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Biol160 Chemistry The Basic Chemistry of Life In order to understand living organisms and how they function, it is important to recognize the basic properties of the fundamental substances of which we are composed. All of the “stuff” that exists in the universe, living and nonliving, is composed of matter. Matter can be found in solid, liquid, and gaseous forms, and is composed of chemical elements. An element is a substance that cannot be broken down into anything else by basic chemical means. There are almost 100 elements that occur naturally on Earth. However, very few elements exist in large quantities in pure form. Through chemical bonding, elements frequently combine with others to form compounds. A compound may consist of just two elements, or it may contain more. A single atom is the smallest stable subdivision of a pure element. A molecule is the smallest independently-existing unit of a compound. The number of atoms in a molecule depends on what type of molecule it is. Small molecules may be composed of only 2 or 3 atoms, while large, complex ones may have thousands of atoms. Please note that this lab exercise includes a pre-lab worksheet, which you should complete and turn in to your instructor before the start of your lab session. I. Atomic structure and chemical bonds Atoms are made up of subatomic particles, including protons, neutrons, and electrons. The table below summarizes their properties. The central region of an atom is called the nucleus. Do not confuse this with the nucleus of a cell, which we’ll examine in the next lab session. The electrons orbit around the nucleus very rapidly. If an atom were modeled to scale, the nucleus might be represented as a pea hovering in the center of a professional sports stadium, and the electrons would be tiny specks of dust zipping around somewhere within the confines of the stadium. The number of protons determines the type of element. For example, if an atom has 12 protons, then it is a magnesium (Mg) atom. If it were possible for that atom to lose one proton, so that it only had 11, then we would be dealing with the element sodium (Na). The number of neutrons and electrons present in an atom is often the same as the number of protons. For example, a typical helium (He) atom has 2 protons, 2 neutrons, and 2 electrons. However, numbers of neutrons and electrons can vary. Atoms of an element that vary in mass because they have different numbers of neutrons are called isotopes. The element oxygen (O) has seven known isotopes. For example, there are oxygen atoms that contain 8 protons and 8 neutrons, while others contain 8 protons and 10 neutrons. Atoms of some elements gain or lose electrons readily. Sodium (Na) atoms frequently lose one of their 11 electrons. That leaves them with 10 electrons and 11 protons, and thus one net unit of positive charge. These charged atoms are called ions, and are symbolized like this: Na+, Ca2+ (calcium atom that has lost two electrons), Cl- (chlorine atom that has gained one electron). proton Location Mass Electrical charge If the number present is changed… center of the atom (nucleus) 1 atomic unit positive it is no longer the same element it is a different isotope of the same element neutron center of the atom (nucleus) 1 atomic unit neutral electron orbiting around the nucleus almost none negative it gains a net charge and is called an ion, but is still the same element Biol160 Chemistry Within an atom, electrons are located at different energy levels, called electron shells, based on how far they are from the nucleus. The number of electrons in the outermost shell determines how an atom will interact with others. Each electron shell can “hold” a certain number of electrons. The first shell, closest to the nucleus, can hold two electrons. Thus, helium (He), which has only two electrons, has a full outer shell. The second electron shell can hold up to eight. Therefore, neon (Ne), which has 10 electrons, also has a full outer shell, since it has 2 electrons in the first shell and 8 in the second. The third and fourth shells can hold more than 8 electrons, but they generally become unstable when more are present. Because of this, a general rule of thumb is that atoms react with one another so as to end up with eight electrons in their outermost shells. In order to achieve this, atoms can gain, lose, or share electrons. In order to picture how this will occur, it is helpful to draw shell models to depict different types of atoms. The examples below have a grey circle for the nucleus, and black dots as electrons. O H Na Ionic bonds Ionic bonds form between atoms due to the attraction of opposite charges that result from the gain or loss of electrons. Typically, atoms with 1, 2 or 3 electrons in their outer shell lose these electrons to other atoms. Look at the shell model of sodium (Na) above. Why will losing one electron make it more stable? Atoms with 6 or 7 electrons in their outer shells are usually the ones to receive or gain those electrons. Table salt, or sodium chloride (NaCl), is a classic example of a compound held together by ionic bonds. Covalent bonds Most biological macromolecules are held together primarily by covalent bonds. These occur when atoms share electrons. One or more pairs of electrons spend part of their time in the outer shells of both atoms involved, thus the outer shells are effectively filled. Water (H2O) is an excellent example of a molecule held together by the sharing of electrons. Oxygen (O) has 6 electrons in its outer shell; thus, to be satisfied, it requires 2 more. Hydrogen (H) atoms require one more, since the capacity of their outer shell is only two electrons total. So the oxygen shares one pair of electrons with one hydrogen atom, and another pair of electrons with a second hydrogen atom. Each pair of shared electrons consists of one that was originally part of the oxygen atom, and one from the hydrogen. A water molecule is comprised of three atoms held together by two single bonds. Single bonds are represented in structural formulas as single lines: H—O—H, for example. If two atoms share two pairs of electrons, they are held together by a double bond. Oxygen gas (O2) is a good example of this, and it would be written as O==O. The table below illustrates the bonding capabilities of the four elements that make up over 90% of the mass of most living organisms. Element Number of protons Number of outer shell electrons Total number of potential bonds hydrogen carbon nitrogen oxygen 1 6 7 8 1 4 5 6 1 4 3 2 Biol160 Chemistry Be sure to answer all of the questions on the worksheet relating to atomic structure and chemical bonds. Unfortunately, your textbook does not provide a periodic table of the elements, so I have listed elements with atomic numbers 1-30 on the last page of this lab write-up. II. pH - DEMO Because water is such a large part of living organisms and our environment, another important chemical aspect to consider is how compounds behave when placed in water. Acids are substances that release hydrogen ions (H+) when dissolved in water. Hydrogen chloride (HCl) is considered a strong acid, because it dissociates almost completely into H+ and Cl- ions. Bases, also called alkaline compounds, release hydroxyl ions (OH-) when dissolved in water. In a sample of pure water, some of the water molecules themselves dissociate into H+ and OH- ions, but since the amounts are precisely the same, water is not considered an acid or a base. Substances that do not release any hydrogen or hydroxyl ions when dissolved are not considered acids or bases, either. The pH scale is used to indicate the concentration of hydrogen ions (H+) in a solution. The values range from 0 to 14. Lower pH values indicate a higher concentration of H+ ions and are considered more acidic. Each unit represents a 10-fold change in H+ concentration. Thus, a solution with a pH of 5 has 10 times more H+ ions than something with a pH of 6, while something with a pH of 4 is 100 times more acidic. At a pH of 7, a solution is considered neutral, because the concentration of H+ ions is equal to the concentration of OH- ions. Note: Although you would expect the pH of pure water to be exactly 7, when water is exposed to air, carbon dioxide from the air dissolves in it and forms a weak acid. Thus the pH of distilled water is usually about 5.7. Solutions with pH values greater than 7 are called basic or alkaline. In lab today, work in groups of 4 for ALL of the following exercises. ACTIVITY 1: Using pH test strips, determine the pH of the 8 different solutions listed below. These are all substances you might encounter in your daily lives. Record their pH values here, and then on your worksheet, list them in order from the most acidic to most basic, along with the pH value obtained. Solutions for pH testing: Alka-Seltzer solution Ammonia detergent solution cola (soda pop) vinegar lemon juice milk water that has been filtered through potting soil The processes that occur inside living organisms are often very sensitive to pH, and therefore pH must be maintained within narrow limits in order for things to function properly. Buffers are compounds that can release or grab H+ ions and thereby keep the pH from changing drastically. Buffers within our blood, for example, maintain a slightly basic pH (about 7.4). The following exercise will illustrate how a buffered solution resists changes in pH. ACTIVITY 2: 1. Use pH strips to determine the pH of distilled water (dH2O) from your dropper bottle, and of the pH 6 buffer solution. Record the values on the table in your worksheet. 2. Place 40 drops of dH2O in one small labeled beaker, and 40 drops of pH 6 buffer solution in another. 3. Add 1 drop of 1% hydrogen chloride (HCl) to each, and determine the pH of each by dipping a test strip into the solution in the beaker. Record the value on your worksheet. 4. Add 4 more drops of HCl, and test the pH. Record the value. 5. Add 5 more drops of HCl, and test the pH. Record the value. 6. Add 10 more drops of HCl, and test the pH. Record the pH value. 7. Wash and dry the beakers. Repeat the experiment, using 10% sodium hydroxide (NaOH) instead of HCl.*Handle the HCl and NaOH carefully, and rinse your hands under water if some gets on your skin.* Biol160 Chemistry III. Testing for carbohydrates, lipids, and proteins Carbohydrates, lipids, and proteins are relatively large molecules that play important roles in living organisms. Carbon atoms and the four covalent bonds they can form provide the basic framework for ALL of these macromolecules. In this lab, we will use a suite of chemical tests to detect the types of molecules present in various food items. Carbohydrates come in several basic forms. The simplest sugars are called monosaccharides, and they contain 3-7 carbon atoms, often bonded together in a ring structure. Glucose is a very common 6-carbon sugar. When two monosaccharides bond together, they form a disaccharide, such as sucrose. Many monosaccharide subunits bonded together form a polysaccharide, a much more complex carbohydrate. Starch, cellulose and glycogen are common examples of polysaccharides. Our tests will allow us to easily detect starch and also reducing sugars (most 6-carbon and some 12-carbon sugars). There are many different types of lipids. They are all grouped together as lipids because at least some part of the molecule does not dissolve in water. Oil and vinegar salad dressing requires vigorous shaking, because the oil and vinegar (which is a water-based solution) do not mix well. We will use two different tests to help determine whether or not lipids are present in our samples. Most proteins are very large molecules with complex shapes. While monosaccharides are the subunits of polysaccharides, amino acids are the subunits of proteins. Amino acids connect via a specific type of covalent bond called a peptide bond. The test we will perform responds to the presence of peptide bonds, and thus indicates the presence of proteins. ACTIVITY: Use the methods described below to test for the presence of starch, reducing sugars, lipids, and proteins in a variety of foods. To do this, you must first prepare a set of standards. The standards demonstrate strongly positive and strongly negative results for each test. For example, when you perform the iodine test and the “liquid to be tested” is a solution of dissolved starch, the liquid should turn a dark blue-black color, indicating the presence of a lot of starch (strongly positive result). When you perform the iodine test and the “liquid to be tested” is distilled water, which contains no starch, the clear water should turn a yellow-brown color. Despite the color change due to the addition of the iodine solution, this is a negative result—starch is not present. SAVE YOUR PAIRS OF POSITIVE AND NEGATIVE STANDARDS FROM EACH TEST UNTIL YOU COMPLETE THE ENTIRE LAB EXERCISE. You’ll need to compare them to the results you obtain when testing various foods. The next table explains the substances we’ll use for creating the standards. Test Positive standard Negative standard iodine test for starch Benedict’s test for reducing sugars paper spot test for lipids Sudan IV test for lipids biuret test for proteins 0.1 % soluble starch 0.1% glucose vegetable oil vegetable oil 1% albumin (protein from egg whites) distilled water distilled water distilled water distilled water distilled water To create your set of standards, you must perform each of the five tests twice. One test will utilize the item listed in the Positive standard column, and one will use the item in the Negative standard column (dH2O) as the “liquid to be tested.” Specific directions for performing each of the tests are given below. Iodine test for starch 1. Place 25 drops (about 1 mL) of the liquid to be tested into a clean test tube. 2. Add 3 drops of iodine solution (often labeled “Lugol’s IKI”) to the liquid. Swirl gently to mix. 3. Examine the resulting color: graygrayish blueblue-black = presence of starch (low concentrationhigh concentration) Biol160 Chemistry Benedict’s test for reducing sugars 1. Prepare a hot water bath using a glass beaker, water, and a hot plate: Fill a 250 mL beaker halfway with water. Place it on the hot plate, and set the hot plate to medium-high; the water should be near boiling. 2. Place 25 drops (about 1 mL) of the liquid to be tested into a clean test tube. 3. Add 3 drops of Benedict’s solution to the liquid. Swirl gently to mix. 4. Place the test tube in the beaker of near-boiling water for 4-5 minutes. 5. Examine the resulting color: light greenyelloworangebrick red = presence of reducing sugars (low concentrationhigh concentration) Paper spot test for lipids 1. Place a drop of the liquid to be tested on a brown piece of paper. If the substance to be tested is a solid, it can be rubbed into the paper. Blot or wipe away any excess material. 2. Allow the paper to dry. 3. Once dry, hold the paper up to the light. A permanent translucent spot indicates the presence of lipids. Sudan IV test for lipids 1. 2. 3. 4. Place 25 drops (about 1 mL) of distilled water into a clean test tube. Add 3 drops of Sudan IV dye. Swirl gently to mix. Add 10 drops of the liquid to be tested, and mix again. Wait for 5-6 minutes. If lipids are present, they will stain darker red than the water, and will usually form small globules floating on the top of the mixture. Biuret test for proteins 1. Place 25 drops (about 1 mL) of the liquid to be tested into a clean test tube. 2. Add 25 drops (about 1 mL) of 10% NaOH. Swirl gently to mix. Recall from the pH exercise that NaOH is a strong base, and can be caustic. If you get any on your skin, wash it off immediately with a lot of water. 3. Add 5 drops of copper sulfate (CuSO4) to the liquid, mix, and wait 3 minutes. 4. A pale purple color indicates that proteins are present. 5. CuSO4 should not be dumped down the drain. Dispose of all liquid from these test tubes into the specially marked container under the hood for proper disposal. Once your set of standards has been created, choose at least three of the eight food items available for testing. Perform all five tests on each food, and in each case, compare your results to the appropriate pair of positive & negative standards to evaluate the presence or absence of that type of molecule in the food. Food items available for testing: sweet onion potato Sprite or 7-Up soft tofu salad dressing energy bar apple cooked rice In some cases, the food will need to be ground up using a mortar and pestle in order to obtain liquid to use in the tests. In those cases, use the following procedure: 1. 2. 3. 4. 5. Clean and dry the mortar and pestle before you begin. Cut the food into very small chunks and place a spoonful-sized amount into the mortar. Add about 5 mL of distilled water. Use the pestle to grind the material firmly, until soupy. Add more water if necessary. Pour the liquid into a test tube; this is your “liquid to be tested.” Note that to perform all the tests, you will need several milliliters (mL) of solution, so be sure you obtain enough liquid when processing the food. Biol160 Chemistry Atomic number (indicates the number of protons in an atom) 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 Name of element Abbreviation hydrogen helium lithium beryllium boron carbon nitrogen oxygen fluorine neon sodium magnesium aluminum silicon phosphorus sulfur chlorine argon potassium calcium scandium titanium vanadium chromium manganese iron cobalt nickel copper zinc H He Li Be B C N O F Ne Na Mg Al Si P S Cl Ar K Ca Sc Ti V Cr Mn Fe Co Ni Cu Zn Biol160 Chemistry Name ______________________________ Section ______ Chemistry Pre-Lab Worksheet (complete this sheet individually, and turn it in at the start of lab) Write the term or short phrase that best completes each sentence. Large ___ may be composed of hundreds or thousands of atoms. ______________________________ Isotopes of an element contain different numbers of… ______________________________ Covalent bonds occur when atoms share… ______________________________ A solution with a pH value of ___ is strongly basic. ______________________________ Solution from the pH testing section that you would not want to eat ______________________________ Once you complete step 6 of the pH buffer experiment, indicate how many total drops of HCl you will have added to the beakers of… distilled water __________ pH 6 buffer solution __________ For each of the following substances, identify whether it is a carbohydrate, lipid, or protein. If it is not one of those, write “other.” vegetable oil _____________________ albumin ________________________ glucose ______________________ starch sucrose ________________________ distilled water ________________ _________________________ Each of the following statements applies to one of the five tests we will perform in Part III of this lab. Give the name of the appropriate test. Can be performed without using a test tube ________________________________ The solution being tested may turn pale purple ________________________________ Requires that the test solution be heated ________________________________ Tests for the same type of molecule as the paper spot test ________________________________ A very dark colored solution indicates that a lot of starch is present ________________________________ Do not dump solutions from this test down the drain ________________________________ Table sugar (sucrose) might also work as a positive standard for this test ________________________________ One of the food items we will have available for testing will be an energy bar. It is likely to be lemon-flavored, coated with yogurt. Why would a dark chocolate energy bar be more challenging to work with in this exercise? (And since we don’t consume food in lab, the answer is not that it would be eaten before we could use it!) Biol160 Chemistry Biol160 Chemistry Name _____________________________________ Section ______ Basic Chemistry of Life Homework (complete and turn in this sheet—front and back—as an individual) I. Atomic structure and chemical bonds Give at least two examples of things that fit each description. You are encouraged to use tables and figures from your textbook to help you answer these questions. Element whose number of protons found in its atoms is a multiple of four ___________________ Element whose outer electron shell is full without gaining, losing, or sharing electrons ___________________ Element where the atoms require exactly two more electrons to fill their outer shell ___________________ Example of a molecule made of 2 atoms ___________________ Example of a molecule made of more than 2 atoms ___________________ Example of a compound composed of 2 different elements ___________________ Example of a compound composed of more than 2 different elements ___________________ A positively charged ion ___________________ Draw shell models of an atom of carbon (C) and an atom of potassium (K). Carbon: Potassium: Use shell model drawings and words to explain how the compound calcium chloride (CaCl2) is held together (what type of bond is involved), and why there are always two Cl for every one Ca. (Hint: You may find Figure 2.7A in your text helpful.) Biol160 Chemistry Use shell model drawings and words to explain why the element hydrogen (H) is found in the form H2 and what type of bond holds it together. Using the information provided about hydrogen, carbon, nitrogen, and oxygen, draw your prediction of the structural formula for each of these molecules. methane (CH4) carbon dioxide (CO2) formaldehyde (CH2O) Ammonia is a compound formed by the elements nitrogen and hydrogen. How many atoms of each would you predict would be present in a single molecule? In the space below, draw a shell model diagram that shows a molecule of ammonia. nitrogen ______ hydrogen ______ Biol160 Chemistry Name ______________________________ Name ______________________________ Section ______ Name ______________________________ Name ______________________________ Basic Chemistry of Life Worksheet (complete and turn in this sheet—front and back—as a group of four) II. pH - DEMO Solutions tested: (give the pH value on the line to the right) most acidic most basic ____________________________ ________ ____________________________ ________ ____________________________ ________ ____________________________ ________ ____________________________ ________ ____________________________ ________ ____________________________ ________ ____________________________ ________ Buffer solution and addition of an acid: total drops of 1% HCl added 0 1 5 10 20 pH of distilled water Buffer solution and addition of a base: pH of buffer solution total drops of 10% NaOH added pH of distilled water pH of buffer solution 0 1 5 10 20 Did this buffer solution do a better job of maintaining stable pH with the addition of an acid or a base? Explain. Soils vary in their acidity and in the amount of buffering compounds contained in them. Suppose that acid rain falls in an area where the soil has a high buffering capacity. Do you expect that there will be much effect on the soil organisms living in the area? Explain your answer. Biol160 Chemistry III. Testing for carbohydrates, lipids, and proteins In this table, record the results from your preparation of the set of standards. Describe the appearance of the liquid in each tube (or the brown paper) when the test was complete. Test Positive standard Negative standard iodine test for starch Benedict’s test for reducing sugars paper spot test for lipids Sudan IV test for lipids biuret test for proteins Record the results from testing the various food samples. Use this notation to indicate your results: strongly positive (i.e. looked very much like positive standard) moderately positive weakly positive negative Substance tested Iodine Benedict’s Test results Paper spot Sudan IV Biuret +++ ++ + 0 Your conclusions: Compounds present in sample Please note that these tests are not exceptionally sensitive, so keep in mind that negative results only mean that there were not large quantities of that type of macromolecule present. For example, all living cells contain lipids in their cell membranes, but that amount alone is too small for either of our lipid tests to detect. 14