ASTM_D_3978_-_04_2012.pdf

Designation:D397804(Reapproved 2012)Standard Practice forAlgal Growth Potential Testing with Pseudokirchneriellasubcapitata1,2This standard is issued under the fixed designation D3978;the number immediately following the designation indicates the year oforiginal adoption or,in the case of revision,the year of last revision.A number in parentheses indicates the year of last reapproval.Asuperscript epsilon()indicates an editorial change since the last revision or reapproval.INTRODUCTIONAlgae are natural inhabitants of surface waters and are found in almost every water environmentthat is exposed to sunlight.The algae contribute to self purification(both organic and inorganic)ofstreams and lakes and are necessary as food for fish and fish food organisms.When large amounts ofnutrients are available,excessive growths referred to as“blooms”can occur.Some algal bloomsrelease substances toxic to fish,birds,domestic animals,and other alga.When nutrients are exhausted,the growth of algae and production of oxygen by photosynthesis decreases.The respiration of bacteriadecomposing the large quantity of algal cells can deplete dissolved oxygen to the extent that fish andother oxygen consumers die.Both the abundance and composition of algae are related to water quality,with algal growth primarily influenced by the availability of nutrients.The presence of indigenous algae in a water sample suggests that they are the most fit to survivein the environment from which the sample was taken.The indigenous algae should produce biomassuntil limited from further growth by some essential nutrient.If the indigenous algal production islimited from further growth by an essential nutrient,the laboratory test alga cultured in anoncompetitive environment and responding to the same limiting nutrient will produce parallelmaximum yield growth responses.Generally,indigenous phytoplankton bioassays are not necessaryunless there is strong evidence of the presence of long-term sublethal toxicants to which indigenouspopulations might have developed tolerance(1)3.A single-indigenous algal species,dominant at the time of sampling,may not be more indicative ofnatural conditions than a laboratory species that is not indigenous to the system.The dynamics ofnatural phytoplankton blooms,in which the dominant algal species changes throughout the growthseason,makes it quite certain that even if the indigenous algal isolate was dominant at the time ofcollection,many other species will dominate the standing crop as the season progresses.When comparing algal growth potentials from a number of widely different water sources there areadvantages in using a single species of alga.The alga to be used must be readily available and itsgrowth measured easily and accurately.It must also respond to growth substances uniformly.Becausesome algae are capable of concentrating certain nutrients in excess of their present need when they aregrown in media with surplus nutrients,this factor must be taken into account in selecting the culturemedia and in determining the type and amount of algae to use.(2)showed that a blue-green algaeMicrocystis aeruginosa,cultured in a low-nitrogen concentration medium,would not grow whentransferred to medium lacking nitrogen.However,when the alga was cultured in medium containingfour times as much nitrogen it was able to increase growth two-fold after transfer into nitrogen-freemedium.A green alga Pseudokirchnereilla subcapitata(formerly known as Selenastrumcapricornutum,gave a similar response.In an analogous experiment with phosphorus,both organismsincreased four-fold when transferred to medium lacking phosphorus.However,if algae are culturedin relatively dilute medium as recommended in the Algal Assay Procedure:Bottle Test(3)forculturing Pseudokirchnereilla subcapitata,disclosed no significant further growth in medium lackingnitrogen or phosphorus when these were transferred from the initial medium over a wide range ofinoculum sizes(4).There are several methods available for determining algal growth.Measurements of optical density,oxygen production,carbon dioxide uptake,microscopical cell counts,and gravimetric cell massdeterminations have been used,but often lack sensitivity when the number of cells is low.Measurement of the uptake of carbon-14 in the form of bicarbonate is a sensitive method but can alsoCopyright ASTM International,100 Barr Harbor Drive,PO Box C700,West Conshohocken,PA 19428-2959.United States1 be time-consuming.In vivo fluorescence of algal chlorophyll has been used with many types of algaeand has proved particularly useful with indigenous algae or filamentous forms not easily measured atlow concentrations by other methods.The method is sensitive and measurements can be quicklyperformed.However,chlorophyll to cell mass ratio may vary significantly with growth in watersamples of different chemical composition(5).The electronic particle counter has been used forcounting and sizing nonfilamentous unialgal species(6,7).Shi