tracked hourly rates of change on the order of 0.18 and 0.10 units, respectively. In Jakolof Bay nondirectional variability within a 12‐h period was > 1 pH unit, exposing organisms to unstable, nonstatic pH conditions on tidal and diurnal cycles. Consistent frequency patterns detailing the magnitude of pH variability was correlated to tidal and O2 signatures, elucidating the dynamics and drivers of pH variability. This first year of observations is the first step in quantifying the anthropogenic contribution to acidification for Kachemak Bay in the forthcoming years.

Continue reading ‘Seasonality and biological forcing modify the diel frequency of nearshore pH extremes in a subarctic Alaskan estuary’

Mind your methods: acidification degrades total nitrogen and stable isotopic values within calcified marine macroalgae

Nitrogen and carbon are commonly used to determine nutrient regimes and trophic structures within marine ecosystems. Macroalgae are convenient for assessing nutrient conditions via stable isotopes and tissue nutrient levels because of their ability to absorb and integrate ambient nutrients over extended time periods. Calcified macroalgae, such as Halimeda and Udotea spp, are common constituents of tropical marine ecosystems, making them ideal candidates for nutrient-based and food web analyses. However, calcified genera require acidification to remove calcium carbonate to accurately determine δ13C and percentage of N (by weight); the overall effect of acidification on the tissue nutrients and stable isotopes of calcified genera is unresolved. Individuals of Halimeda kanaloana (n = 10) and Udotea geppiorum (n = 9) were collected from Maui, O‘ahu, and Lāna‘i. Each specimen was split into two samples and either decalcified using liquid-phase HCl (acidified) or left unaltered (control). We found that liquid-phase HCl acidification resulted in significantly lower percentage of N in both Halimeda kanaloana and Udotea geppiorum. Whereas δ13C values in acidified samples of both species were predictably lowered, the δ15N in acidified U. geppiorum was significantly increased. Acidification may have unpredictable consequences on both the percentage of nutrients in calcified algal tissue and their δ15N, suggesting that the use of acidification in calcified algal nutrient studies may produce erroneous conclusions. Analysing two sets of samples as calcified (for δ15N) and acidified (for δ13C) would eliminate these errors. However, the use of calcified macroalgae to assess percentage of N should be avoided.

Continue reading ‘Mind your methods: acidification degrades total nitrogen and stable isotopic values within calcified marine macroalgae’

Coastal Ocean Data Analysis Product in North America (CODAP-NA) – An internally consistent data product for discrete inorganic carbon, oxygen, and nutrients on the U.S. North American ocean margins

Internally-consistent, quality-controlled data products play a very important role in promoting regional to global research efforts to understand societal vulnerabilities to ocean acidification (OA). However, there are currently no such data products for the coastal ocean where most of the OA-susceptible commercial and recreational fisheries and aquaculture industries are located. In this collaborative effort, we compiled, quality controlled (QC), and synthesized two decades of discrete measurements of inorganic carbon system parameters, oxygen, and nutrient chemistry data from the U.S. North American continental shelves, to generate a data product called the Coastal Ocean Data Analysis Product for North America (CODAP-NA). There are few deep-water (> 1500 m) sampling locations in the current data product. As a result, cross-over analyses, which rely on comparisons between measurements on different cruises in the stable deep ocean, could not form the basis for cruise-to-cruise adjustments. For this reason, care was taken in the selection of data sets to include in this initial release of CODAP-NA, and only data sets from laboratories with known quality assurance practices were included. New consistency checks and outlier detections were used to QC the data. Future releases of this CODAP-NA product will use this core data product as the basis for secondary QC. We worked closely with the investigators who collected and measured these data during the QC process. This version of the CODAP-NA is comprised of 3,292 oceanographic profiles from 61 research cruises covering all continental shelves of North America, from Alaska to Mexico in the west and from Canada to the Caribbean in the east. Data for 14 variables (temperature; salinity; dissolved oxygen concentration; dissolved inorganic carbon concentration; total alkalinity; pH on the Total Scale; carbonate ion concentration; fugacity of carbon dioxide; and concentrations of silicate, phosphate, nitrate, nitrite, nitrate plus nitrite, and ammonium) have been subjected to extensive QC. CODAP-NA is available as a merged data product (Excel, CSV, MATLAB, and NetCDF, (Jiang et al., 2020). The original cruise data have also been updated with data providers’ consent and summarized in a table with links to NOAA’s National Centers for Environmental Information (NCEI) archives (

Continue reading ‘Coastal Ocean Data Analysis Product in North America (CODAP-NA) – An internally consistent data product for discrete inorganic carbon, oxygen, and nutrients on the U.S. North American ocean margins’

Coccolithophore calcification studied by single-cell impedance cytometry: towards single-cell PIC:POC measurements

Since the industrial revolution 30% of the anthropogenic CO2 is absorbed by oceans, resulting in ocean acidification, which is a threat to calcifying algae. As a result, there has been profound interest in the study of calcifying algae, because of their important role in the global carbon cycle. The coccolithophore Emiliania huxleyi is considered to be globally the most dominant calcifying algal species, which creates a unique exoskeleton from inorganic calcium carbonate platelets. The PIC (particulate inorganic carbon): POC (particulate organic carbon) ratio describes the relative amount of inorganic carbon in the algae and is a critical parameter in the ocean carbon cycle.

In this research we explore the use of microfluidic single-cell impedance spectroscopy in the field of calcifying algae. Microfluidic impedance spectroscopy enables us to characterize single-cell electrical properties in a noninvasive and label-free way. We use the ratio of the impedance at high frequency vs. low frequency, known as opacity, to discriminate between calcified coccolithophores and coccolithophores with a calcite exoskeleton dissolved by acidification (decalcified).

We have demonstrated that using opacity we can discriminate between calcified and decalcified coccolithophores with an accuracy of 94.1%. We have observed a correlation between the measured opacity and the cell height in the channel, which is supported by FEM simulations. The difference in cell density between calcified and decalcified cells can explain the difference in cell height and therefore the measured opacity.

Continue reading ‘Coccolithophore calcification studied by single-cell impedance cytometry: towards single-cell PIC:POC measurements’

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Ocean acidification in the IPCC AR5 WG II

OUP book