Chapter 2 Cell Structure and function

Preparing and studying slides of plant cells Materials - An Onion bulb, iodine and distilled water Apparatus Glass slide, cover slips, a light microscope, forceps, glass dropper, scalpel, mounting needle , filter paper

Procedure A piece of onion scale is cut from an onion bulb using a scalpel. The transparent epidermis from the inner surface of the onion is gently peeled off using a pair of forceps. Put a drop of water onto the middle of the slide and place the epidermis on the drop of water. With the help of a mounting needle, cover the specimen with a cover slip lowering it slowly. Add a drop of iodine solution onto one side of the cover slip. Place a filter paper at the opposite end to allow iodine to spread through the epidermis Excess iodine solution is absorbed using another filter paper. The slide is examined.

Observation

Conclusion A typical plant cell has a fixed shape , cell wall, nucleus, cytoplalsm and a large vacuole.

Preparing and studying slides of animal cells Materials cheek cells, methylene blue solution, iodine solution and distilled water Apparatus Glass slide, cover slips, a light microscope, glass dropper, mounting needle , filter paper, toothe pick

Procedure A blunt end of a clean toothpick is used to gently scrape the inner lining of the cheek. The scrapings are mounted in a drop of water on a clean glass slide. The specimen is the slowly covered with a cover slip using a mounting needle A drop of methylene blue is laced onto one side of the cover slip. A filter paper is then placed at the opposite side of the cover slip to draw the solution across the specimen. The slide is observed. The steps are repeated using iodine solution. Observation

Conclusion A typical plant cell does not have a fixed shape. But has nucleus and cytoplalsm

Chapter 3 Movement of subtances across plasma membrane Studying the movement of subtances across the plasma membrane Problem Statement What factor influences the diffusion of substances across a semi permeable membrane Variable Manipulated- Size of solute molecules Responding- Colour of solution in the visking tube Constant- Surrounding temperature

Hypothesis The diffusion of molecules through a semi permeable membrane is based on the size of the molecules Material- Benedicts solution , 1% Starch suspension, Iodine solution, 30% glucose solution, visking tubing, cotton thread Apparatus- Test tubes, beakers, Bunsen burners

Procedure A visking tubing is soaked in the water for 5 minutes to soften it. One end of the visking tubing is tied firmly with a piece of cotton thread to prevent leakage. The visking tubing is filled with 15ml of glucose solution and 15ml of starch suspension. other end of the visking tubing is tied with another piece of cotton thread. The colour is recorded. The outer surface of the visking tubing is rinsed with water. 400ml of disttiled water and 15 ml of iodine are mixed in a beaker. The colour is recorded. The visking tubing is immersed in the beaker solution for 40 minutes. After 40 minutes, the visking tubing is transferred to a dry beaker. The colour of the solutions in the visking tubing and beaker is observed and recorded. Both solutions are tested for the presence of glucose using benedicts solution. 2ml of each solution are poured into the separate tubes and 1 ml of benedicts is then added to each test tube. The solution is heated in a boling water bath for 5 minutes and the change in colour is recorded.

Results Visking tubing 15 ml of glucose + 15 ml of starch suspension 400ml of distilled water + 15 ml of iodine solution Colourless Blue black Positive. A brick red precipitate is formed Positive. A brick red precipitate is formed

Beaker

Yellow

Yellow

Conclusion The diffusion of molecules across the semi permeable membraneis based on the size of molecules. The Hyphothesis is accepted.

Chapter 3 Studying the osmosis using osmometer Problem Statement What substances can diffuse across a semi permeable membrane Variable Manipulated- Time Responding- Increase in level of sucrose solution Constant- Surrounding temperature and concentration of sucrose

Hypothesis Water molecules can diffuse thorugh a semi permeable membrane Material- 30% sucrose solution, distilled water, visking tubing, cottong thread Apparatus- Retort stand, Cappilary tube, a ruler, marker pen, scissors, 250ml beaker, syringe, stopwatch

Procedure A visking tubing about 8cm long is cut The visking tubing is soaked in water for about 5 minutes to soften it. One end of the tube is tightly tied with a piece of cotton thread. The visking tubing is filled with 30% sucrose solution using a syringe. The other end of the visking tubing Is tied to the bottom of the capillary tube. The outer surface of the visking tubing is rinsed with distilled water The capillary is the clamped vertically to the retort stand. The visking tubing is then immersed into a beaker filled with distilled water. The initial level of the sucrose solution is marked with a marker pen at the beginning of the experiment. The level of sucrose solution is then marked and recorded every 10minutes for a duration of 40 minutes .

Results Time The increase in height of sucrose solution from the initial level 0 0 10 30 20 60 30 90 40 108

Conclusion Small molecules like water can easily duffuse through semi permeable membranes such as the visking tubing. The increase in the level of the sucrose solution is due to the process of osmosis. Hyphothesis is accepeted.

Effects of hypotonic, hypertonic and isotonic solutions on animal cells. Aim To study the effects of hypotonic, hypertonic and isotonic solutions on animal cells. MaterialsFresh chicken blood, 0.15 M sodium chloride solutions, 0.50 M sodium chloride solutions, Distilled water, Filter paper Apparatus- Glass slides, Cover slips, Filter papers, Light microscope, Test tubes, Beakers, Dropper Procedure 4 slides were labeled as A, B, C and D. A drop of blood was dropped on slide A and covered with a cover slip and is observed under a light microscope. A drop of distilled water was dropped on slide B and covered with a cover slip. A drop of blood was put at one side of the cover slip. The blood is allowed to spread by placing a filter paper on the opposite side. The slide was observed under a light microscope . Steps 3 and 4 were repeated using 0.15 and 0.50M sodium chloride solutions on slide C and D respectively.

Observations Slide A Observation Red blood cell have biconcave shape Discussion Shape is normal The red blood cell are placed in a hypotonic solution Water diffuses into the red blood cells by osmosis causing cells to swell and burst The contents are released to the surroundings Haemolysis occurs The concentration is isotonic to the cytoplasmic fluid No net movement of water in and out The concentration is hypertonic to the cytoplasmis fluid of red blood cells There is a net of outflow of water from the cells by osmosis. The cell have become shrivelled.

B(red blood cells in distilled Cells swell and burst water)

C(red blood cells in 0.15m sodium chloride)

Red blood cells retain their shape and size

D(redblood cells in 0.50 sodium chloride)

Red blood cells shrink and crenate

Conclusion Observation on animal cells in above .

Effects of hypotonic, hypertonic and isotonic solutions on plant cells. Aim To study the effects of hypotonic, hypertonic and isotonic solutions on plant cells. Material Onion bulb, 0.5 M sucrose solutions, 1.0 M sucrose solutions, Distilled water Apparatus Light microscope, Glass slides, Cover slips, Razor blade / scalpel, A pair of forceps, Mounting needle, Filter papers Procedure Four slides are prepared and labelled A,B,C and D A thin layer of epidermal cell of onion was stripped using a pair of forceps and mounted on slide A. A drop of distilled water was dropped on it and covered with a cover slip. The slide was examined under a light microscope using low power and then high power. The epidermal cells are drawn and labelled. A drop of 0.5M sucrose solution is placed at one side of the cover slip. The solution is allowed to spread by placing a filter place at the opposite end of the cover slip. The epidermal cells are then observed under the microscope. The epidermal cells are drawn and labelled. Steps 6 and 7 are repeated by substituting 0.5 m with 1.0 m sucrose solution

Observations Slide A(plant cells in distilled water) Observation Cells appear turgid. The vacuoles swell up into larger sizes. Discussion The distilled water is hypotonic to the cell sap of the epidermal cells The net flow of water by osmosis into the vacuoles cause them to swell At this point, the cells are highly turgid. The vacuoles and cytoplasm exert an outward force against the plasma membrane and cell wall. The cells do not burst because of their rigid cell walls. 0.5M is isotonic to the cell sap The cells do not lose or gain water. 1.0M is hypertonic to the cell sap of the epidermal cells. There is net outflow of water from the vacuoles by osmosis. The plant cells have undergone plasmolysis.

B(plant cells in 0.5M sucrose solution)

The shape and the structure remains the same. The epidermal cells looks flaccid, the vacuoles loses water and becomes smaller. The cytoplasm shrinks and the plasma membrane pulls away from the cell wall.

C(plant cells in 1.0M sucrose solution)

Conclusion Observation on plant cells in above .

Concentration of an external solution which is isotonic to the cell sap of a plant Aim To determine the concentration of an external solution which is isotonic to the cell sap of a plant Problem statement What is the concentration of an external solution which is isotonic to the cell sap of a plant? Hypothesis When the external solution is isotonic to the cell sap of the plant, there is no net gain in the size of the plant cell Variable Constant : Manipulated : Responding

Material Potato, distilled water, filter paper and sucrose solutions with concentration of 0.1M, 0.2M, 0.3M, 0.4M, 0.5 and 0.6M Apparatus Petri dishes, knife, forceps, ruler and cork borer

Procedure 7 petri dishes were labeled as A G. Petri dish A was filled with distilled water Each petri dish was filled with different solution according to the table below: Petri dish B C D E F G Sucrose solution (M) 0.1 0.2 0.3 0.4 0.5 0.6 A cork borer was used to bore 21 potatoes in cylindrical strips. Each strip was cut to a length of 5cm. 3 strips were placed in each petri dish and left for an hour. After an hour, the strips were removed. The potato strips were wiped dry and the length of each strip was measured. Measurements were recorded. A graph of elongation of potato strip against the concentration of sucrose solution was plotted. Observations

Differentiating between reducing and non-reducing sugars

Material Benedicts solution, dilute hydrochloric acid, sodium hydrogen carbonate powder, 10% glucose solution and 10% sucrose solution. Apparatus Test tubes, beaker, dropper, tripod stand, wire gauze, Bunsen burner Procedure 2ml of benedicts solution is added to an equal volume of glucose solution in a test tube. The mixture is shaken and then heated by placing the test tube in a beaker of boiling water for a few minutes. The colour of the precipitate that is formed is observed. The test is repeated by replacing the glucose solution with sucrose solution. If the sucrose solution produces a negative result, the experiment is continued, using another sample of sucrose solution. This time the sucrose solution is added to 1ml of dilute hydrochloric acid and bolied for a few minutes. The mixture is then left to cool A small amount of sodium hydrogen carbonat is added ti neutralise the acid. Steps 1 tlo 3 are repeated when the effervescence stops. The colour of the precipitate formed is observed and recorded in the table below. Observations Sugar Glucose Sucrose (without HCL) Observation A bric red precipitate is formed The colour of the benedicts solution remains unchanged A brick red precipitate is formed Inference Glucose is a reducing sugar Sucrose is a non reducing sugar

Sucrose (bolied w HCL)

A positive result with Benedicts solution can only be obtained if the sucrose solution is first bolied with HCL

Conclusion Reducing sugar produce red brick precipitate when tested with Benedicts solution.

Effect of temperature on the activity of enzyme amylase on starch Aim To study the effect of temperature on the activity of enzyme amylase on starch Problem statement What is the effect of temperature on the activity of enzyme amylase on starch? Hypothesis The rate of the activity on salivary amylase on starch increase with the increase in temperature until it reaches the optimum temperature of 37. Variables Constant :Volum of saliva, volume and concentration of starch suspension, Ph of medium Manipulated : Temperature of medium Responding : Rate of enzymatic reaction Materials 1 % starch solution, iodine solution, ice, distilled water Apparatus 5 beakers, 10 test tubes, syringe, glass rod, dropper, white tile, thermometer, Bunsen burner, tripod stand, wire gauze, stop watch Procedure 5 ml of saliva was collected in a beaker after rinsing ones mouth with clean water. 5 ml of distilled water was added into the beaker to dilute the saliva. 5 test tubes were labeled as A, B, C, D and E and each of it were filled with 2 ml of saliva. 5 test tubes were labeled as A1, B1, C1, D1 and E1 and each of it were filled with 5ml of starch solution. Test tubes A and A1, B and B1 , C and C1, D and D1, E and E1 areimmeredsed respectively into five different water baths with temperature which are kept constant at 0, 28 37 ,45 and 60 The test tubes are left for 5 minutes A drop of iodine solution Is placed into each groove of the white tile. After five minutes, the starch suspension in test tube A1 is poured into the saliva solution in A. The mixture is stirred using a glass rod. A stopwatch is started immediately

Use a dropper to remove a drop of mixture from the test tube and place It into iodine solution in the first groove. ( counted as zero minute) The iodine test is repeated every minute 10 minutes. The dropper is rinsed in a beaker distilled water after each sampling. Record the time taken for the completion of the hydrolysis of starch that is when the mixture gives a negative iodine test (no longer turns blue black) The test tubes containing mixture are kept in their respective water baths throughout experiment. Steps are repeated for test tubes B,C,D,E

Observation Test Tubes Temperature Time take for Hydrolysis of starch To be completed Not completed after 10 minutes 2 1 3 Not completed after 10 minutes Rate of enzymatic reaction 1/t (minute) 0 0.50 1 0.33 0

A B

0 28 37

C 45 D E 60

Conclusion Changes in temperature affect the activity of salivary amylase on starch. Salivary amylase is inactive at 0 and denatures at 60 The rate of reaction catalysed by the salvary amylase is highest at 37 which is optimum temperature. Hyphothesis is accepted.

Effect of pH on the enzyme activity Aim To study the effect of pH on the enzyme activity Problemstatement What is the effect of pH on the enzyme activity Hypothesis Pepsin works best in acidic medium Variables Constant : Volume and concentration of albumen suspension, volume and concentration of pepsin solution and temperature of medium. Manipulated : pH of Medium Responding Rate of enzymatic reaction Materials Albumen suspension, 1% pepsin solution, 0.1M hydrochloric acid, 0.1 M sodium hydroxide solution, pH paper, distilled water Apparatus Test tube, syringe, thermometer, stopwatch, beaker, Bunsen burner tripod stand, filter funnel,wire gauze Procedure Prepare and albumen suspension by mixing egg white with 500ml of ditilled water. Boil the suspension and leave it to cool. Discharge large particles using glass wool. Label the three test tubes P,Q and R 5ml of albumin suspension was poured into each test tube, P, Q and R. The following solutions were added into each test tube. Test tube Solution P 1 ml hydrochloric acid + 1 ml pepsin solution Q 1 ml sodium hydroxide + 1 ml pepsin solution R 1 ml distilled water + 1 ml pepsin solution A piece of pH paper was dipped into each test tube. The pH value was recorded. All test tubes were placed into a beaker of water bath at 370C for 20 minutes. The change in the content of the test tube was observed and recorded.

Observation

Test tubes

Ph

Mixture

Beginning

End

Cloudy

Clear

Cloudy

Cloudy

Cloudy

Cloudy

Conclusion The activity of pepsin is affected by the Ph of its medium. An acidic medium pH3 is the most suitable medium for pepsin to function effieciently The hyphothesis is accepted

Effects of substrate concentration on the activity of salivary amylase Problem statement What is the effect of substrate concentration on the activity of salivary amylase Hyphothesis The rate of enzymatic reaction increases with the increase in substarte concentration until it reaches a maximum rate. Constant Manipulated: concentration of starch Responding Time taken for hydrolysis of starch to be completed Constant Enzyme concentration, Temperature and pH of medium Material Starch suspensions at various concentrations, 0.1% amylase or saliva suspension, iodine, distilled water Apparatus test tubes, syringe, dropper, glass rod, white tile, wire gauze, measuring cylinder stopwatch Procedure Six test tubes were labeled as A, B, C, D, E and F. 4ml of starch suspension of various concentrations are poured into the following test tubes A,B,C,D,E,F Test tubes are immeredsed respectively in water bath of 37 A drop of iodine solution is placed into each groove of the white tile. 1ml of 0.1% amylase is added to test tube A using a syringe The stopwatch is started immediately. The contents are stirred with a glass rod. A drop of mixture is tested with the iodine solution on the white tile. This step is repeated every 30 second interval until the mixture stops turning blue black. The time take for the starch to be completely hydrolysed is recorded. Steps were repeated for test tubes B, C, D, E and F. At every sampling, the dropper must be rinsed with clean distilled water.

Observations

Test Tube

Concentration of starch suspension

Time taken for hydrolysed of starch to be completed

Rate of Reaction = (substrate concentration/time) (% minute)

0.1

240

4.0

0.025

0.2

240

4.0

0.050

0.3

240

4.0

0.075

0.4

240

4.0

0.100

0.5

300

5.0

0.100

0.6

360

6.0

0.100

Conclusion The rate of enzymatic reaction increases with the increase in substrate concentration until it reaches maximum rate. The hyphothesis is accepted.

Effect of enzyme concnetration on the activity of salivary amylase Problem statement What are the effects of enzyme concentration on the activity of salivary amylase Hypothesis The rate of enzymatic reaction increase with the increase in enzymatic concentration as long as there are no factors limiting the rate of reaction. Variables Constant : Substrate concentration, Temperature and Ph of medium Manipulated : Concentration of enzyme Responding : time taken for the hydrolysis of starch to be completed Materials 1 % starch solution, 0.8% amylase or saliva suspension ,iodine solution, distilled water Apparatus test tubes, syringe, dropper, glass rod, white tile, wire gauze, measuring cylinder stopwatch Procedure Six test tubes were labeled as A, B, C, D, E and F. The test tubes contain the following mixtures A B C D E F : 0.5ml of 0.8% amylase + 2.5ml distilled water : 1.0ml of 0.8% amylase + 2.0ml distilled water : 1.5ml of 0.8% amylase + 1.5ml distilled water : 2.0ml of 0.8% amylase + 1.0ml distilled water : 2.5ml of 0.8% amylase + 0.5ml distilled water : 3.0ml of 0.8% amylase

Test tubes are immersed respectively in water bath of 37 A drop of iodine solution is placed into each groove of the white tile. 4ml of 1% of starch suspension is added to test tube A using a syringe The stopwatch is started immediately. The contents are stirred with a glass rod. A drop of mixture is tested with the iodine solution on the white tile. This step is repeated every 30 second interval until the mixture stops turning blue black. The time take for the starch to be completely hydrolysed is recorded. All results are recorded and tabulated

Observation Test Tubes Concentration of amylas Time take for Hydrolysis of starch To be completed Seconds minutes 330 150 90 60 60 60 5.5 2.5 1.5 1.0 1.0 1.0 Rate of enzymatic reaction 1/t (minute)

A B C D E F

0.17 0.33 0.50 0.67 0.83 1.00

0.18 0.40 0.67 1.00 1.00 1.00

Conclusion The rate of reaction increases with the increase in enzyme concentration until a certain concentration of enzyme is achieved. The hyphothesis is accepted.

Conclusion Changes in temperature affect the activity of salivary amylase on starch. Salivary amylase is inactive at 0 and denatures at 60 The rate of reaction catalysed by the salvary amylase is highest at 37 which is optimum temperature. Hyphothesis is accepted.