• What are phytoplankton?
• How do they photosynthesize?
• How do we measure photosynthesis in the sea?

Overview

In our discussion of light in the sea, we discovered that the growth and dynamics of phytoplankton radically influence the color of the oceans and affect the depth of light penetration through the water column. We now take a closer look at these "grasses" of the sea and discuss the major groups that comprise their ranks. No discussion of phytoplankton would be complete without paying homage to the means by which they assimilate carbon and synthesize organic compounds using light energy, arguably the most important process on earth, photosynthesis.

What are phytoplankton?

Phytoplankton have been called the grasses of the sea. Through the process of photosynthesis (photo=light; synthesis=build), these microscopic, single-celled plants nourish the entire food web of the oceans. The lives of all animals that live in the sea -- with the exception of hydrothermal vent organisms -- depend on phytoplankton for energy and minerals. The global carbon cycle, which regulates the temperature of our planet, and life-sustaining oxygen, essential to the metabolism of all aerobic organisms, are controlled by the actions of the phytoplankton. Perhaps no other group of organisms plays such a major role in the maintenance of life on Earth.

Collectively, phytoplankton are known as microalgae, a designation given to all algae, of which phytoplankton are a part, that are too small to be seen with the naked eye. The other group of algae that inhabit the seas are the macroalgae, the kelps and fucoids that grow along the shore. We'll study macroalgae in more detail when we examine rocky shores.

The largest phytoplankton include the diatoms, the coccolithophorids, and the dinoflagellates. The smallest phytoplankton, the ultraphytoplankton (which includes picophytoplankton), include a single-cell planktonic form of the blue-green algae known as the cyanobacteria. This latter group was not really appreciated until the mid-1970s when it was "discovered" that this group of phytoplankton may contribute up to 70% of the world ocean's entire primary production. At a concentration of approximately 100,000 cells per milliliter (about a drop of water), this group of fascinating photosynthetic bacteria has received much attention in recent years. Despite their "bacterial" nature, they are eukaryotic and they do contain chlorophyll. For that reason, they are included among the algae.

Phytoplankton also come in many different shapes. With the naked eye, they appear as tiny specks floating in the water. Under a microscope, however, an incredible variety of bizarre and beautiful forms make themselves known: opalescent ovals punctuated with thousands of tiny holes, pill-box-like chains with protruding spines, plated-and-grooved "spaceships" with flickering flagella. These are just a few of the myriad forms that greet an interested visitor to their world.

Three principal characteristics distinguish the major groups of phytoplankton from each other. These are:

1. size and shape
2. photosynthetic pigments
3. cell wall composition

We've already talked about size. Let's examine a few of the other differences for a moment.

The diatoms are perhaps the most well-known and beautiful of all the phytoplankton. They typically inhabit cold, nutrient-rich water, but, in fact, they are found just about everywhere. Benthic forms (cells that live attached to the bottom) are common in rivers, lakes, and even the seashore. Their characteristic yellow-brown color is caused by fucoxanthin, an accessory pigment. Accessory pigments assist chlorophyll in catching radiant energy by having absorption bands at different wavelengths than chlorophyll. In this way, changes in the spectral distribution of light can be compensated by making more accessory pigments. This ability of diatoms and other phytoplankton to change the amount and type of pigments in response to changes in the intensity and spectral distribution of light is called photoadaptation.

Diatoms come in two major shapes: centric and pennate. Centric diatoms are radially symmetrical, appearing round like pillboxes or baskets. Pennate diatoms have bilateral symmetry, appearing slender and curved like rods. Because of their shape, centric diatoms float better than pennate diatoms and, thus, centric diatoms tend to be strictly planktonic while pennate diatoms tend to be primarily benthic.

The cell wall of diatoms is known as a frustule. The frustule is composed of pectin, a jelly-like carbohydrate substance, and of course, silica. It is the siliceous part of the frustule that gives diatoms their characteristic beauty. Every diatom frustule is split into two pill-box-like valves, an upper valve and a lower valve. The upper valve overlaps with the lower valve giving the frustule a box-like appearance. This pill-box-like arrangement makes asexual reproduction somewhat of a bother. The two valves split and a new lower half is formed. This means that the original lower valve now becomes the upper valve (of the daughter cell) and that the size of the diatom is reduced. If this process continued, the diatom would shrink out of existence. However, when a certain minimum size is reached, diatoms form a reproductive spore, known as an auxospore. This spore gives rise to a full-size diatom and the process starts all over again.

The other major group of phytoplankton that we have studied are the coccolithophorids. Coccolithophorids have cell walls made of calcium carbonate, which is often built into elaborate "life-saver-like" plates called coccoliths. Sometimes, these coccoliths are raised on stems, making them appear like a conglomeration of trumpets. When present in large numbers, i.e. when there is a coccolithophorid bloom, the water may actually turn milky white as a result of the large number of coccoliths. The white cliffs of Dover are largely composed of the remains of coccolithophorids.

The other group of phytoplankton we must mention are the dinoflagellates. These bizarre armored denizens of the lighted depths must surely be among the most remarkable organisms on Earth. Equipped with a battery of talents, there is just about nothing that these organisms don't seem to be able to do. Consider: they photosynthesize; they eat other organisms; they can assimilate dissolved organic substances; they migrate up and down the water column; they have biorhythms; they bioluminesce; they produce toxic substances that prevent them from being eaten. These many characteristics must surely make them one of the more successful organisms. If they had any drawback, it would be that they are slow-growing.

Dinoflagellates have cell walls that come in two varieties: naked or armored. Naked dinoflagellates, such as the bioluminescent Noctiluca, have smooth and flexible cells walls. Other dinoflagellates, such as the red-tide-causing Gonyaulax, has a cell wall that is composed of cellulose plates, given it an armored or helmeted appearance. Naked or armored, most dinoflagellates have two flagella, whiplike structures that provide locomotion, that rest within grooves, one groove rounding the periphery of the cell and the other groove running the perpendicular along the length of the cell. This arrangement of the flagella allows them to spin and move forward (or backwards, who can tell?).

The reddish color of dinoflagellates, most prominent when they bloom profusely in a phenomenon known as red tides, is caused by an accessory pigment known as peridinin. This peridinin, like fucoxanthin, has absorption bands in the 500-560 nanometer range. It functions to capture the green wavelengths of light when blue wavelengths are less available (i.e. when phytoplankton are so numerous that they absorb most of the blue light).

How do they photosynthesize?

The principal "ingredient" that enables phytoplankton and all photosynthetic organisms to use light energy is chlorophyll. This amazing molecule captures photons from the sun and transfers them down a chain of electron-transfer components that assist in the manufacture of energy in the form of ATP that can be used to synthesize cellular components from carbon dioxide. In the process, electrons are "stolen" from water, resulting in the production of oxygen as a byproduct. It is worth our while to briefly examine this process of photosynthesis and to understand how light capture results in the uptake of carbon dioxide and the synthesis of organic compounds.

The basic equation that describes photosynthesis can be written as follows:

light energy

6CO2 + 6H20 ------------------ C6H12O6 + 6O2

chlorophyll

or, 6 molecules of carbon dioxide plus 6 molecules of water in the presence of light energy and chlorophyll makes one molecule of sugar (glucose) and 6 molecules of oxygen. Note that inorganic molecules, namely carbon dioxide and water are used to "create" organic molecules. It is this interface between the nonliving and the living world, this creation of living matter from nonliving matter, governed by the spark of light, that so fascinates me. In the grasp of a plant, that which was once dead is now living. The transfer of energy and mass from the physical domain to the biological domain is one of the most intriguing processes on this planet. Without it, life does not exist. Through this alleyway is derived all the organic matter and energy on which we and other animals depend. Now that's a large responsibility!

The reactions symbolized above constitute include both the light and dark reactions of photosynthesis. As mentioned above, chlorophyll captures the energy of the sun and transfers it down an electron transfer chain that results in the synthesis of ATP and NADPH2, which are energy-carrying molecules. These are the light reactions. In the dark reactions, ATP and NADPH provide energy to break apart carbon dioxide and manufacture glucose.

Our discussion of photosynthesis here will be limited to the light reactions, which are involved in the transfer of light energy from the sea to the phytoplankton. Within each phytoplankton cell, the light-capturing molecule chlorophyll is housed in a special organelle called a chloroplast. On the surface of the chloroplasts, chlorophyll molecules extend like tiny umbrellas to capture photons thrown at them by the sun. These "umbrellas" are called reaction centers and they involve a couple different kinds of chlorophyll and all the accessory pigments we talked about earlier. Two types of reaction centers are important in the light reactions of photosynthesis. They are called photosystem I and photosystem II.

Now I don't expect you to memorize all this, nor do I even expect you to completely (or even remotely) understand it, but I think you will find it fascinating, as I have, so I'm going to take a shot at explaining it to you here. Let's start with a description of the photosystems. At the center of each photosystem is a "mother" chlorophyll molecule. You may think of it as the handle of the umbrella. Surrounding this "reaction center molecule" are a 50 - 300 other chlorophyll molecules. Linked to these chlorophyll molecules (perhaps interspersed within them) are the accessory pigments, which may be thought of as different color umbrellas trained at catching different wavelengths of light. For every photosystem I reaction center there is a photosystem II reaction center; thus, they work together to perform the light reactions in photosynthesis.

Photosystem II, stretched like an umbrella across the chloroplast of a phytoplankton cell, is the larger of the photosystems. Most of the accessory pigments "feed" their photons towards the photosystem II "mother" chlorophyll, known as P680. As shown in the diagram handed out in class, photosystem II is responsible for breaking apart water, which liberates oxygen. This breaking apart of water also liberates a hydrogen. Thus, photosystem II performs the "water-splitting" reactions. To accomplish this feat, photosystem II must absorb 4 photon hits, the number required to split water. As a photon of light is absorbed by photosystem II, which includes all of the associated chlorophyll molecules and accessory pigments, the energy from this photon is sent to the reaction center, the P680. This photon hit on P680 and the splitting of water provides an electron that puts P680 in an excited state, often symbolized as P680*. This excited state allows P680* to transfer an electron through an electron transfer system to P700, the "mother" molecule of photosystem I.

Photosystem I performs the NADPH2-making reactions. The transfer of an electron from photosystem II to P700 sets up the photosystem I reactions. Another photon hit on photosystem I is required to transfer the electron from water to an electron transfer chain that produces NADPH2. In the process of transferring an electron through the photosystem I electron transfer chain, 1 to 2 molecules of ATP are produced.

Because 4 photons are required to split water, in photosystem II, and 4 photons are required to transfer that electron through photosystem I, a total of 8 photons are required to produce the energy need to manufacture one molecule of glucose in the dark reactions. This was a question for my qualifying exams in graduate school, so now all of you are trained for at least one question in graduate school (if you decided to pursue science). Feel honored!

How do we measure photosynthesis in the ocean?

The rate at which photosynthesis occurs in the ocean is termed gross primary productivity. Of course, phytoplankton cells must respire to grow, so while they are photosynthesizing, they may be respiring too. The difference between the gross primary production of phytoplankton and the respiration of phytoplankton is termed net primary productivity. In instances where phytoplankton are being eaten while they are photosynthesizing and respiring, the term net community productivity is used. This term better describes the sum of all the processes that contribute to the production of carbon compounds in the ocean.

Traditional measurements of primary productivity in the ocean have concentrated on two separate aspects of photosynthesis, namely, the uptake (or consumption) of carbon dioxide or the evolution (or release) of oxygen. Both of these processes take place as a result of photosynthesis and, thus, provide some indication of how fast plants are growing. The cornerstone of either of these methods is the light-dark bottle method. As shown in your book, transparent and dark bottles are filled with seawater samples, strung on a line, and incubated in seawater for a given amount of time. (Today, polycarbonate bottles are used, which are much cleaner; not glass, as described in your book.) The difference between values of oxygen (or carbon dioxide) at the beginning of the experiment and the end of the experiment (typically an increase if photosynthesis is occurring) in the light bottles gives a value for net primary production at each depth. The start-finish difference in oxygen (or carbon dioxide) values in the dark bottles give some idea of the amount of respiration.

Using the values for net primary productivity determined in the light bottles and respiration determined in the dark bottles, the gross primary productivity is calculated according to:

net primary production + respiration = gross primary production

Can you see how this must be true given the equations of photosynthesis? Only respiration, the opposite of photosynthesis, occurs in the dark bottle. Respiration produces carbon dioxide (we breathe out) and consumes oxygen. Thus, if we are to accurately determine the total amount of oxygen produced or carbon consumed solely as a result of photosynthesis, we must account for the respiration that occurs during the incubation period. Otherwise, we are only measuring net production. Gross primary production is calculated by adding the respiration values determined in the dark bottles to the net primary production values determined in the light bottles.

This light-dark bottle method may be used for productivity determinations based on oxygen or carbon. Typically, however, oxygen measurements are used only where big differences between the starting and finishing concentrations are expected, such as in lakes or coastal waters rich in phytoplankton. In the open ocean, more sensitive methods, such as the C-14 technique, must be used to obtain accurate measurements.

In this method, samples of seawater are typically collected before dawn, carefully placed in clean polycarbonate bottles, inoculated with radioactive "carbon dioxide" (actually, C-14 bicarbonate is added), and suspended at different depths in the euphotic zone of the water column. After a period of time, typically one day (sun-up to sun-down) but sometimes as short as 2 hours, the bottles are collected, the phytoplankton are filtered onto glass fiber filters, and the filters are placed in a liquid that emits a photon of light for every radioactive decay. Using a scintillation counter, an instrument that measures these flashes of light, oceanographers can determine how much carbon was "fixed" into the cells of phytoplankton during photosynthesis.

The carbon-14 technique is the most widely used method for determining primary productivity today. Moreover, the method of hanging bottles from a line (attached to a buoy) is preferred over other means (such as incubating bottles on deck under screens that simulate the amount of sunlight). This method of placing bottles on a line is known as in situ productivity measurements. In situ methods are preferred because they duplicate most closely the underwater light field experienced by phytoplankton. Both the intensity of light, which varies with depth and can be affected by clouds or waves, and the spectral distribution of light, which may be affected by other particles or phytoplankton suspended in the ocean, are reproduced using in situ methods.

However, the C-14 method is not without its problems. In the real ocean, phytoplankton do not live in a bottle. Potential artifacts are introduced as phytoplankton are collected, poured into a bottle, and thrown overboard on a line. Contamination of the sample, container effects (i.e. the presence of a surface on which bacteria may proliferate), rapid decline of sensitive organisms (such as small gelatinous zooplankton) and other potential problems throw into question whether the bottle incubation method gives a realistic number for phytoplankton production. In addition, the bottle method fails to duplicate mixing within the water column, a process that could have considerable effect on the chemical and light environments phytoplankton experience throughout the day.

For these reasons, oceanographers at the University of Southern California set out to find a non-intrusive method for measuring the photosynthetic rate of phytoplankton. Nonintrusive methods rely on measuring ocean properties as they exist, not by subjecting the ocean to any perturbations. These passive measurements work much like the tricorders on Star Trek. By pointing your instrument at a particular spot, you can determine certain properties of that spot, such as whether "signs of life" are present. Along those lines, you might say that the USC oceanographers were attempting to develop oceanic tricorders!

As we just discussed, a photon of light hitting a chlorophyll molecule puts it into an excited state. An excited chlorophyll molecule may transfer its energy either to another chlorophyll molecule (i.e. through the umbrella) or to an electron transfer chain. However, some of that energy gets lost along the way through a process known as fluorescence. As a chlorophyll molecule fluoresces, it loses energy at a less energetic wavelength, namely red light. Thus, chlorophyll molecules fluoresce red when stimulated by light, namely, blue light (since blue light is the color best absorbed by chlorophyll).

Oceanographers have used chlorophyll fluorescence for decades as an accurate and reliable means of measuring the concentrations of chlorophyll in seawater. This type of fluorescence measurement, developed by Carl Lorenzen at the University of Washington, artificially stimulates chlorophyll fluorescence using a high-intensity blue light and measures the fluorescence produced using a detector that only responds to red light. The oceanographers at USC wondered whether chlorophyll fluorescence stimulated by solar radiation in the water column, called natural, or solar-stimulated, fluorescence, could be used as a means of measuring rates of photosynthesis.

The first task was to determine whether natural fluorescence could be detected in the water column. You may remember that red light from the sun is removed in the surface waters of the oceans. Thus, any light detected at depth in the water column could only come from chlorophyll fluorescence. In the early 1980s, scientists measuring the underwater light spectrum noted abnormal amounts of red light at depth, an observation they attributed to faulty equipment. Using optical instruments specially developed to measure natural fluorescence, the oceanographers at USC determined that red light at depth was indeed from solar stimulation of chlorophyll. For the first time, the dream of measuring photosynthesis without putting phytoplankton in a bottle was becoming a reality.

The next task was to determine whether these measurements of natural fluorescence corresponded to measurements of natural fluorescence. If a direct relationship could be found between natural fluorescence and primary production, a much more rapid and non-intrusive means for measuring productivity would be established.

The first set of experiments took place in 1987 aboard the Calypso. By comparing traditional in situ measurements of primary production, as described above, with optical measurements of natural fluorescence, as developed by oceanographers at USC, the relationship between these two processes was established. Subsequent measurements in different oceans, including the North Atlantic and Antarctic Oceans further substantiated that natural fluorescence was a reliable means for measuring primary production. This work, as reported by Chamberlin et al. in 1989, set the stage for a much greater understanding of primary production in the sea.

A Few Final Thoughts

It is most difficult to restrain from expanding on the many fascinating aspects of photosynthetic research in the sea. Both optical and biological processes present challenges to our understanding of how phytoplankton grow and reproduce. Yet, if we are ever to gain an understanding of how man impacts the sea, if we are ever to determine the role of the oceans in the global carbon cycle, then we must strive to improve our methods to measure the sea. The vital role that phytoplankton play, both as moderators of our atmosphere and as the base of the oceanic food web, certainly speaks for our need to understand them. By educating students and the public as to the nature of these "grasses of the sea", perhaps our interest in studying them can be sustained.