Chlorophyll in Water

What is Chlorophyll?

Chlorophyll, in various forms, is a pigment bound within the living cells of algae and other phytoplankton found in surface water. Chlorophyll assists in transferring energy during photosynthesis—the process by which sunlight is converted into energy and oxygen—by facilitating the reduction of carbon dioxide by water.

Chlorophyll a is the most abundant form of chlorophyll within photosynthetic organisms and, for the most part, gives plants their green color. While chlorophyll a is most prominent, other forms (b, c, and d) contribute to the overall fluorescence signal used in chlorophyll measurement. These various forms allow different species to absorb sunlight across different wavelengths.

Why Measure Chlorophyll?

Measuring chlorophyll in surface water provides valuable insights into the abundance and distribution of microscopic plant matter, commonly referred to as phytoplankton or algae. These measurements are crucial for assessing the health, composition, and ecological status of water bodies. Since chlorophyll is a key component of photosynthesis, its presence in water serves as a direct indicator of algal growth, making chlorophyll monitoring a practical way to track shifts in aquatic ecosystems.

Algae and phytoplankton are often referred to interchangeably, but they have distinct meanings. Algae include simple aquatic organisms, such as seaweed and pond scum, which contain chlorophyll. Phytoplankton, a subset of algae, are suspended microorganisms that perform photosynthesis. Monitoring chlorophyll within phytoplankton provides researchers and water managers with essential data on water quality, as these organisms are highly responsive to changes in nutrient levels.

In nutrient-rich waters—generally high in phosphorus and nitrogen—chlorophyll levels tend to be elevated. These nutrients encourage algal blooms, which can deplete dissolved oxygen as the algae die off and decompose—a primary factor in fish kills. High nutrient levels are often linked to pollution from sources like septic systems, wastewater treatment plants, and agricultural runoff. Therefore, chlorophyll measurement is also an effective, indirect indicator of nutrient pollution.

Key Applications for Chlorophyll Monitoring:

• Rivers and Streams: Monitoring chlorophyll in rivers and streams helps identify nutrient-driven algal growth, which can lead to eutrophication. Excessive algae can reduce oxygen levels, impacting fish and other aquatic organisms and sometimes resulting in fish kills.
• Lakes and Reservoirs: Chlorophyll measurement in lakes and reservoirs aids in characterizing the water body’s ecological health. High chlorophyll levels can indicate nutrient pollution from runoff, while regular monitoring helps manage algae levels, safeguarding water quality and recreational use.
• Ponds and Small Bodies of Water: Excessive algal growth can quickly disrupt the ecosystem in smaller, contained water bodies. Chlorophyll data helps managers control algae populations, maintaining a balanced environment for fish and plant life.
• Drinking Water Reservoirs: Chlorophyll monitoring in drinking water reservoirs is essential for identifying nuisance algal blooms early. Elevated chlorophyll levels can indicate blooms that may clog filtration systems or affect taste and odor, making early detection a priority for treatment plants.
• Ocean and Coastal Waters: Chlorophyll measurement in ocean and coastal waters helps track phytoplankton distribution and predict harmful algal blooms. These blooms reduce dissolved oxygen levels and can create hypoxic zones, endangering marine life and impacting fisheries.

In summary, chlorophyll monitoring is a practical, reliable method for assessing water quality, revealing nutrient pollution, and preventing ecological disturbances caused by algal blooms. This makes it an essential practice for environmental management and water resource protection.

How is Chlorophyll Measured?

There are various techniques to measure chlorophyll, including spectrophotometry, high-performance liquid chromatography (HPLC), and fluorometry. All of these methods are published in Standard Methods for the Examination of Water and Wastewater.

Spectrophotometry is the classical method of determining the quantity of chlorophyll in surface water. It involves collecting and filtering a water sample, rupturing the collected cells, extracting chlorophyll, and analyzing the extract via spectrophotometry or HPLC.

This general method, detailed in Section 10200 H. of Standard Methods, has been shown to be accurate in multiple tests and applications and is the procedure generally accepted for reporting in scientific literature.

The fluorometric method also requires the same extraction methods used with spectrophotometry, then uses a fluorometer to measure discrete molecular chlorophyll fluorescence.

However, these spectrophotometric methods have significant disadvantages. They are time-consuming and require an experienced, efficient analyst to generate consistently accurate and reproducible results. In addition, they do not lend themselves readily to continuous monitoring of chlorophyll (and thus phytoplankton).

YSI has developed optical sensors for measuring chlorophyll in spot sampling and continuous monitoring applications. Built for the field, our rugged and portable sensors determine chlorophyll in situ without disrupting the cells, allowing for the collection of large quantities of chlorophyll data.

In situ analysis with optical sensors will not be as accurate as spectrophotometry. Some sources of inaccuracy can be minimized by combining extractive analysis of a few samples during a study with the optical sensor data. Still, it is not possible for optical sensors to be as accurate as the spectrophotometric methods.

It is also important to consider that optical sensors are sensitive to all chlorophyll present in the environmental water sample, including that within living and nonviable cells and free-floating chlorophyll released by lysed cells. For this reason, optical chlorophyll measurements may not be directly representative of algal biomass.

Despite these limitations, optical in situ sensors are ideal for algal growth trend analysis, for visualizing seasonal and event-based fluctuations, and for understanding the effects of different inputs on the abundance of algae on a fine temporal scale.

How YSI Sensors Measure Chlorophyll

One key characteristic of chlorophyll is that it fluoresces. When irradiated with light of a particular wavelength, it emits light of a higher wavelength (or lower energy). The ability of chlorophyll to fluoresce is the basis for all commercial fluorometers capable of measuring the analyte in situ. These instruments induce chlorophyll to fluoresce by shining a beam of light of the proper wavelength into the sample and then measuring the higher wavelength light emitted due to the fluorescence process.

Most chlorophyll systems use a light-emitting diode (LED) as the source of the irradiating light; it has a peak wavelength of approximately 470 nm. LEDs with this specification produce radiation in the blue region of the visible spectrum. On irradiation with this blue light, chlorophyll emits light in the 650-700 nm region of the spectrum. To quantify the fluorescence, the system detector is usually a photodiode of high sensitivity that is screened by an optical filter that restricts the detected light. The filter prevents the 470 nm exciting light from being detected when it is backscattered off particles in the water. Without the filter, turbid (cloudy) water would appear to contain fluorescent phytoplankton, even though none were present.

Most commercial fluorometers fit into two categories:

• Benchtop instruments generally have superior optical flexibility and capability. Still, they are relatively expensive and can be difficult to use in the field.
• The less expensive field-type fluorometers with a fixed optical configuration can be more easily used in the field and are usually compatible with data collection platforms.

Using a pump is generally recommended for benchtop and field-type fluorometers, which can require large-capacity batteries for field use. These types of fluorometers have other disadvantages. For example, they can only give chlorophyll (fluorescence) readings and cannot measure any other water quality parameter. Also, the output is in mV or fluorescence units, not μg/L.

YSI’s unique Total Algae sensors consist of a probe similar in concept to the field-type fluorometers but much smaller, making them compatible with the probe ports of the EXO and ProDSS. These instrument platforms offer the ability to measure multiple water quality parameters alongside chlorophyll measurements—such as dissolved oxygen, temperature, pH, and more—for manual, discrete sampling or unattended, continuous monitoring. EXO and ProDSS Total Algae sensors are also two sensors in one, combining chlorophyll measurements with either phycocyanin or phycoerythrin, pigments common in cyanobacteria, using dual-channel LED technology.

The output of the Total Algae sensor is automatically processed via the instrument software to provide readings in relative fluorescence units (RFU) or µg/L of chlorophyll. A more quantitative output of cells/mL can also be achieved by generating a correlation coefficient in Kor Software using laboratory-analyzed grab samples. In addition, no pump is required, allowing the sensor to operate from either the sonde internal batteries or the batteries in a display/logger.

To learn more, check out our blog on How Algae Sensors Work | Answers to Four Challenging Questions.

Fouling Effects on Optical Measurements

Field optical measurements are particularly susceptible to fouling. This can occur from the long-term buildup of biological and chemical debris and/or the short-term formation of bubbles from the outgassing of the environmental water. In short-term sampling applications, these bubbles can generally be removed by simply agitating the probe manually.

A mechanical wiper—such as the Central Wiper for the EXO—is ideal for unattended applications. The wiper can be activated in real time during discrete sampling operations or function automatically during long-term unattended monitoring studies. The number of wiper movements and the cleaning cycle frequency for the unattended mode can be set in the EXO Sonde software. Generally, one wipe cycle per measurement is sufficient for most applications, but more frequent cleaning cycles may be necessary in environments with particularly heavy fouling.

Do you have questions about chlorophyll or need help selecting a measurement solution? Ask our experts or schedule a free virtual consultation today!