General Purpose
Conceptual
Procedural
Many abiotic factors can be measured directly using basic scientific instruments (thermometer, pH meter, etc.). However, some abiotic and biotic factors require extraction or chemical reactions to be analyzed. In this pre-lab, three of the assays that require more complicated analysis will be explored. The dissolved oxygen test and the chemical testing are routinely done when investigating fish kills and when monitoring water quality.
Chlorophyll Analysis
Measuring chlorophyll is an indirect measurement of the potential for photosynthesis that can be done by the photoautotrophs in an ecosystem. Since chlorophyll is a pigment in the photosynthetic membranes of the organisms, it must be extracted from those membranes before it can be measured.
The extraction of membrane pigments like chlorophyll often requires the use of organic solvents, and any time you are using an organic solvent proper care should be used and proper procedures should be followed. After extraction of the pigment from the membrane, the amount of pigment is determined using the spectrophotometer.
Please view the following video to see some of the aspects of the chlorophyll analysis. This video does not have audio.
Dissolved Oxygen Test
Ecologists are interested in the productivity of a community as an estimate of the system’s health. Two important measures of productivity are gross primary production (GPP), the rate that a community converts light energy to biomass, and net primary production (NPP), the GPP minus the amount of energy used in the community for respiration (R). Examining dissolved oxygen is a means of looking indirectly at the members of an aquatic community and their productivity, based on their metabolic products.
In the presence of light, autotrophic members of the community produce oxygen as a by-product of photosynthesis. Heterotrophic and autotrophic members of the community both utilize oxygen during respiration, a process that is independent of light. Because of this, dissolved oxygen is used as a means of monitoring ecosystems.
Algae blooms alter the amount of oxygen in the aquatic ecosystem and can result in fish kills through multiple means. Some algae “secrete” toxins that can result in lesions on the fish and directly kill fish. Alternatively, if the algae “overgrows” it can block light to other photosynthetic organisms. The algae will produce oxygen while it is growing, but when it dies in large amounts oxygen can be depleted when the algae decays. Dissolved oxygen levels below 5 ppm indicate that the body of water is in danger of a fish kill.
Oxygen in water is determined through a series of chemical reactions that involve color changes. Oxygen present in the sample water is fixed by adding manganese sulfate and that in turn oxidizes added iodide ions to iodine. These chemical reactions result in turning the water brown (the presence of iodine). The more dissolved oxygen, the darker brown the water turns.
Three bottles are filled with lake water. One bottle is assayed for dissolved oxygen immediately (initial bottle). One bottle is covered in foil (dark bottle) and one bottle is left uncovered (light bottle) and put in the lake for the next 24 hours. The next day, the light and dark bottles are assayed for dissolved oxygen.
In a closed bottle that is exposed to light (light bottle), both respiration and photosynthesis occur. In absence of light (dark bottle), only respiration (R) will occur. The changes in dissolved oxygen in the incubated light and dark bottles inform us about the prevalence of plants (autotrophic) and animals (autotrophic) in the tested water.
To determine the amount of oxygen produced in a known time period, the oxygen in a bottle collected and measured at the beginning of the time period (Initial bottle) is subtracted from the light and dark bottles. You will determine the dissolved oxygen in the Initial bottle for the class that takes place 24 hours after your class.
To determine the oxygen in a sample, reagents 1 and 2 of the oxygen assay kit are added and the sample is incubated 10 minutes with intermittent shaking. The measuring tube is used to add 5.8 mL of solution to the viewing vial, and the solution is titrated by adding drops of sodium thiosulfate in the presence of a starch indicator until the solution turns clear. The drops of sodium thiosulfate added are counted and the amount of oxygen can be calculated from the amount of sodium thiosulfate needed to turn the solution clear. Each drop correlates with one ppm of dissolved oxygen which is equivalent to 1 mg/L.
Calculations for the NPP, GPP, and R
Result units are mg/L/hr dissolved oxygen:
NPP = Light − Initial
GPP = Light − Dark
R = Initial − Dark
Please view the following video for further explanation of the Dissolved Oxygen Assay:
Dissolved Oxygen from Hayden-McNeil on Vimeo.
Proceed to the Pre-Lab Quiz