THE EFFECT OF TEMPERATURE TO THE RESPIRATION RATE OF BEAN SPROUT

NUR RIZQI AKHFIANI (093204028) INTERNATIONAL CLASS OF BIOLOGY EDUCATION 2009

CHAPTER 1 PREFACE 1.1 Background Although we often think of respiration in terms of humans and breathing, it is important to note that it is one of the 7 characteristic processes shared by all living organisms, so it must have a universal definition: "processes leading to, and including the chemical breakdown (oxidation) of food materials to provide energy for life" (Beth, Asher.2010). It is important to realise that green plants respire at all times, like all living organisms. However, they are unique in that they can also make their own food by photosynthesis . As a by-product of this process, they produce a waste substance, oxygen, which they release from the cells of their leaves into the air. Of course, they may re-use some of this oxygen in aerobic respiration, so some people wrongly assume that plants do not need oxygen, or do not respire in the same way as other living organisms. What happens in these plants is merely a question of balance between the processes of photosynthesis and respiration. Seeds are dormant stages of living organisms, and contain embryo plants, ready to grow when conditions are right. Most of the time, they appear to be doing nothing much, but they are respiring only slowly because they do not need much energy, and need to conserve the food reserves they contain. However, before they start to grow into plants (and continue the life cycle by flowering and producing seeds again), seeds must germinate. In order to do this, seeds absorb water which they need in order to mobilise their food reserves using enzymes (basically the same process as digestion in animals), then they speed up their rate of respiration quite dramatically. They then use the energy released in order to sprout roots to absorb more water and minerals, and grow a shoot which takes the leaves above ground, so as to make food by photosynthesis. Of course this usually happens under the ground, but it is not necessarily so.

The respiration rate of plant is affected by some environmental factors; such as: temperature, humidity, pH, and so on. For this activity, we will observe about the effect of temperature to the repiration rate of bean sprout. 1.2 Problem Based on the background above, the problem which we got is How does the effect of temperature to the respiration rate of bean sprout? 1.3 Purpose Based on the problem above, the problem which we got is observing the effect of temperature to the respiration rate of bean sprout.

CHAPTER 2 THEORY Definition and The Process of Plants Respiration Respiration is a catabolic reaction that breaks down glucose and other nutrients. Germinating seeds take in oxygen from the surrounding air. The oxygen is used to convert nutrients stored in the seeds endosperm into energy that the seed uses to sprout. Enzymes activated when the seed imbibes (takes in water) make the energy conversion possible. Carbon dioxide is given off as a waste product during respiration. Plant respiration is the mechanism by which energy stored in the form of glucose, a product of photosynthesis, is released for use in plant metabolism. Photosynthesis uses light energy from the sun along with water and carbon dioxide to produce glucose, or sugar, and releases oxygen into the atmosphere. Respiration releases carbon dioxide (Asher,Beth.2010). Respiration begins when a seed absorbs water. The enzymes inside seeds have to be in suspension to function. Activated enzymes start respiration and use organic matter stored in the seed during respiration to make ATP molecules needed for growth. Respiration continues in the seed after the sprout has emerged until the seeds endospermic material is used up and the cotyledons have produced the sprout (Schlesinger, 1997). Soaking seeds before planting has been a common practice and seeds need water uptake to begin respiring. Dry seeds do respire, but at an extremely low, nearly undetectable rate. Stored gases in seeds are released when water is first taken up. This is not true respiration. True respiration starts when the seeds testa (seed coating) is softened by water and enough water passes the softened coating to activate seed enzymes. The seed swells and begins to consume oxygen (Gifford, 1994; Amthor, 1995). Respiration goes in spurts, like seedling growth. A root and a shoot develop as the result of the biochemical reactions in the seed. Seed respiration levels off after this point. When the root and shoot begin the next growth spurt leading up to true leaves, the respiration rate increases again to fuel the growth (Amthor, 1991).

Respiration process begins with the arrest of O2 from the environment. Gas transport processes in plants as a whole takes place in diffusion. Oxygen used in respiration entry into any plant cell by diffusion through intercellular spaces, cell wall, cytoplasm and cell membrane. Similarly, the CO2 produced by respiration will diffuse out of the cell and into the space between cells. This is because the plasma membrane and the protoplasm of plant cells are highly permeable to both gas. After taking O2 from the air, O2 and then used in the respiration process with several stages, among which glycolysis, oxidative decarboxylation, Krebs cycle and electron transport. The first stage is glycolysis, which stages the conversion of glucose into two molecules of pyruvic acid (atom C3), this event took place in the cytosol. Pyruvate acid produced will then be processed in the stage of oxidative decarboxylation. Besides glycolysis also produces 2 molecules of ATP as energy, and 2 NADH molecules that will be used in electron transport. In anaerobic conditions, glycolysis results Pyruvate acid is converted into carbon dioxide and ethyl alcohol. This conversion process is catalyzed by enzymes in the cytoplasm. In anaerobic respiration the amount of ATP produced only two molecules for every one molecule of glucose, these results differ greatly with ATP generated from the overall results of aerobic respiration is 36 ATP. The second stage of respiration is oxidative decarboxylation, namely the conversion of pyruvic acid (C3 atom) into acetyl CoA (atom C2) with the release of CO2, this event took place in the cytosol. Acetyl CoA is produced will be processed in the Krebs cycle. Other results of NADH that will be used in electron transport. The next stage is the citric acid cycle (Krebs cycle) that occurs in the matrix and inner mitochondrial membrane, namely the processing stages of acetyl CoA by the citric acid compound as the compound which was first formed. Some of the compounds generated in this phase, including the one molecule of ATP as energy, one molecule of FADH and three molecules of NADH to be used in electron transfer, as well as two molecules of CO2. Last stage is the transfer of electrons, ie a series of reactions involving the career system of electrons (electron carriers). This process occurs in the inner mitochondrial membrane. In this reaction electrons are transferred in a series of redox reactions and assisted by the enzyme cytochrome, quinone, pyridoxine, and flavoprotein. This electron transfer reaction will produce H2O (I Komang Jaya Santika Yasa, 2009). The process of respiration can be described as follows:

1.) glycolysis: Glucose 2 pyruvic acid + 2 NADH + 2 ATP 2. Krebs Cycle: 2 acetyl pyruvate acetyl CoA 2 + 2 CO2 + 2 NADH + 2 ATP 2 acetyl CoA 4 CO2 + 6 NADH + 2 FADH2 3. Electron transport chain: 10 NADH + 5O2 10 10 NAD + + H2O + 30 ATP 2 FADH2 + O2 2 FAD + 2 H2O + 4 ATP Thus, the total energy generated from the process of respiration is 38 ATP. (Danang, 2008).

Function of Plants Respiration (Davey et al., 2004). During respiration, glucose is broken down to become products that will fuel other processes that the plant will use. Respiration is a multi-stage process that can take different pathways depending on the presence or absence of oxygen. Glycolysis breaks glucose into two molecules initially, in the cytoplasm, or the main body of the cell enclosed within the plant cell wall. The presence or absence of oxygen determines how the process will progress. If oxygen is present, products from glycolysis will be used within the mitochondria, or energy centers, of the cell, to continue respiration. If oxygen is not present, fermentation, a less efficient use of energy, occurs. Effects of Plants Respiration (Davey et al., 2004). During photosynthesis, plants store energy captured during the day, the light reaction, to fuel the process of creating sugar chiefly at night, during the dark reaction of photosynthesis. As the name implies, the dark reaction is independent of the presence of light. Glucose is produced. Glucose will in turn fuel respiration.

Factors Affecting Respiration Rate 1. Availability of substrate Respiration dependent on substrate availability. Starch content of plants, fruktan, or low sugar, do respiration at a low rate. Scarcity of food plants carry on respiration of sugar often faster when sugar is provided. Even the leaf respiration rate is often much faster soon after sunset, when the high sugar content as compared to when the sun rises, while a lower sugar content. In addition, the shaded leaf or leaves the bottom usually slower respirated than the upper leaves are exposed to more light. If this does not happen, then the lower leaves will die sooner. Differences due to the sugar content may not equal of rate of photosynthesis which causes a lower respiration rate in the shaded leaves. (Salisbury & Ross, 1995) 2. Availability of oxygen The availability of oxygen will affect the rate of respiration, but the magnitude of these effects are different for each different species and even between the same organs in plants. Normal fluctuations in the oxygen content of air is not much affect the rate of respiration, because the amount of oxygen required for berespirasi plants is much lower than the available oxygen in the air. (I Komang Jaya Santika Yasa, 2009) 3. Temperature Effect of temperature factors for respiration rate of plants is strongly associated with factors Q10, which is generally the reaction rate of respiration will increase for every increase in temperature of 10 C, but this depends on each species. For most parts of the plant and plant species, Q10 respiration is usually 2.0 to 2.5 at temperatures between 5 and 25 C. When the temperature is increased further to 30 or 35 C, respiration rate remained elevated, but more slowly, so Q10 begin to decline. An explanation of the decline in Q10 at high temperatures is that the rate of penetration of O2 into the cell through the cuticle or periderma start inhibit respiration during chemical reactions take place quickly. Diffusion of O2 and CO2 are also accelerated with increasing temperature, but the Q10 for the physics of this process is only 1.1, so the temperature does not significantly accelerate the diffusion of solution through the water. Increasing the temperature to 40 C or more, in fact decreased respiration rate, especially when plants are in this state in a long time. It seems that

the enzymes necessary to quickly start having denaturation at elevated temperatures, preventing an increase in metabolic proper place. In pea sprouts, increase in temperature from 25 to 45 C initially rapidly increasing respiration, but after two hours of its speed began to decrease. Possible explanation is that a period of two hours is long enough to damage some respiratory enzymes. (Salisbury & Ross, 1995) 4. Type and Age of Plant Each plant species has metabolsme differences, thus the need for berespirasi plants will be different in each species. Young plants showed a higher respiration rate than older plants. Similarly the plant organ that is in its infancy. (I Komang Jaya Santika Yasa, 2009).

CHAPTER 3 RESEARCH METHOD 3.1 Kind of Research The kind of research in this laboratory activity is experimental, because there is some treatment to solve the problem formula and there are variables used, such as manipulation variable, control variable, and response variable. 3.2 Variables Variables which is used in this laboratory activity are: 3.2.1 Manipulation variables : The temperature (incubation temperature: 37C; room temperature: 32C). 3.2.1 Control variables : Weigh of bean sprout (5gr), Volume of NaOH 0.5M (30mL), Kind of bean sprout (green bean), Amount droping of PP solution (two drops), Age of bean sprout (2 days), Volume of BaCl2 (2.5mL),distance between hanging bean sprout with NaOH solution in the Erlenmeyer tube (half of tube), time of saving (24hours). 3.2.3 Response Variables 3.3 Tools and Materials 3.3.1 Tools Erlenmeyer tube 250mL Kasa silk Rope Pipettes Transparent plastic Rubber Balance Burret 6 pieces As enough As enough As enough 6 pieces 6 pieces 1 piece 1 piece : The respiration rate of bean sprout (ml/hour)

 Stative and clem Cut

1 piece 1 piece

3.3.2 Materials Bean sprout age 2 days NaOH solution 0.5M HCl solution 0.5N BaCl2 solution 0.5N Phenolftalin solution 3.4 Procedures 1. Prepare 6 pieces of Erlenmeyer tube, and fill it with 30mL of NaOH 0.5M. 2. Measure 5gr of bean sprout for each Erlenmeyer tube. Need 4 erlenmeyer tube that is contained of bean sprout. 3. Covering the each 5 gr of bean sprout with kasa silk, and tie it using rope. 4. Entering the covered bean sprout into Erlenmeyer. 5. Hanging the covered bean sprout using rope as half of height of tube. The Erlenmeyer which is filled by bean sprout is 4 tubes. Two tubes for placed in room temperature, two others placed in incubator temperature. Covering the Erlenmeyer tubes hole with transparent plastic and rubber. 6. The last two of Erlenmeyer tube is not filled by bean sprout, only filled by NaOH 30mL as the control. 7. Put one of control tube, and two of tube filled bean sprout into incubator (37 C). wait until 24 hours. 8. Put one of control tube, and two of tube filled bean sprout into usual room (32 C). Wait until 24 hours. 9. After 24 hours. Doing titration. Firstly, fill the burret with HCl solution until the scale of burret shown 0 number 10. Take 5mL of NaOH solution in Erlenmeyer. Then, added by 2.5 mL of BaCl2 11. Drop it with 2 drops of PP till the solution is young red. 12. Titration the red solution with HCl in the burret. Titrate it drop by drop of HCl until the color of solution is changing to be clear (the red color is disappear). 13. Doing the titration until 6 tube of erlenmyer is titrated. Note, the volume of HCl that used for changing the color of solution.

3.5 Design Experiment

1. Prepare 6 pieces of Erlenmeyer tube, and fill it with 30mL of NaOH 0.5M.

2. Measure 5gr of bean sprout for each Erlenmeyer tube. Need 4 erlenmeyer tube that is contained of bean sprout.

3. Covering the each 5 gr of bean sprout with kasa silk, and tie it using rope.

4. Entering the covered bean sprout into Erlenmeyer.

5. Hanging the covered bean sprout using rope as half of height of tube. The Erlenmeyer which is filled by bean sprout is 4 tubes. Two tubes for placed in room temperature, two others placed in incubator temperature. Covering the Erlenmeyer tubes hole with transparent plastic and rubber.

6. The last two of Erlenmeyer tube is not filled by bean sprout, only filled by NaOH 30mL as the control.

7. Put one of control tube, and two of tube filled bean sprout into incubator (37 C). wait until 24 hours.

8. Put one of control tube, and two of tube filled bean sprout into usual room (32 C). Wait until 24 hours.

9. After 24 hours. Doing titration. Firstly, fill the burret with HCl solution until the scale of burret shown 0 number.

10. Take 5mL of NaOH solution in Erlenmeyer. Then, added by 2.5 mL of BaCl2

11. Drop it with 2 drops of PP till the solution is young red.

13. Titration the red solution with HCl in the burret. Titrate it drop by drop of HCl until the color of solution is changing to be clear (the red color is disappear).

12. Doing the titration until 6 tube of erlenmyer is titrated. Note, the volume of HCl that used for changing the color of solution.

CHAPTER 4 DATA AND ANALYSIS 4.1 Data After did the experiment, we got the problem as follow: Table of Data Table 1.The Effect of Temperature to The Respiration Rate of Bean Sprout Measurement No. Result from Room Temperature 32C Control 1st bean sprout 2nd bean sprout 1. Volume HCl (mL) Volume of 2. NaOH which isnt binded by CO2 (mL) Volume of 3. NaOH which is binded by CO2 (mL) Volume of 4. CO2 that is respiration product (mL) Respiration 5. Rate (mL/hour) 0.125 0.2375 3 5.7 21 25.2 22.8 19.2 25.2 24.6 9.0 4.8 7.2 10.8 4.8 5.4 1.5 0.8 1.2 1.8 0.8 Incubator Temperature 37C Control 1st bean sprout 2nd bean sprout 0.9

Graphic1.The Relationship of Temperature to The Respiration Rate of Bean Sprout

4.2 Analysis of Data Based on the data above, we can see the relationship of temperature in room to the respiration rate of bean sprout. In incubation room with temperature 37 C has respiration rate 0.2375 mL/hour. In room temperature with 32 C has respiration rate 0.125 mL/hour. So, the higher of temperature, the faster of respiration rate but just until the optimum temperature to do respiration. In usual room temperature 32 C, the volume of HCl used for titration of the 5mL of NaOH (until the pink color is being colorless/white) is 1.5mL in control Erlenmeyer, 0.8mL in 1st Erlenmeyer of bean sprout, and 1.2 mL in 2nd Erlenmeyer of bean sprout. The volume of NaOH (from 30mL of NaOH volume) which isnt binded by CO2 is 9.0mL in control Erlenmeyer, 4.8mL in 1st Erlenmeyer of bean sprout, and 7.2 mL in 2nd Erlenmeyer of bean sprout.The volume of NaOH (from 30mL of NaOH volume) which is binded by CO2 is 21mL in control Erlenmeyer, 25.2mL in 1st

Erlenmeyer of bean sprout, and 22.8 mL in 2nd Erlenmeyer of bean sprout. The volume of CO2 that as respiration product is 3mL. And the respiration rate is the comparison between volume CO2 as the respiration product with the time of saving (24 hours), as much 0.125mL/hour. In incubation temperature 37 C, the volume of HCl used for titration of the 5mL of NaOH (until the pink color is being colorless/white) is 1.8mL in control Erlenmeyer, 0.8mL in 1st Erlenmeyer of bean sprout, and 0.9 mL in 2nd Erlenmeyer of bean sprout. The volume of NaOH (from 30mL of NaOH volume) which isnt binded by CO2 is 10.8mL in control Erlenmeyer, 4.8mL in 1st Erlenmeyer of bean sprout, and 5.4 mL in 2nd Erlenmeyer of bean sprout. The volume of NaOH (from 30mL of NaOH volume) which is binded by CO2 is 19.2mL in control Erlenmeyer, 25.2mL in 1st Erlenmeyer of bean sprout, and 24.6 mL in 2nd Erlenmeyer of bean sprout. The volume of CO2 that as respiration product is 5.7 mL. And the respiration rate is the comparison between volume CO2 as the respiration product with the time of saving (24 hours), as much 0.2375mL/hour 4.3 Explanation Based on the data analysis above we can see that, the higher of temperature, the faster of respiration rate but just until the optimum temperature to do reaction rate. If the optimum temperature has reached, so the reaction rate will be decreased. It because respiration is the enzymatic reaction which is involving a lot of enzymes work inside. So, if the temperature is so high and in long time in that condition, the enzyme which is needed for doing respiration rate is denaturated. In this experiment we used bean sprout of green bean with age 2 days, because the bean sprout is very active doing metabolism for growth, and there is still no photosynthesize process in these bean sprout because they have not chloroplast organ yet in their body, so the energy from metabolism is concerning for growing activity of these bean. In other hand, the bean sprout in age 2 days starts to have cotiledone as the storage tissue (food storage) which contains of a lot of carbohydrate. The carbohydrate is the substrate in repiration process to get energy by carbohydrate combine with oxygen. So, the carbohydrate contain is decreasing for bean sprout growth. The reaction of respiration in bean sprout, as follow: C6H12O6 (carbohydrate) + 6 O2 6 CO2 + 6H2O + Energi

Bean sprouts do breathing to get the energy carried by involving gaseous oxygen (O2) as the material is absorbed / required and produce the gas carbon dioxide (CO2), water (H2O) and the amount of energy. Basically, the process of respiration has functions to get the energy used in metabolism and growth and developmental processes to become a mature plant. The larger a plant, then the greater needed for energy, so in respiration process is requiring a lot of oxygen too. The tested bean sprout us placed in the Erlenmeyer tube which is filled by NaOH solution. These NaOH has function to catch the carbon dioxide which is resulted by these bean sprout respiration. The respiration process that we tested to sprout is 24 hours, then the NaOH will be reacted with BaCl2 solution, and droped by PP solution as indicator. After it, these NaOH solution will be titrated using HCl to produce salt. The whole reaction of our practice activity to know the respiration rate, as follow:
-

When NaOH catch the carbon dioxide as the result of respiration process: NaOH + CO2 NaHCO3

When 5mL NaOH is reacted with 2.5 mL of BaCl2 : 2NaOH + BaCl2 2NaCl+ Ba(OH)2 Base properties because when it reacted again with indicator PP, the color is being redish purple.

When doing titration with HCl: 2Ba(OH)2 + 2HCl 2BaCl + H2O Base + Acid Salt + water

The amount of HCl needed to change the color of Ba(OH)2 reddish purple to the recent disappear (become white or transparent) is the amount of NaOH which is not binding the CO2 , or can be written like this: HCl needed = NaOH which is not binding CO2. So, from the volume of HCl needed, we can know how many NaOH which isnt binding the carbon dioxide. NaOH has ability to bind the Carbon dioxide as the product of respiration, but not all of CO2 can be binded by NaOH. So, the NaOH which is not binded the carbon dioxide is not all of them can be reacted with BaCl2 and produce Ba(OH)2 that has white color. Ba(OH)2 is tested with indicator PP, the color is changed to be redish purple that indicates the properties of that solution is base. When we titrates the 5 mL of Ba(OH)2 with HCl, it results the salt of BaCl2. We can know the solution has changed to be salt BaCl2 from the changing of color, if the redish purple color is

changed to be disappear recently of color being transparent, so the solution has been the salt. In the temperature range of 0C to 45C, a temperature increase will be followed by an increase in respiration rate. In incubation room with temperature 37 C has respiration rate 0.2375 mL/hour. In room temperature with 32 C has respiration rate 0.125 mL/hour. It means that in incubator temperature, the respiration rate is faster than in room temperature. Its because in incubator, the temperature is stable or constant. The enzyme work is more optimal if it is placed in stable temperature without the denaturation of enzyme. As we know, all the respiration reaction is involving the enzymes. If the enzyme is not denaturate, so the work of enzyme will be optimum in respiration. It means that the enzyme will make faster the glucose changing to be carbon dioxide, and the number of carbon dioxide produced in respiration of bean sprout is more number. If the temperature is higher, the volume of carbon dioxide is big binded by NaOH, so that the concentration of carbon dioxide which is produced in respiration is bigger too. In room temperature the volume of carbon dioxide as the product of respiration is fewer than in incubator room. It is because if the temperature is lower, the enzyme work is not optimal. It means that the enzyme will make slower the glucose changing to be carbon dioxide, and the number of carbon dioxide produced in respiration of bean sprout is fewer. If the temperature is lower, the volume of carbon dioxide is few binded by NaOH, so that the concentration of carbon dioxide which is produced in respiration is fewer too. In control Erlenmeyer which is not filled by bean sprout is also showing the respiration process. Because there is another microorganism which is did respiration except the bean sprout. We can know it because there is NaOH which bind the carbon dioxide, it means there is carbon dioxide production in the Erlenmeyer. Maybe from microorganism which live in the Erlenmeyer because we didnt sterilize the Erlenmeyer firstly before doing those practice. The respiration rate of Erlenmeyer with bean sprout is faster than without bean sporut and the carbon dioxide product is more than in without bean sprout, it because of the oxidizing substrate in respiratory metabolism. And generally substrates for respiration are substances that accumulate in number and relatively many metabolic

processes involves a series of enzymatic reactions that involve enzymes, the respiration rate on the existing Erlenmeyer sprouts are also influenced by the enzymes contained in the sprouts and enzymes will increase when the temperature also high when the temperature is too high but will also damage the enzyme. While the erlenmeyer containing only NaOH, the process of respiration is slower and less CO2 emitted. This is because it is not influenced by enzyme

CHAPTER 5 CONCLUSION From the experiment we did and the explanation above, we conclude that temperature is affecting the respiration rate of bean sprout. The higher of temperature, the faster of respiration rate happen, but just until the optimum temperature to do reaction rate. In the temperature range of 0C to 45C, a temperature increase will be followed by an increase in respiration rate. More of CO2 is produced by the bean sprout, the faster of respiration rate.

REFERENCES Rahayu, Yuni Sri. 2008. Petunjuk Praktikum Fisiologi Tumbuhan. Surabaya: Unesa Press Kimball, John W. 1983. Biologi Edisi 5. Bogor : Airlangga Asimov, Isaac (1968). Respiration. New York, London: Basic Books, Inc. Campbell, Neil A., Lawrence G. Mitchell, Jane B. Reece. 1999. Biology, 5th Ed. Benjamin/Cummings Publ. Co., Inc. Menlo Park, CA. (plus earlier editions) Affourtit C, Krab K, Moore AL. 2001. Control of plant mitochondrial respiration. Biochimica Biophysica Acta Bioenergetics 1504: 5869 Amthor JS. 1991. Respiration in a future, higher CO2 world. Plant, Cell and Environment 14: 1320. H,Lambers.2008. Plants respiration. Online at: http://books.google.com/books/about/Plant_respiration.html?id=TE9_bfFw-D8C, accessed at 6th November 2011. Fussi,annisa.2010.Respirasi pada Kecambah.Online at: http://annisanfushie.wordpress.com/2008/12/07/respirasi-pada-tumbuhan/, accessed at 6th November 2011.