BIO 111L

Environmental Science Laboratory

Biological and chemical Analysis of Habitats

As a part of the assessment of various habitats, you will be performing tests on the chemical characteristics of soil and water. Water tests will generally be performed by several groups of four to five persons each, with each group performing most of the same tests and sharing the collective data. This gives each person the opportunity to participate in each test. Students should observe tests carefully so that they can perform them again in a subsequent lab. The groups will collect samples from different locations, and since the values for different locations may vary, and since technique will vary from one person to another, sharing the data will allow an average to be determined which more accurately reflects the actual value than a

single result. Data should be recorded on the data sheet for the specific field lab, as well as in the data tables for the Field Evaluation lab near semester's end.

I. Water Testing

Most of the tests are in kit form and follow a "cookbook" procedure. The instructor will describe basic procedures the first time tests are performed. Pay attention so that you can do the tests later. The tests are of two types:

A. Titration is a process by which the precise amount of a chemical, called the reagent (short for reactive agent), which is used in a chemical reaction indicates the amount present of a certain substance. An instrument called a titrator, which looks like a hypodermic syringe is used to deliver the reagent. The titrator is marked in units which are read directly as PPM (parts per million) for the substance being tested. The procedure requires particular attention paid to the exact endpoint of the reaction which is characterized by a color change. At the instant the color changes the reaction is over and the amount can be determined. An example is the dissolved oxygen test in which a substance called sodium thiosulphate is the titrating reagent and at the endpoint the solution changes from blue-black to clear. At that instant titration should stop and the total amount of sodium thiosulphate used indicates the PPM (parts per million) of dissolved oxygen. The critical point in titration technique is making certain to mix the solution by shaking gently after each drop of titrating reagent is added in order to see the color change and not go past the endpoint. Another example is the test for hardness (calcium carbonate) in which the color changes from pinkish to blue. The endpoint occurs at the instant the solution is no longer pink.

B. The other type of test is called a colorimetric reaction in which the result of the test gives a color which is compared to standards which illustrate various levels in PPM. If the color appears to be intermediate between standard values interpolation is used to estimate the value. This test leaves less room for error than titration. A critical point may be finding more of the substance in your sample than the standards, as evidenced by a much darker color than the standards. In that event dilute your sample with distilled water to 1110 (one drop of sample added to 9 drops of distilled water), compare to the standards, and multiply your result by 10. The nitrogen, phosphorus, and silica tests are of this type.

C. The Tests:

1. dissolved oxygen (D.O.) - Oxygen levels are critical for normal

biological health in a body of water and oxygen is usually the limiting factor in polluted water.

Dissolved oxygen is measured in PPM (parts oxygen per million parts of water) and a level of 8

PPM is considered the minimum necessary for biological health. Since oxygen levels are

impacted by both plant and animal physiology, they are quite volatile and begin to change as

soon as a sample is taken. For this reason the test must be done, or the sample fixed,

immediately in the field.

2. biochemical oxygen demand (also biological oxygen demand, BOD)

This represents the demand for oxygen placed on the system by the normal flora and fauna as

well as that contributed by eutrophication. The BOD of a body of water must be less than the

dissolved oxygen available plus a safety margin.

3. nitrates, phosphates, ammonia nitrogen - These are nutrients

contributed by biological processes such as decomposition, and by weathering of minerals from

rock. They also come in larger amounts from eutrophication sources such as sewage effluent,

runoff from agriculture, livestock, etc. Their detectable presence usually indicates such

contamination.

4. pH - The measure which indicates the acidity and an important factor in

determining normalcy for biological systems.

5. hardness - Indicates levels of carbonates, mostly calcium, which reflect

geological factors and determine the base conditions for living organisms.

6. heavy metals - Lead, mercury, arsenic, chromate etc. which are toxic to

life and represent contaminants.

II. Soil Testing

Just as with water the tests are in kit form with "cookbook" instructions provided

with each kit. Pay attention to modifications announced by the instructor. Samples of the upper

few inches of organic soil (raw litter from the surface is not used) are taken at each site and

tested later in the laboratory. The soil may need to be screened to remove larger chunks and

produce a uniform ~me consistency. For most tests an extract of the soil is made first which will

then contain the mineral to be tested. It may be more efficient to make one extract in great

enough quantity to do several tests.

The Tests:

1. pH - levels of acidity are important in the ability of the soil to support

plant growth other biological processes, and also reflect the impact of geology and weathering,

as well as man's activities.

2. nitrates, phosphates - These are critical soil nutrients which derive

from decomposition but in larger quantities may reflect importation from fertilizers, manure, etc.

3. nitrites - Their presence may indicate lack of aerobic processes

necessary in the formation of nitrates.

4. ammonia - Generally low in quantity, except in forest soils, its

presence may indicate additives or inhibition of biological processes.

5. humus - This represents the soil's organic matter, it is critical to

maintain normal biological processes and productivity and to support nutrient retention.

III. Sampling of Aquatic Organisms

As a measure of the biological health of an aquatic system, as well as a tool for measuring biological diversity, sampling is unmatched. Sampling can take may forms from fine-mesh nets which filter the water, or fine screens to filter the mud, to a series of chronologically collected samples which identify the successional stages by which an organic or inert substrate is colonized by various organisms.

We will use a variation of the latter technique which itself is quite simple. Upon visiting a lake, river, etc we leave a tied bundle of leaves or other substrate in the water for algae, insects, larvae, and the like to latch onto. The nature of the substrate itself, whether organic or inert, whether deciduous leaves or evergreen needles, can be an important and interesting variable in the study. At prescribed intervals (the peak number or organisms usually occurs at between 7 and 10 days after insertion) the packets are removed from the water and the organisms are fixed with AFA solution* and taken back to the lab. Once fixed, the samples are preserved and can be viewed at the class's or researcher's leisure. Using guides to the identification and classification of aquatic insects we will compare those taken at different times or in different habitats.

*AFA = alcohol-formalin-acetic acid. CAUTION: Adequate ventilation is important.

Those wearing contact lenses should consider not wearing these on the day the specimens are

viewed. Ask your instructor.

IV. Biodiversity

Ecologists have devised a number of indices to compare the biodiversity of habitats. All of these methods are complicated by the nature of the problem and its mathematical expression. Since this is not an ecology course and statistical analysis and mathematical procedures are not our focus, we will perform a very rudimentary analysis of diversity which will allow us to compare the places we visit. Called Simpson's Diversity Index, this analysis considers both the number of different species and their evenness of distribution. For example, if two different habitats each have twelve different species and a total 120 organisms each, but habitat A's biota consists of ten each of the twelve, while habitat B has 109 of one dominant species plus l each of the other 11, then habitat A would be considered more diverse. For practical purposes our study will be confined to plants.

Calculation of Simpson's Diversity Index:

1) Select and measure a suitable area for study. Depending on habitat type, an area might range from 1 m² to 100 m² in size. Randomness is important, but variability and suitability are important as well.

2) Identify, the total number of plants and the number of' each specific type of plant species in the area. Record these in the table.

3) Determine the proportion of' each different species, Pi, where i is the iteration number or number of the sample. For example, if there were 100 total plants in the sample, and of those 100 twelve were ponderosa pines the proportion of ponderosas is 12/100 or .12. You need not actually identify each species. You may simply describe it and count the number.

4) Square each Pi and sum all of the squares. Then divide this sum into 1. This will give you D, the diversity index. Simpson's Diversity Index is useful only to compare different areas to which the calculation has been applied.