The aim of this report is to investigate the effect of conditions: temperature, light, moisture, sugar and there effect on the size and density of stomatal pores. The experimental hypothesis is that the greater the light intensity the bigger size and lower density of stomatal pores and the more water there is the smaller the pores and the higher number of stomatal pores. Samples from two plants were experimented on, the Geranium plant and the Kalanchoe daigremontiana plant. From both plants 1cm by 1cm samples of the leaves were taken to measure the stomata sizes and to count the number of stomatal pores. The results showed that the condition with the greatest effect is the light conditions which showed greater stomatal size and density.
Introduction
Although sotmatal opening may seem like … it could contribute in climate change, as we know climate change is due to the rapid increase of green house gasses and in particular carbon dioxide (CO2) in our atmosphere and thus this is a serious current global concern.
Stomata
Stomata are tiny structures found on the epidermis of a plant. The tiny pore called the stoma is surrounded by guard cells which are specialised cells. The main function of the stomata is to allow gases such as carbon dioxide, water vapour and oxygen to move rapidly into and out of the leaf. Since most of the water (90%) is lost through stomata, plants regulate the degree of stomatal opening to reduce the water loss. The guard cells have unevenly thickened walls. The cell wall around stoma is tough and flexible and the one away from stoma is thinner.
Stomata are openings generally present on the lower surface of the leaves through which the gases and water vapour diffuse in and out easily. The oxygen diffuses in through the stomata and then enters the leaf cells. Similarly, the carbon dioxide produced by the leaf cells diffuses out through the stomata.
Stomata are generally open during the day so that COâ‚‚ can enter he plant for photosynthesis (which needs light). They are usually closed at night to save water, because COâ‚‚ is no longer needed (photosynthesis can’t happen in the dark).
What is photosynthesis?
Photosynthesis is the process by which plants, some bacteria, and some protistans use the energy from sunlight to produce sugar, which cellular respiration converts into ATP, the “fuel” used by all living things. The conversion of unusable sunlight energy into usable chemical energy, is associated with the actions of the green pigment chlorophyll. Most of the time, the photosynthetic process uses water and releases the oxygen.
Plants are the only photosynthetic organisms to have leaves (and not all plants have leaves). A leaf may be viewed as a solar collector crammed full of photosynthetic cells.
The raw materials of photosynthesis, water and carbon dioxide, enter the cells of the leaf, and the products of photosynthesis, sugar and oxygen, leave the leaf.
Cross section of a leaf, showing the anatomical features important to the study of photosynthesis: stoma, guard cell, mesophyll cells, and vein (Farabee, 2007)Water enters the root and is transported up to the leaves through specialized plant cells known as xylem (pronounces zigh-lem). Land plants must guard against drying out (desiccation) and so have evolved specialized structures known as stomata to allow gas to enter and leave the leaf. Carbon dioxide cannot pass through the protective waxy layer covering the leaf (cuticle), but it can enter the leaf through an opening (the stoma; plural = stomata; Greek for hole) flanked by two guard cells. Likewise, oxygen produced during photosynthesis can only pass out of the leaf through the opened stomata. Unfortunately for the plant, while these gases are moving between the inside and outside of the leaf, a great deal water is also lost. Cottonwood trees, for example, will lose 100 gallons of water per hour during hot desert days. Carbon dioxide enters single-celled and aquatic autotrophs through no specialized structures.
Stomata opening and closing
most plants do not have the aforementioned facility and must therefore open and close their stomata during the daytime in response to changing conditions, such as light intensity, humidity, and carbon dioxide concentration. It is not entirely certain how these responses work. However, the basic mechanism involves regulation of osmotic pressure.
When conditions are conducive to stomatal opening (e.g., high light intensity and high humidity), a proton pump drives protons (H+) from the guard cells. This means that the cells’ electrical potential becomes increasingly negative. The negative potential opens potassium voltage – gated channels and so an uptake of potassium ions (K+) occurs. To maintain this internal negative voltage so that entry of potassium ions does not stop, negative ions balance the influx of potassium. In some cases chloride ions enter, while in other plants the organic ion malate is produced in guard cells. This in turn increases the osmotic pressure inside the cell, drawing in water through osmosis. This increases the cell’s volume and turgor pressure. Then, because of rings of cellulose microfibrils that prevent the width of the guard cells from swelling, and thus only allow the extra turgor pressure to elongate the guard cells, whose ends are held firmly in place by surrounding epidermal cells, the two guard cells lengthen by bowing apart from one another, creating an open pore through which gas can move.

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When the roots begin to sense a water shortage in the soil, abscisic acid (ABA) is released. ABA binds to receptor proteins in the guard cells’ plasma membrane and cytosol, which first raises the pH of the cytosol of the cells and cause the concentration of free Ca2+ to increase in the cytosol due to influx from outside the cell and release of Ca2+ from internal stores such as the endoplasmic reticulum and vacuoles. This causes the chloride (Cl-) and inorganic ions to exit the cells. Secondly, this stops the uptake of any further K+ into the cells and subsequentally the loss of K+. The loss of these solutes causes a reduction in osmotic pressure, thus making the cell flaccid and so closing the stomatal pores.
Interestingly, guard cells have more chloroplasts than the other epidermal cells from which guard cells are derived. Their function is controversial. (stomata opening, 2008)
This diagram shows normal responses of stomata to light, CO2, pH, K+ ion and Water Deficiency
Transpiration
Transpiration is the term used to describe the transport of water through an actual, vegetated plant into the atmosphere. Transpiration is an important part of the evapotranspiration process, and a major mechanism of the water cycle in the atmosphere. Transpiration may also refer to the rate of the water vapor transport through the whole vegetative canopy (that is, through the group of plants).
Just as you release water vapor when you breathe, plants do, too-although the term “transpire” is more appropriate than “breath.” During this process individual water molecules are released from the surface of the plant body through tiny structures called stomata. There are many more individual water vapor molecules inside the air spaces between the tissues of a plant than in the air surrounding the plant body. Consequently water vapor will always exit the plant along a concentration gradient. As more water vapor molecules exit the plant, the remaining water molecules tug on each other and will pull an entire column of water throughout the plant body through special tissues called xylem during the process of transpiration. One way to visualize transpiration is to put a plastic bag around some plant leaves. As Figure 1 shows, transpired water will condense on the inside of the bag. If the bag had been wrapped around the soil below it, too, then even more water vapor would have been released, as water also evaporates from the soil. During a growing season, a leaf will transpire many times more water than its own weight. An acre of corn gives off about 3,000-4,000 gallons (11,400-15,100 liters) of water each day, and a large oak tree can transpire 40,000 gallons (151,000 liters) per year.
Factors affecting transpiration
The amount of water that plants transpire varies greatly geographically and over time. There are a number of factors that determine transpiration rates:
Temperature: Transpiration rates go up as the temperature goes up, especially during the growing season, when the air is warmer due to stronger sunlight and warmer air masses. Higher temperatures cause the plant cells which control the openings (stoma), where water is released to the atmosphere, to open, whereas colder temperatures cause the openings to close.
Relative humidity: As the relative humidity of the air surrounding the plant rises the transpiration rate falls. It is easier for water to evaporate into dryer air than into more saturated air.
Wind and air movement: Increased movement of the air around a plant will result in a higher transpiration rate. This is somewhat related to the relative humidity of the air, in that as water transpires from a leaf, the water saturates the air surrounding the leaf. If there is no wind, the air around the leaf may not move very much, raising the humidity of the air around the leaf. Wind will move the air around, with the result that the more saturated air close to the leaf is replaced by drier air.
Soil-moisture availability: When soil moisture is lacking, plants can begin to senesce (premature ageing, which can result in leaf loss) and transpire less water.
Type of plant: Plants transpire water at different rates. Some plants which grow in arid regions-for example, cacti and succulents-conserve precious water by transpiring less water than other plants.
(Burba & Pidwirny, 2007)
Kalanchoe Daigremontiana
Kalanchoe daigremontiana and also sometimes called Mother of Thousands, is a plant from southwest Madagascar. The plant generally would reach up to 3 feet (1m) tall and consist of leaves that reach 6-8 inches long and about 1.25 inches wide. These are medium green above and blotched with purple underneath. The most interesting feature of this plant is the fact that it has spoon-shaped bulbiliferous spurs that bear young plants on its margins. This plant is distinguished by its ability to propagate via vegetative propagation. All parts of the plant are poisonous, which can even be fatal if ingested by infants or small pets.

Blooming: In the greenhouse, the plants bloom sporadically in late winter. The compound cymes have 1 inch (2.5 cm) long purplish flowers.
Culture: Kalanchoe daigremontiana needs full sun to partial shade with a well-drained soil mix. In the greenhouse, soil is mixed consisting of 1 part peat moss to 2 parts loam and sand. The plants are watered and allowed to dry slightly before watering again. They are generally fertilised only once during the season with a balanced fertiliser. During the winter months, water is somewhat restricted, but the plants are not allowed to dry out completely. The plants can become very weedy, so these plants should not be used around other plants. Plantlets are drought resistant, root readily, and if allowed to establish, can easily create a plant epidemic wherever the plantlets land (hence their common name).
Propagation: Kalanchoe daigremontiana is easily propagated from plantlets formed on the edges of leaves or from cuttings. Cuttings must be kept very dry to root.

(Kalanchoe daigremontiana – Mother of Thousands, 2006)
Geranium
Geraniums are just one of the members of the family Geraniaceae. Pelargonium are also a member of the same family so that confusion has arisen by both of them being referred to as Geraniums
The Geranium as it is commonly known, is actually of the genus Pelargonium. Geranium is the correct botanical name of the separate genus that contains the related Cranesbills. Both genera are in the Family Geraniaceae. Which originally included all the species in one genus, Geranium, but they were later separated into two genera by Charles L’Héritier in 1789. The first species of Pelargonium known to be cultivated was Pelargonium triste, a native of South Africa. The cranesbills make up the genus Geranium of 422 species of annual, biennial, and perennial plants found throughout the temperate regions of the world and the mountains of the tropics, but mostly in the eastern part of the Mediterranean. One can make the distinction between the two by looking at the flowers : Geranium has symmetrical flowers, while Pelargonium has irregular or maculate petals. The name “cranesbill” derives from the appearance of the seed-heads, which have the same shape as the bill of a Crane. The genus name is derived from the Greek word geranos, meaning ‘crane’.
All geranium species are perennials and generally winter hardy plants, and are generaly grown for their attractive flowers. They generally have a long lifespan and most have a mounding habit, with palmately lobed foliage. Some species have spreading rhizomes. They are normally grown in part shade to full sun, in well draining but moisture retentive soils, that are rich in humus. They are generally found in Mediterranean areas. (The Origin of the Geranium, 2006)
Propagation: is by semi-ripe cuttings in summer, by seed, or by division in autumn or spring. (Geranium, 2009)
Experiment
Aim
To investigate the effect of conditions: temperature, light, moisture, sugar, can effect on the size and density of stomatal pores.
Hypothesis
The greater the light intensity the bigger size and lower density of stomatal pores; the more water there is the smaller the pores and the higher number of stomatal pores.
Apparatus

Microscope
Slides
stage micrometer
Leaves
Sugar solution
Iodine (optional)
Tweezers
Petri dish
Clear nail vanish
Beakers

Method

Select the leaves which will be used
Put them in the different conditions over night
Next day, polish all the leaves with a rarther thick layer of clear nail vanish
Allow to dry for about 2 hours
When completely dry, gently peal of the nail polish
Put it on the slide and observe with the microscope, also insert the eye piece graticule to measure the size of the stomata
(optional) if not very visible, apply iodine on the slide put the piece of nail polish on top and press down with tissue to clear of excess solution
Count the number of stomata visible and measure the size

Results
We see here that in the conditions with light and sugar the stomata appears to have the greatest size whereas in water conditions and dark condition stomatal size appear to be very small.
Here we see that stomatal size appear to be generally larger with the lowest size to be 0.02 mm which is for the condition with sugar placed in the dark. The condition with the highest stomatal size is the condition with water placed in the light with 0.08mm in size, followed by the condition with sugar placed in light with a stomatal size of 0.07.
The graph shows that the condition with sugar placed in the dark had the lowest stomatal density followed by the dark condition with 55 stomatal pores in the 1 cm by 1 cm sample. The conditions with light seem to have the highest number of stomatal pores with the highest condition, water placed in light having 120 stomatal pores followed by the condition in purely light having 98 stomatal pores. On the other hand the condition with sugar placed in light has the lowest conditions with 30 pores per 1cm square.
Generally the number of stomatal pores in the 1 cm by 1 cm sample are fairly quite low with the highest number of pores present in the condition presented with light with a number of 99 pores. The condition with the lowest number of pores was the condition in water placed in the dark with 25 followed by the condition in the water placed in the light with followed by the condition in the water placed in the light with 28 pores.
Discussion
From the results it is clear that the stomatal size is greatly dependent on the conditions in which it is located. For the geranium we can see that the stomata has the greatest size of 0.1 mm (see appendix 1) in the condition with light, here it is possible to accept the hypothesis, the fact that it has a greater stomatal density in conditions with the most light. On the other hand the Mexican hat showed lower numbers such as 0.04 mm. This may be because of the origin of this sort of plant and the fact that they are generally found in hot countries where there is generally more light could have ment that these sort of plants undergo various adaptation and therefore have adapted to have smaller stomatal sizes even thought there is a lot of light present.
We also notice that the geranium has the most stomatal numbers (120 stomatal pores in a 1cm by 1cm sample of leaf) in the water condition; this shows that stomatal numbers increase in the presence of more water. Here also the Mexican hat leaf shows the opposite, here it has a very low stomatal number, this may be an anomaly or may be purely due to the fact that the plant is found in areas with water shortages and therefore here again it may have adapted to maintain low numbers of stomata pores in water.
In the conditions with sugar we see that sugar doesn’t have much effect on the Mexican Hat since the results for the conditions with sugar placed in a area with light had results very close to the results for the conditions in water placed in light conditions. Bearing in mind the fact that the sugar solution was made up of water mixed with sugar, for the results to be similar to the results with water only we can assume that what is causing the effect is in fact the water and not so much of the sugar. On the other hand from the graphs we see that it may have some effect on the Geranium plant since the results alter for conditions in only water than the results for conditions in sugar.
The conditions in the fridge showed quite average to low results, this is because in the fridge it had lower temperatures and therefore colder temperatures cause the openings to close.
During the investigation it was almost impossible to find the actual stomatal pore size between the guard cells and you rarely get the chance to measure the diameter of the hole therefore vague assumptions had to be drawn by just having to measure the length of the closed hole and make some assumptions from there.
Another way to find out whether stomata are open or closed, or more accurately, how open they are, is by measuring leaf gas exchange. A leaf is enclosed in a sealed chamber and air is driven through the chamber. By measuring the concentrations of carbon dioxide and water vapor in the air before and after it flows through the chamber, one can calculate the rate of carbon gain (photosynthesis) and water loss (transpiration) by the leaf.
However, because water loss occurs by diffusion, the transpiration rate depends on two things: the gradient in humidity from the leaf’s internal air spaces to the outside air, and the diffusion resistance provided by the stomatal pores. Stomatal resistance (or its inverse, stomatal conductance) can therefore be calculated from the transpiration rate and humidity gradient. (The humidity gradient is the humidity inside the leaf, determined from leaf temperature based on the assumption that the leaf’s air spaces are saturated with vapor, minus the humidity of the ambient air, which is measured directly.) This allows scientists to learn how stomata respond to changes in environmental conditions, such as light intensity and concentrations of gases such as water vapor, carbon dioxide, and ozone.
Reference
Image 1: http://www.tutorvista.com/search/stomata-opening
Image 2: http://www.tutorvista.com/search/stomata-opening
Image 3: http://www.tutorvista.com/search/stomata-opening
Bibliography
Burba, G., & Pidwirny, M. (2007, April 1). Transpiration. Retrieved December 15, 2009, from Encyclopedia of Earth : http://www.eoearth.org/article/Transpiration
Farabee, M. (2007, June 6). PHOTOSYNTHESIS. Retrieved December 15, 2009, from http://www.emc.maricopa.edu/faculty/farabee/BIOBK/BioBookPS.html
Geranium. (2009, November 10). Retrieved December 15, 2009, from Wikipedia, the free encyclopedia: http://en.wikipedia.org/wiki/Geranium
Kalanchoe daigremontiana – Mother of Thousands. (2006, September 21). Retrieved December 15, 2009, from Plant of the week: http://www.plantoftheweek.org/week375.shtml
stomata opening. (2008, – -). Retrieved December 15, 2009, from tutorvista.com: http://www.tutorvista.com/search/stomata-opening
The Origin of the Geranium. (2006, October 8). Retrieved December 15, 2009, from A SouthWest Oasis: http://aswoasis.wordpress.com/2006/10/08/the-origin-of-the-geranium/