Introduction to Transpiration (A-level Biology)

Introduction to  Transpiration

Transpiration

Transpiration is where plants absorb water through the roots. It then travels up through the plant, in the xylem vessels, getting released into the atmosphere as water vapour through the leaves’ pores. Whilst water and oxygen are exiting through the leaf’s stomata, carbon dioxide enters.

The transpiration stream is the movement of water up the stem. It enables processes, for example, photosynthesis, growth and elongation. This is because it supplies the plant with water. This is required for all of these processes to take place.

It also supplies the plant with minerals () that are needed, as well as aiding in it controlling its temperature via evaporation of water.

The process of transpiration involves osmosis. This is because the water moves from the xylem cells to the mesophyll cells.

Evaporation and diffusion occurs in order to aid transpiration. Evaporation occurs at the mesophyll cell surface and moves into the intercellular space around it. Diffusion, in this case, is where water vapour moves out of the stomata, down a water vapour potential gradient.

The use of a potometer can help investigate the rate of transpiration. The water lost by the leaf gets replaced from a capillary tube, enabling you to measure the bubble’s movement, and therefore, the rate of transpiration.

Factors that can effect the rate of transpiration:

• Air movement – removes still air from the area surrounding the leaf, increasing the concentration gradient and rate of diffusion.
• Humidity – the rate of diffusion decreases in response to the concentration gradient decreasing.
• Temperature – water air can hold more water vapour, which increases the rate of evaporation and so also transpiration. Random motion is also increased.
• Light intensity / Amount of light – can cause more stoma to open for gas exchange, though this plateaus when all of the stomata become open. There is also importance in the size and position of the stomata which affects the rate of transpiration.
• Number of leaves
• Waxy cuticle presence
• Water availability

Xerophytes

Xerophytes are plants that are adapted to live in very dry conditions by mechanisms which minimise water loss and/or store available water.

Two examples of xerophytes are cacti and agaves.

Adaptations can include:

• Smaller leaves – reduces surface area
• Rolling leaves – reduces exposure of lower epidermis to the atmosphere
• Densely packed mesophyll – prevents water loss via evaporation
• Thick, waxy cuticles
• Close stomata – in response to low water availability
• Hair and pits – reduces water vapour potential gradient by trapping moist air
• Deep roots or wide-spreading shallow roots

Hydrophytes

Hydrophytes are plants which can live in water (e.g., water lilies). They  do not need many adaptation to live in this environment.

Hydrophytes adaptations:

• Thin or absent waxy cuticle – no need for conserving water supplies
• Lots of stomata open constantly – maximises gas exchange by being placed on the upper surfaces
• Flat and wide leaves – gives larger surface area to increase light absorption
• Air sacs (found in some) – aids in flotation

Practical 1

Observing and Drawing a Stained Section of Plant Tissue Under a Light Microscope

It is important to understand what you are seeing and how to appropriately annotate a diagram of a stained section of a plant tissue under a light microscope.

An eyepiece graticule allows structures to be measured whilst under a microscope. It acts as a ruler and can be fitted to the microscope directly.

A stage graticule (also known as a stage micrometer) can be used in place of a regular microscope slide. It has a measuring scale on it in order to be able to calibrate the eyepiece’s value at different magnifications.

Calibrating an Eyepiece Graticule

1. Set up the light microscope as usual.
2. Stage. Insert a stage graticule onto the stage.
3. Line up graticules. Using the diagram, try to line the stage and the eyepiece graticules as closely as possible.
4. Identify. Identify the number of divisions on the eyepiece which are equal to every division on the stage graticule.
5. Calculate. Calculate the length on one eyepiece division as the division on the stage graticule is now known as we know the length equivalent.

If the microscope’s magnification changes, the eyepiece graticule needs to be recalibrated.

Repeat the process for every objective lens to have a calibration factor for all of them.

Slide Preparation

1. Cut cross-section. On a white tile, cut a sample of the plant, as thinly as possible using a sharp scalpel. It should be perpendicular to the stem.

2. Place in water (2 minutes). This prevents drying out of the specimen.

3. Set up the light microscope a usual.

4. Staining. Making sure to use the appropriate dye, place the specimen into a watch glass which contains the chosen dye.

Dyes are crucial to distinguishing between structures by providing contrast, it is important to make sure you choose the right one(s).

Multiple dyes can be used as they each bind to different structures. This is known as differential staining.

• • Acetic-orcein – Stains chromosomes and connective tissues dark red. It is a biological stain and commonly bind to the DNA (hence staining the chromosomes).
  • Eosin Y – Stains alkaline structures (e.g., cytoplasm) pink.
  • Iodine – Stains starch in carbohydrates blue-back / brown. Stains glycogen red.
  • Iodine in potassium iodine solution – Stains cellulose yellow.
  • Haematoxylin – Stains DNA (and RNA) purple-blue
  • Methylene blue – Stains acidic structures (e.g., nucleus) blue. Often used to stain DNA.
  • Toluidine – Stains DNA in meristem cells. Lignin and tannins turn blue-green, pectins turn pink-purple whilst nucleic acids turn purple or green-blue. Most commonly used dye.

5. Mounting the sample. There are two techniques you could use (we use a different technique here):

• Wet mount
• Dry mount

The wet mount technique is often used on live specimens, such a those that live in water.

1. Water. Place one drop of water on the slide via a pipette.
2. Specimen. Place it in the drop of water, using tweezers.
3. Cover slip. Carefully place the cover slip on the specimen without creating bubbles.
4. Stain. On one side of the cover slip, place a drop the chosen stain. On the other side, place a paper towel to absorb the stain, pulling it under the cover slip. Make sure the stain is not toxic to  live specimens.

The dry mount technique is used for specimens such as hairs, insect parts and pollen.

1. Specimen. Slide it into a thin enough piece for light to be able to shine through.
2. Place. Put it on the middle of the slide, using tweezers.
3. Cover slip. Place the coverslip on the specimen.

6. Wash. Rinse under the tap, by holding on using tweezers, to rid of any excess stain.

7. Place. Putting the specimen on the slide, add a drop of water and a coverslip. (Repeat for the thinnest sections)

8. View. Using the lowest magnification lens first, adjust the knobs until the image becomes clear.

9. Draw. Create an annotated diagram of what you can see under the microscope. How to draw a scientific diagram explained at the end.

10. Repeat. In order to view a longitudinal section instead of the transverse sections we used, repeat the process.

What you should see:

• Phloem
• Xylem
• Collenchyma
• Parenchyma
• Sieve tubes
• Sclerenchyma

Practical 2

Light Microscope

Now it is time to view the specimens under the microscope.

1. Stage. Place slide under the clips on the stage.
2. Choose objective lens. Selected the lowest powered one first and work your way up.
3. Adjustment. Use the coarse adjustment knob to move the lens just above the slide. Make sure to not do this looking down the microscope, or you may break the slide.
4. Focus. Use the fine adjustment knob until you begin to see a clear picture of the specimens through the lens.
5. Increase magnification. Restart the process of adjustment and focus with a higher magnification lens.

Measuring

Using a calibrated eyepiece graticule, measure the structures by counting the number of eyepiece divisions. Multiple this by the length on one division to find the actual length.

If the diameter is an irregular shape (e.g., a cell), use the length of the greatest distance.

Drawing and Annotations

Using a sharp pencil, draw what you see, making sure it covers at least half of the page.

Instead of shading an area (which you shouldn’t do), use a ruler to draw the annotation lines, pointing to the exact area, as these should be straight and horizontal, without overlapping.

Always make sure to give a scale.

Practical 3

Using a potometer

A potometer can measure the rate of transpiration as an estimate, by measuring the water uptake of a plant. It assumes that water uptake by the plant and water loss by the leaves are directly related.

It is set up underwater to help avoid unwanted air bubbles in the xylem vessels.

It has some limitations:

• Other water uses. Due to some of the water being using up by the plant for other uses, such as turgidity maintenance, not all of the water taken up by the plant is measure, hence, we can calculate an estimate.
• Other processes. Some of the water is used in photosynthesis.
• Death. Due to the plant being a living specimen, as soon as its roots are removed, is begins dying. In response to this, less water may be taken up.

The process:

1. Cut shoot. Making sure to cut the shoot underwater, as prevents air entering the xylem, whilst cutting at a slant increases the surface area.
2. Insert the shoot. The assembly of the potometer underwater also helps to prevent any air entry.
3. Remove apparatus. Whilst keep the capillary tube in a beaker full of water, remove the apparatus from the water.
4. Check for gaps. Use screws or petroleum jelly to make sure the apparatus is both watertight and airtight.
5. Dry. Make sure the leaves are all dried.
6. Acclimatisation. After making sure the plant has had time to acclimatise, shut the tap.
7. Test for air bubbles. By removing the capillarity tube from the beaker, wait for an air bubble to form, then put it back into the water.
8. Note. Make sure to write down a reading of the starting position of the air bubble.
9. Record. Using a stopwatch, record how far the bubble moves at a regular time interval (r.g., every 15 minutes).
10. Calculate. Divide the distance the bubble moved the the time it took to move there to calculates an estimation of the rate of transpiration.

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