The factors affecting the rate of permeability in a cell membrane?

- - Mohammed Faisal

The factors affecting the rate of permeability in a cell membrane?

The plasma membrane is a permeable membrane, and it controls exchanges between the cell inner and outer environment. The rate of membrane permeability is dependent on a many factors. These are:

Molecular size of the solute as the permeability mostly decreases with increasing size
Lipid solubility’s permeability usually increases with increasing fat or oil solubility
The degree of ionization as permeability mostly decreases with increased ionization
The pH and temperature also have great effect on the permeability of the plasma membrane and in the experiment I will test how temperature has an effect on the rate of permeability.

Roles of the components of cell membranes:

My study has shown me that cell membranes contain many different types of molecules and they each have different parts to play in the overall structure and function of the membrane, these molecules are;

Phospholipids – these forms a bilayer which is the basic structure of the membrane, they have non – polar tails which means that it is difficult for polar molecules or ions to get passed them it can be said they are a barrier to most water soluble substances.

Cholesterol – these molecules will help to get the fluidity of the membrane regular and it will prevent it from becoming too rigid or too fluid. Cholesterol is used also for the mechanical stability of membranes this is very important because without this membranes would break rapidly and cells would burst.

Proteins – The transmembrane proteins act as carriers and also channels for ions and glucose. Proteins also act as a receptor for hormones this will detect which hormone it is and what it does.

Glycolipids and Glycoprotein’s – these are only found on the exterior surface of the cell membrane, they are formed by carbohydrates attaching to lipids to form Glycolipids and to proteins to form glycoproteins, they act as receptors and aid in recognition of cells.

Features of the fluid mosaic model

The membrane is double layer (bilayer) of phospholipids molecules. The individual molecules move about by diffusion within there own monolayer.

The phospholipids tail point inwards, facing each other and forming a non-polar hydrophobic interior. The phospholipids heads face the aqueous solution (water containing) medium that surrounds it.
The phospholipids tails can be saturated or unsaturated.
If they are unsaturated they have more fluid in the membrane and if they are saturated then there is less fluid.
Most of the protein molecules float in the phospholipids bilayer
The average thickness is about 7nm
Proteins and lipids have branching carbohydrates which help to detect things.

Transport across plasma membrane

A phospholipids bilayer around cells makes a very effective barrier; this is mostly against the movement of water soluble things.

There are four main mechanisms, diffusion, osmosis, active transport and bulk transport by exchange.

Osmosis

Water molecules are small enough to pass between phospholipids molecules and they also pass through pore proteins. The movement of water into and out of cells is influenced by a variety of factors:

How much water is present in the cytoplasm and in the exterior environment.
The concentration of solutes, such as ions and sugars, on either side of the cell surface membrane.
In plants, the pressure exerted on cell contents by the cell wall

Water potential is the tendency for the water to move from one place to another and is determined by the factors listed above. Solutions with high water potentials have few solutes molecules; solutions with low water potentials have many dissolved solute molecules. Water moves from a solution with a high water potential to one with a lower water potential. The diffusion of water through a partially permeable membrane down a water potential gradient is osmosis.

Diagram : http://www.okc.cc.ok.us/biologylabs/Images/Cells_Membranes/osmosis.gif

Diffusion

Diffusion is the net movement of molecules from a place of higher concentration to a place of lower concentration. The molecules move down a concentration gradient. This is because of the kinetic energy which is the energy of movement in molecules or ions; this makes them move at random.

After diffusion molecules reach an equilibrium situation where they are evenly spread in a volume of space.

Facilitated Diffusion

Many molecules that cells require are too large to pass between phospholipids molecules. They have also been charged and therefore unable to pass through the hydrophobic region in the centre of the bilayer. Protein molecules exist in membranes to help (or facilitate) the diffusion of these substances.

Channel Proteins – are transmembrane proteins that form tunnels, or pores, through the bilayer for water soluble molecules. Some channels are open all the time, others open when triggered by the presence of a chemical such as hormone. The lining of the pre is formed by the hydrophilic R groups of the proteins that allow water and polar substances to pass through easily.
Carrier Proteins – change shape to help move molecules into or out the cell. Molecules bind to the protein which stimulates the protein to change its overall shape, so allowing the molecules to diffuse through membrane.

Diagram: http://www.blc.arizona.edu/courses/181gh/grimes/Membranes/graphics2/f5.111.jpg

Active Transport

Some substances required by cells are in a lower concentration outside the cell than inside. Cells cannot obtain there substances by diffusion. Root hair cells absorb nutrients, such as potassium ions, from the water in the soil. Carrier proteins similar to those used in facilitated diffusion move ions across membranes but against the concentration gradient. Root hair cells use energy from respiration to change the shape of the carrier proteins so that they can pump molecules or ions into cells. Active transport is also used to pump molecules and ions out of the cell.

Diagram: http://www.connect.ab.ca/~lburns/active_transport.gif

Beetroot

The red pigment contained in beetroots is called betalain this is what gives the beetroot its dark colour. The betalain is a group of nitrogen containing pigment this is normally yellow, orange, pink, red and purple. Beetroot is good for you, as it contains no fat, very few kilojoules, and is a great source of fibre. The shape of beetroot is spherical and it has some cylinder shaped stalks which stick out of it.

Preliminary Experiment

Before starting the real experiment, a preliminary experiment which will last for about 20 minutes will be carried out, this will help in deciding which pieces of equipment will be essential to use for the experiment. The preliminary experiment will also help decide how long the beetroot should be left in the water, this will be tested by leaving the beetroot for a period of time in distilled water and by seeing what time is sufficient for the beetroot to be left in the water I predict that about 5-10 minutes should be enough but I am still going to test for longer just encase.

The preliminary can also help decide how much water to use as too much water can dilute the colour of the beetroot too much and using too little water could leave too dark colour so the preliminary experiment should help decide how much water to use, I predict that 10cm³ should be suitable amount.

This is a preview of the whole essay

It will also help decide what range of temperature I should use; it will be tested at a high and low temperature. The preliminary will also give a good understanding of what the aim of the experiment is and what you should be more aware of and what you should look out for in the final experiment. The colour of the filter in the colourimeter will be decided when the preliminary experiment is done.

The preliminary experiment also would help in deciding the size of the beetroot and which cork borer size should be used.

Hypothesis:

These two hypotheses are based on scientific knowledge, the accuracy of the hypothesis will be found out when the experiment is carried out fully.

1. Will increasing the temperature mean that more colour is released from the beetroot?

The higher temperature will have more effect on the releasing of colour from the beetroot as the higher temperature will break down the membrane and so more colour will be released. The temperature must be kept constant or as much as possible because this will defiantly effect the permeability of the beetroot as if the temperature is increased or decreased this could effect how much pigment is released. My research show that the optimum temperature for the beetroot membrane is at about 30°C – 40°C and temperatures above this temperature going from 50°C Onwards will begin to denature the beetroot membrane.

2. Will leaving the beetroot in the heated water for a long period of time have more effects on the permeability of the cell membrane?

Leaving the beetroot in the heated water for longer then 5 – 10 minutes will be sufficient enough time to make a difference to the permeability of the membrane because the colour is released during that time and leaving it in the water for further periods of time will not make much more difference maybe 15 minutes should be the most that it should be left in for.

Method:

Pour 10cm³ of distilled water through a 10cm³ syringe into a 25cm³ test tubes, because distilled water is pure and has been distilled to get rid of impurities.
Cut the beetroot with a cork borer size 6 into equal pieces to make it a fair experiment because uneven pieces of beetroot can mean that some pieces will leak more than the smaller pieces.
Wash the beetroot with distilled water because it is clearer than tap water as it can maybe affect the experiment. This is done to get rid of excess red pigment and leakage from the beetroot.
Place the test tube in a water bath this will be done at two temperatures 20°C and 70°C this is because we know that the room temperature is about 23°C so we will use a temperature which is at about there and 70°C because 30°C - 40°C is about the optimum temperature for the membrane for releasing the pigment without being damaged or denatured. I am going to go above 30°C - 40°C to see what happens, and to see at which temperature the proteins denature.
Put the cut beetroots into four test tubes of distilled water. This is done when the test tubes are hot enough (this is tested with the thermometer).
The test tubes will be in the water bath for 5, 10, 15 and 20 minutes and a sample will be taken out of each test tube after each has lasted its time in the water bath. These will be left in the water bath in 20°C and 70°C. A water bath is better than a Bunsen burner because it can keep a more consistent even though it does go a bit higher or lower it is generally more reliable. A Bunsen burner cannot keep the constant temperature without having to switch it off and then turning it back on when the water has cooled down too much. The Bunsen burner if heating a test tube will continuously heat it up and the temperature will consistently rise as it gets hotter.
A stop clock will be used because it is digital it will count down in seconds and it is possible to do a countdown so a sound will be heard when the time is up this is very precise. It is much better than normal clocks because it counts in seconds so it is much more precise. A normal clock might not function properly e.g. going slower you might not be able to tell. The normal does not count digitally so it will be less precise at getting the tubes out at the write time as the clocks second hand might be going too fast or slow.
A thermometer was used to check the temperature, these are an accurate piece of equipment, because its principle is simple, it works as the liquid in it changes its volume relative to its temperature so basically liquids would take up less room when they are cold and take up more room when they are warm. The water bath dial though set at e.g. 50°C it might not be fully on 50°C so the thermometer is their to double check it and make sure that the water is heated to the desired temperature as close as possible.
To release the colour and to spread it I gently shook the test tube, this was done gently so liquid didn’t fall out.
Then some pure distilled water was placed in a beaker in a colourimeter this was done to see if the colourimeter was working and also to get the reading to the 100% clear mark.
Then a sample of each test tube was taken and was placed in the colourimeter.
The colour of the filter which was used was green or 550.

Results:

This table shows the percentage of light passing through solutions at 20°C.

This table shows the percentage of light passing through solutions at 70°C

The preliminary experiment helped in deciding how to carry out the final experiment. After the preliminary I decided to use 7 different temperatures these are going to be 20, 30, 40, 50, 60, 70 because between 20 and 30 is the room temperature and 70 is a very high temperature but is not too high were the water will start to boil. I also decided that 5 minutes was sufficient time to let the beetroot in the water because by then what was going to happen has happened and leaving it in for longer made no difference. The equipment list was decided after the preliminary was finished and the list has all the necessary equipment needed to do the experiment. The preliminary was vital in discovering which variable were to be controlled so that the experiment could be as accurate as possible.

Colourimeter:

The diagram below shows how a colourimeter works

Prediction:

I think that if the temperature does not exceed 40ºC - 50ºC which is the optimum temperature then the permeability of the plasma membrane shouldn’t be affected. When the temperature goes over the limits then the water expands the cholesterol, Glycolipids and phospholipids which put pressure on the membranes from inside. The part of the membrane that is lipid will become liquid which would make it open to leakage. The proteins in the membrane will denature and this will increase the permeability in the surface. All of this will mean that compounds could exit the cell because high temperatures will make the molecules shake and vibrate more frequent. All this fast movement will mean that there the organised structure will be disrupted and eventually the structure will be ruined, this is because the higher temperatures break the hydrogen bonds in the structure of the protein and the structure is lost.

If the membrane is damaged because then the structure will all fall apart and will not work anymore. As the water molecules will rapidly enter the cell membrane by osmosis and the cells will swell up and burst this will release the pigment and it will spread.

Equipment List:

Beetroot
Distilled water
Scalpel
Ruler
White tile
Cork borer for size
Colourimeter
Stop clock
Syringes 10cm³
Water bath
Thermometer
Test tube
Test tube racket
Measuring cylinder 25cm³
Pipette
Spatula
Tripod

Safety:

In order to carry out the experiment safely and precisely the following safety procedures must be performed.

Wear goggles
Wear protective overcoat
Tie back long hair
Do not eat in the laboratory
Put all coats and bags out of the way
Put all coats and bags out of the way
Put all chairs under the table
Do not run in the laboratory
Wash hands before and after the experiment
Carefully handle hot water
Do not throw things in the laboratory
Be aware of all open flames
Clear up water spillage on floors and desks.

Method:

I poured 10cm³ of distilled water in a 25cm³ test tube.
I cut the beetroot with a cork borer size 6 into equal pieces to make it a fair experiment because uneven pieces of beetroot can mean that some pieces will leak more than the smaller pieces, to the length of 20mm.
I then washed the beetroot with distilled water because it is clearer then tap water as it can maybe affect the experiment. This is done to get rid of excess red pigment and leakage.
I will place the test tube in a water bath this will be done at the following temperatures 20, 30, 40, 50, 60, 70°C
I put the cut beetroot into the test tube of distilled water this is done when the test tubes are hot enough this is tested with the thermometer
This will be left in the water bath in all the different temperatures for 5 minutes each.
I will use a stop clock.
To release the colour I shook the test tube, this was done gently so liquid didn’t fall out.
I then used pure distilled water in a colourimeter this was done to see if the colourimeter was working and also to get the reading to the 100% clear mark.
I then took sample of each test tube and placed them in the colourimeter, the water is set at 100% it registers and then the other is compared to it so how much light is transmitted through the coloured water. 100% goes through on clear water.. I used the green filter because you get the best set of results. Red filter is the same colour as the pigment. 550 green is at the other end of the colour wheel. Green is a complimentary colour to red and it is on the other end of the colour wheel.
I then repeated this method again this was done for the reliability of the results, and so that the results were also more accurate.

Results:

This table shows the percentage of light passing through solutions when the membranes of the beetroot have been emersed in different temperature.

Fair Test:

The reason I used a stop clock is because it is digital. It is much better than normal clocks because it counts in 1/10 then 1/100 so it is much more precise. Whereas a normal clock doesn’t count in microseconds and if it is not functioning properly e.g. going slower you might not be able to tell
The colourimeter will give exact results on the colour of the water as it will scan it and give accurate results.
I am using a syringe because the units are in cm³ and it is 10 cm³ which was exactly how much water I am going to use in the test tubes for the experiment. And it is much more accurate then a measuring cylinder as it will duck up the water when you pull the tail back and it will immediately stop sucking up the water when you stop pulling the tail backwards.

Calibration Curve

This Table Shows the Figures for Calibration Curve

This was provided to me by the technician in the graph I will draw a calibration curve.

The calibration curve gives an accurate measure of the percentage of pigment released.

If you take a sample of a liquid such as in my experiment I had beetroot in water and shine light through it, it will give you a percentage of transmission that is how much light has been able to pass through it. The transmission percentage points if you then plot them on a calibration curve our starting points were provided by the school technician the calibration curve will give a good estimate of how much pigment is released.

This table shows the pigment released according to my transmission results when plotted on the calibration curve

The table and graph show a trend that as the transmission percentage is decreased the pigment being released is increased. This means that at higher temperatures, the membrane protein breaks down and denatures this leads to more pigment being released. The result show as the transmission is 55% the pigment released is 2% this is at the 20°C but when the transmission is 1% the pigment released is 50% this is at 70°C.;

Analysis:

My graph show as the temperature increased the amount of pigment released increased, the proteins in the membranes which are the phospholipids, cholesterol begin to denature when the temperature increases from 60 ºC - 70 º ºC. I know this because when the temperature is increased to these amounts the reading of the colourimeter goes to 1%.

In my results I had an anomaly at 50ºC on the first reading and second reading this result is not go in the trend of the rest of the results so it is anomalous this could have been done through a few reasons:

My results show that, generally as you increase the temperature of the water that the beetroot is placed in, then the colourimeter reading decreases as you can see in the graph at 20ºC the colourimeter is 58% but as it gets to 50ºC it has dropped down to 21% in the first reading and in the second reading 20ºC is at 52% but at 50ºC it is again much lower at 24%.

This is because as my first hypothesis was that as the temperature increased more red pigment was released this is because the higher temperature breaks down the phospholipids, cholesterol and proteins in the membrane and so more colour is released.

So my conclusion is that the increasing of temperature increases the permeability of cell membranes.

My graph shows that when the temperature is low around 58% of light is transmitted through and as the temperature is increased the amount of light transmitted is decreased. When I reached 60ºC as I increased the temperature the light transmitted through begins to remain the same and so creating a straight line on the graph. The lowest amount of light transmitted was 1% which was at the temperature of 60, 70ºC.

Using scientific knowledge I can say that this pattern is right because the permeability should increase as the temperature increases. I have researched that cell membranes are a phospholipids bilayer with proteins attached to them, 45% of the membrane is made of phospholipids. The beetroot cells have the red pigment in them called betalain this is what gives the beetroot its red colour, the pigment molecules cant leave the cell because they are too big, but when placed in hot water parts of the membrane are broken down which will damage it and leave big holes which are big enough for the pigment molecules to escape out and enter the water darkening its colour.

When you increase the temperature of the water there will be more heat and more pressure on the membrane so this gains more chance that the membrane will be broken down which will leave more holes for the red pigment to leak out and so more pigment molecules will enter into the surrounding liquid.

This means that the permeability is increased when more of the molecules have escaped into the water surrounding them there will be much more molecules in the way of the light to block it, this is why the amount of light transmitted is decreased as the temperature is increased.

So if the temperature is kept at room temperature as I tested in my preliminary then fewer pigment molecules should be able to escape because the cell membrane is intact and haven’t been destroyed and so the pigment has no hole from which it can escape from and so the rate of permeability won’t increase form this.

Evaluation:

Overall the data I obtained in this experiment was pleasing and tends to follow the trend in the graph. There were a few anomalies. As I conducted two experiments instead of three it is much more difficult for me to say which result on the graph is the anomaly. If I had done three experiments then I could have seen which two results were more inline with each other and this would have determined the anomaly. If I had done three experiments then the results would have been much more reliable but I did not have time to do a third experiment. The anomalies that appeared on the table of results and that were masked on my graph were at the 50 ºC mark this tells me that it is above this point that the proteins in the membrane begin to break down and denature because as the temperature is increased from 50 ºC to 60 ºC the reading on the colourimeter is at 1%. Apart from this anomaly the other points fit the trend it is only 50 ºC that does not fit the trend as it decreases.
The results that I obtained in the two experiments seemed to be accurate for example at 30 ºC the first test tube reading was at 40% and the second test tube was at 43 this shows me that this is quite accurate as the two results only had a very short percentage difference between. As I did only two experiment I can not say whether any of these are an anomaly but looking at these two results it seem to me that they are not anomalies as they do follow the trend of decreasing percentage.

There are some sources of experimental error these could account for some of the anomalies that I had come up in my experiment. One source of error which did not talk about in my plan was the need to extract the beetroot from the distilled water gently with forceps. Rough use of the forceps could have pierced the cell membranes and the tonoplast membranes surrounding the vacuole containing the red pigment, this would cause the red pigment to be instantaneously squeezed out into the distilled water. The result of this could be more pigment being released into the distilled water then should have done.

I could have measured different mounts of water each time – having more water will dilute the pigment of the beetroot more then other test tubes and having less water will mean that the colour will be darker then it should have been this could lead to anomalies in the results and give inaccurate results.
I could have left the test tubes in the water bath longer or shorter then 5 minutes – this could mean that it is not a fair experiment as the test tube is not left for enough time and the pigment that should have been released was either not enough released or too much released this could mean inaccurate reading on the colourimeter.
I might have cut different sizes of beetroot each time – if a beetroot is cut too big them too much pigment could be released as the piece of beetroot is bigger but if it is too small then less pigment is released then should have been this could mean that inaccurate reading on the colourimeter.
I might not have drained the excess fluid off the beetroot after I cut it – if the beetroot is not drained properly then pigment is released from the beetroot that has been damaged when it is cut this means that excess beetroot will be released into the distilled water this would mean that the reading is inaccurate some temperature will have more pigment released then should have been and then leading to a anomaly.
The temperature of the water might have altered after I put the test tubes in if this happens then the water will not be heated to the right temperature and the right amount of pigment might not be released meaning that a lower reading taken rather then the real amount that it should have been.

To improve the reliability of the result I repeated the experiment again this would have improved the accuracy if I had maybe changed the apparatus by using a different colourimeter which measures light over a much narrower range of wavelength. the stop clock that I used could have counted down this would have meant that it would have stopped on exactly 5 minutes and I could taken the test tubes out in better time. The apparatus which I used was what was available and so using some different apparatus was not really possible and also I did not have time to find and change the apparatus.

When timing I used a stop clock it was timed for 5 minutes but I could have let the clock run a bit over the time or under the time and so the beetroot was not taken out at the exact time each time. This could mean that as the beetroot was left in the heated water for longer more pigment could have leaked out this looks to be the case at 50ºC as it does not follow the trend in the graph it is even with 40ºC on the first transmission and on the second transmission it is above 40 ºC this is an anomaly and the cause seems to be that the beetroot was left in the heated water for too long. Hence more pigment being released then should have been released.

The factors affecting the rate of permeability in a cell membrane?