Page 244 - Algae Anatomy, Biochemistry, and Biotechnology

Algal Culturing 227

reactions. Of these metals, the concentrations (or more accurately the biologically available con-
centrations) of Fe, Mn, Zn, Cu and Co (and sometimes Mo and Se) in natural waters may be limiting
to algal growth. Little is known about the complex relationships between chemical speciation of
metals and biological availability. It is thought that molecules that complex with metals (chelators)
influence the availability of these elements. Chelators act as trace metal buffers, maintaining con-
stant concentrations of free ionic metal. It is the free ionic metal, not the chelated metal, which
influences microalgae, either as a nutrient or as a toxin. Without proper chelation some metals
(such as Cu) are often present at toxic concentrations, and others (such as Fe) tend to precipitate
and become unavailable to phytoplankton. In natural seawater, dissolved organic molecules (gen-
erally present at concentrations of 1–10 mg l 21 ) act as chelators. The most widely used chelator in
culture media additions is EDTA, which must be present at high concentrations because most com-
plexes with Ca and Mg, present in large amounts in seawater. EDTA may have an additional benefit
of reducing precipitation during autoclaving. High concentrations have, however, occasionally
been reported to be toxic to microalgae. As an alternative the organic chelator citrate is sometimes
utilized, having the advantage of being less influenced by Ca and Mg. The ratio of chelator:metal in
culture medium ranges from 1:1 in f/2 to 10:1 in K medium. High ratios may result in metal
deficiencies for coastal phytoplankton (i.e., too much metal is complexed), and many media there-
fore use intermediate ratios.
In today’s aerobic ocean, iron is present in the oxidized form as various ferric hydroxides and
thus is rather insoluble in seawater. While concentrations of nitrogen, phosphorus, zinc, and manga-
nese in deep water are similar to plankton elemental composition, there is proportionally 20 times
less iron in deep water than is apparently needed, leading to the suggestion that iron may be the
ultimate geochemically limiting nutrient to phytoplankton in the ocean. Very little is known
about iron in seawater or phytoplankton uptake mechanisms due to the complex chemistry of
the element. Iron availability for microalgal uptake seems to be largely dependent on levels of che-
lation. It is highly recommended that iron be added as the chemically prepared chelated iron salt of
EDTA rather than as iron chloride or other iron salts; the formation of iron chelates is relatively
slow, and iron hydoxides will form first in seawater, leading to precipitation of much of the iron
in the culture medium.
Apparently, as a result of the extreme scarcity of copper in anaerobic waters, copper did not
begin to be utilized by organisms until the earth became aerobic and copper increased in abundance.
Consequently copper does not seem to be an obligate requirement, algae either not needing it, or
needing so little that free ionic copper concentrations in natural seawater are sufficient to maintain
maximum growth rates. Copper may indeed be toxic, particularly to more primitive algae, and
hence copper, if added to culture media at all, should be kept at low concentrations.
The provision of manganese, zinc, and cobalt in culture medium should not be problematical
because even fairly high concentrations are not thought to be toxic to algae.

Vitamins
Roughly all microalgal species tested have been shown to have a requirement for vitamin B 12 ,
which appears to be important in transferring methyl groups and methylating toxic elements
such as arsenic, mercury, tin, thallium, platinum, gold, and tellurium, around 20% need thiamine,
and less than 5% need biotin.
It is recommended that these vitamins are routinely added to seawater media. No other vitamins
have ever been demonstrated to be required by any photosynthetic microalgae.

Soil Extract
Soil extract has historically been an important component of culture media. It is prepared by
heating, boiling, or autoclaving a 5–30% slurry of soil in fresh water or seawater and subsequently
filtering out the soil. The solution provides macronutrients, micronutrients, vitamins, and trace