11 May 2023
Current position while writing (diamond shape). The circles are the positions at previous blog posts, the black line is roughly the route we took (base map from 
ggplot2).
After explaining what I measure a few weeks ago, I now want to explain how we measure. To keep this shorter, I’ll only explain pH and alkalinity. I am most familiar, with those, as we are working on pH measurements in the SapHTies project and the alkalinity system is open enough to partially see what is going on. Additionally, I did semi-automatic alkalinity titrations in Kiel in January.
pH
The pH value (or more precisely the pHT value) of seawater is measured spectrophotometrically most of the time. “Spectrophotometric” sounds quite fancy but just means that you use a pH indicator dye and look at the colour. pH indicators change their colour according to the pH value of a solution. Litmus paper and universal indicator are probably best known. The paper can be put into the solution directly or some drops placed onto it. It then changes its colour. The final colour can then be compared to the colour scale to determine the pH value. Liquid universal indicator is used very similarly, a few drops of the indicator are added to a sample and the colour is then compared to the colour scale.
Spectrophotometric measurements work in the same way. We use a different indicator for seawater but the concept is exactly the same: indicator is added to a sample and the resulting colour is compared to a known colour scale.
The indicator used for seawater is meta-Cresol Purple (often abbreviated as mCP). This is very well suited for seawater because the pH value at which its colour change occurs is close to the pH value of seawater (about pH 8). As the pH value of seawater usually varies in a relatively small range, we therefore only need to measure in this range and mCP is perfect for this..
Interesting to know: Why does a pH indicator change its colour?
If you think about it, it’s fascinating that something changes its colour just because the pH value is different. The pH value is the hydrogen ion concentration, so the indicator has to interact with this somehow. In fact, the indicator itself is an acid-base system. This means its molecules can accept or release a hydrogen ion. Indicators are weak acids which means that often only a part of the molecules has released its hydrogen ion while the other parts kept it. (This is a dynamic equilibrium and is related to the probability of each molecule releasing the ion. It’s not the case that some molecules keep better hold of their ion). The percentage of molecules with and without the hydrogen ion determines the colour.
What’s happening? The colours here originate from delocalised electrons in ring structures of the molecules. These structures interact with the light which changes its colour (= wavelength). We have an organic molecule with a ring structure as our indicator. In this ring structure, we get “clouds of electrons” which interact with the light and produce a colour. Now, if the molecule releases a hydrogen ion, the ring structure changes. For example, the number and position of double bounds changes. This, in turn, changes the electron cloud which then interacts differently with the light producing a different colour.
Each indicator has a specific pH range in which the colour without hydrogen a ion turns into the colour with a hydrogen ion. If a molecule can accept two or even three hydrogen ions, several colour changes occur. These points of colour change are where the indicators are used. For example, in a titration, an indicator is chosen that changes its colour in the range of interest. Then acid is added until the colour change occurs (the pH value of interest is reached), and the original concentration can be calculated.
An indicator you might know from everyday life is red cabbage. Depending on the pH value it is either blue-ish or red/purple.
This was only a very rough overview. Some more information on indicators can be found, for example, on Wikipedia: https://en.wikipedia.org/wiki/Indikator_(Chemie)
Well, let's go on:
To summarise, we take our sample, add mCP and compare the colour to a colour scale. In reality, we don’t compare the colours visually but measure them. The measured value is then compared to known values and the pH is determined. This makes it much easier than sitting in front of samples and trying to see a minimal difference in colour … and it’s faster and more precise. I will link to more detailed measurement and calculation descriptions at the end of the entry.
In addition to the measurement needed to determine the colour, a few other measurements are taken to reduce different influences on the measurement. In the beginning, the sample is measured without an indicator. This allows us to correct the measurement for this specific sample. After the first addition of the indicator, often a second addition and measurement are done. As the indicator itself is an acid, it affects the pH value when added to the sample. The second measurement is there to determine this effect and correct it.
If it was working, this lab-on-chip sensor would measure pH spectrophotometrically … (Picture: Rieke Schäfer)
Even so, the details are quite complex, I hope the concept is clear: take a sample, add an indicator, measure colour, and look up the pH value. This raises the question of who came up with the colour scales for the indicator. It can’t be something someone comes up with randomly and has to be based on something. Of course, this is not only true for mCP used in oceanography but for all pH indicators. pH-colour scales do not come out of nowhere.
And exactly this, to determine the colour scale for mCP more precisely is one of the goals of the SapHTies project which my work is part of. So, what do we do? As part of the project we at PTB, the LNE (France) and IPQ (Portugal) measure the pHT values of buffers with different salinities. Buffers are solutions that have a stable pH value (at least while external influence remains moderate). For seawater experiments, usually TRIS is used as the buffer. We measure different salinities because they influence the pH value and in the end, we want results for a variety of conditions. To make sure we know the salinities precisely, we don’t use real seawater but mix our own. For this artificial seawater, an ion composition very similar to real seawater is used. Some alterations are however made due to experimental constraints. The measurement method is based on voltage differences in Harned cells. Harned cells are quite complex glass instruments in which the pH value of buffers can be measured (more details in the links).
This is a Harned cell. (Picture: Ralf Eberhardt)
Now we have the pH value of our solutions with different salinities (and different temperatures). These same solutions are then used by GEOMAR to do spectrophotometric measurements as described above. The only difference to the previous description is that we already know the pH value and can therefore match the measured colour with this pH value. With several measurements, the colour scale can be created (and traceability ensured). Of course, there is already a colour scale for mCP otherwise it couldn’t be used as an indicator. The goal of the SapHTies project is, in addition to clarifying some details related to the buffers, to determine the measurement uncertainties and reduce them.
Now let’s get back to the seawater pH measurements. Spectrophotometry is not the only method to measure pH, several other methods are available and there still is active development of new methods. You might know pH glass electrodes. They look like a pen and are placed into the sample for a while before the display shows the pH value. This does sound nice and easy, but unfortunately, glass electrodes have a couple of disadvantages. They are fragile, require frequent calibration and are not well suited for seawater. Their big advantage compared to spectrophotometric measurements is that they do not require any chemicals. In recent years, a number of alternatives have been developed that work with different methods to determine pH. For example, the pH sensor I am using here (the one that is working), measures pH with an optode. In this method, an indicator in solid form is activated with red light and its luminescence in near-infrared is measured (concept on the manufacturer's page). We are also trying to get another sensor to work that measures pH based on redox potentials. There is a number of different methods, like ISFET and many others. Each method has its advantages and disadvantages. With an increasing number of autonomous observation platforms, no or low consumption of chemicals, seldom calibration and biofouling resistance are often arguments for a method.
Pyroscience pH sensor which measures pH by luminescence. The measurement happens in the blag cap. With a pump, seawater is continuously pumped into the cap, the sensor measures every minute, and the water leaves the cap through the second tube. (Picture: Rieke Schäfer)
Sensing cap of Pyroscience pH sensor (with USB stick for size comparison). There really is not much to see. The measurement reaction happens at the small, black cylinder in the green „cage“. The cap has to be replaced from time to time. (Pictures: Rieke Schäfer)
Alkalinity
Now we continue with the measurement of alkalinity. Don’t worry, this section will be much shorter. As the details of alkalinity are much more complex and I haven’t worked with it as much as with pH, I’ll mostly give a rough overview of which concepts are used to measure it.
The alkalinity of a sample is usually determined by titration. As mentioned previously, alkalinity describes how much the addition of acid affects the pH value. In the titration, acid is added and it is determined how much acid is needed to reach a certain pH value. This can be done, of course, manually but there are also semi-automatic and automatic systems which add the acid and do the measurement.
On this cruise, we have two alkalinity systems. I am in charge of a system that can measure continuously and that measures by adding a certain amount of indicator-acid-mix to the sample. The pH value of the mixture is then measured spectrophotometrically (this is why the indicator is added) and the alkalinity is calculated from that. Overall, somewhat similar to a titration, however, the amount of acid added is always the same.
I already mentioned previously that using the alkalinity system is not always easy. For example, a few days ago air bubbles got into the tubes. When I noticed this in the evening, I checked everything for the reason but could not really find it. As it seemed to get better, I postponed a more detailed search to the next morning. Over night, a lot more air got in but at least in the morning, I was able to find the problem. After exchanging the tube for a new one I waited for all the air to be flushed out and the system to stabilise again. It seemed fine again after a while, but then suddenly the values were too low. So I checked again everything for bubbles and any other potential problems but could not find anything. After five values that were too low, everything worked fine again. However, as I did not find the reason for the wrong values I was worried that it might not be actually solved, so I am checking the values more often now. Luckily, it seems to be working fine now.
Our second alkalinity measurement system is doing a titration. It is used for samples from the CTD. This system too, is making problems quite often. During the transport, one glass piece broke and after solving that, tube connections start to leak from time to time. And the computer program that runs the system is very particular with which buttons you are “allowed” to push without it crashing.
By the way, the comparability of the alkalinity measurements is ensured through reference solutions. They are usually measured after the calibration and then repeated throughout the measurement time if needed. For the system, I am taking care of, we are also measuring a “standard” roughly once a day to correct for drift and other influences. In this case, the standard is just seawater with mercury chloride to fix its alkalinity. Alkalinity is not influenced by CO2 exchange with the air (at least short term) which makes keeping the standard much easier. Mercury chloride kills microorganisms that might change the alkalinity.
My task(s)
This was a lot of oceanography and you might wonder where the metrology comes in. In fact, the measurements are normal routine measurements in oceanography. What we are interested in from a metrological point of view is how these measurements are done under real conditions and which influences and constraints occur. In the office, we can theorise a lot about measurement uncertainties. The real conditions are however quite different. So on this cruise, I have the opportunity to do the measurements myself, talk to the people and understand how things are done (or not) from a metrological point of view.
Originally, we also wanted to look at the drift of the spectrophotometric pH sensor. As mentioned, I have TRIS buffers with me that were intended to regularly check the sensor. Unfortunately, the sensor does not want to work at all so we had to adapt our plan. Now, I am measuring the buffer with the other pH sensor. As this one needs a lot more solution for each measurement, I can’t do measurements as often as planned but some data points are better than none. We have to make it work with the things we got here. We’ll see what these measurements tell us. At the moment, I am happy to have the TRIS buffer as I don’t quite trust the results the sensor is giving us … We know the pH of the buffer very well, so we’ll at least be able to tell how wrong the sensor is in that case.
I hope this gave a rough overview of how pH and alkalinity are measured in oceanography, which problems might occur and what I am doing here.
Sunset to relax after all that lab work:
As promised, some links on topics I mentioned:
Seawater pH measurements at PTB
Overview on spectrophotometric pH measurements of seawater
Explanations how indicators are used from Khan Academy (assumes some background knowledge which is explained in the previous videos)
Two publications from a previous project that also determined the color scale for mCP:
Description of primary pH measurements at PTB (pHT is measured and calculated slighlty differently but this gives an overview)
Overview on seawater pH measurements from NIST
Detailed (and somewhat complex) overview on alkalinity
PTB doctoral student 
Rieke Schäfer is blogging here directly from the RV "Sonne" on her way west from South America across the Pacific Ocean.
