Analysis of Amino Acids by Paper Chromatography
Introduction-
Proteins may be thought of natural polymers of amino acids, as the
composition of proteins is of amino acids.
The technique known as paper chromatography is used to separate amino
acids for analysis. In this technique small spots of amino acids are
introduced to a piece of porous filter paper. The bottom of the paper
is then placed in a small bath of an appropriate solvent. The solvent
is allowed to rise up the paper. The various amino acids are attracted
to the paper to different degrees due to their differences in
polarity.
Various amino acids travel at different rates.
The location of the amino acids can be determined by spraying the
paper with a 2% solution of Ninhydrin in ethanol. The Ninhydrin forms
a purple complex with an amino acid which is readily identified.
Materials-
Solutions of amino acids used are -
Amino acid mixture 'X'
Aspartic Acid
Leucine
Proline
Asparagine
Phenylalanine
Solvent (ammonia and propan-2-ol in the ratio 2:1)
2 % solution of Ninhydrin in ethanol
Method-
Setting up the apparatus
1. This section of the experiment is done in a fume cupboard as the
solvent is dangerous. Safety glasses should also be worn. The
solvent is poured into a chromatography jar to a depth that is
enough for the chromatography paper end to be dipped into. A lid
is placed over the jar and left in the fume cupboard. This gives
time for the air in the jar to be saturated with the solvent of
the vapour.
2. Before handling the chromatography paper, hands are washed. The
chromatography paper mustn't be touched all over the place
constantly because amino acids that is smeared on the hand may get
For this experiment we have to use physical methods to separate the reaction mixture from the liquid. The physical methods that were used are filtration and evaporation. Filtration is the separation of a solid from a liquid by passing the liquid through a porous material, such as filter paper. Evaporation is when you place the residue and the damp filter paper into a drying oven to draw moisture from it by heating it and leaving only the dry solid portion behind (Lab Guide pg. 33.).
Catecholase is an enzyme formed by catechol and oxygen used to interlock oxygen at relative settings, and it is present in plants and crustaceans (Sanyal et. al, 2014). For example, in most fruits and vegetables, the bruised or exposed area of the pant becomes brown due to the reaction of catechol becoming oxidized and oxygen becoming reduced by gaining hydrogen to form water, which then creates a chain that is is the structural backbone of dark melanoid pigments (Helms et al., 1998). However, not all fruits and plants darken at the same rate. This leads to question the enzymatic strength of catecholase and how nearby surroundings affect its activity. The catecholase enzyme has an optimal temperature of approximately 40°C (Helms et al., 1998). Anything above that level would denature the tertiary or primary structure of the protein and cause it to be inoperable. At low temperatures, enzymes have a slower catalyzing rate. Enzymes also function under optimal pH level or else they will also denature, so an average quantity of ions, not too high or low, present within a solution could determine the efficiency of an enzyme (Helms et al., 1998). Also, if more enzymes were added to the concentration, the solution would have a more active sites available for substrates and allow the reaction rate to increase if excess substrate is present (Helms et al., 1998). However, if more
Abstract/Summary: “Proteins account for more than 50% of the dry weight of most cells, and they are instrumental in almost everything organisms do” (Campbell, 1999). The significance of proteins to the continuation of our biological systems is undeniable, and a study of how to quantify proteins seems an appropriate introduction to our studies of biology. In order to study proteins we must first know how to separate then quantify the amount using basic principles of experimental design such as a standard curve. In this experiment we wish to quantify the amount of previously extracted protein by measuring the absorbance of the unknown amount and determining its concentration by overlaying it against a standard curve of the absorbance of known concentrations of the protein. We used the dye agent Bradford Protein Assay to get an absorbance of 0.078, 0.143, 0.393, 0.473, and 0.527 at the protein’s respective concentrations of 0.28, 0.56, 0.84, 1.12, and 1.40 mg/mL. When a best-fit line was applied to the standard curve, and the absorbance of our unknown concentration (0.317 A) plotted, we estimated a concentration of around 0.84 mg/mL of protein. Our calculations indicated a quantity of 168 mg of protein, which was an approximately 8.96% yield of the projected 1875 mg that was expected. Errors that may have led to this small yield percentage may have stemmed from our previous lab and our initial attempts to extract the desired amount of protein.
J.C. Biro, B. Benyó, C. Sansom, Á. Szlávecz, G. Fördös, T. Micsik, and Z. Benyó; A common periodic table of codons and amino acids. Biochemical and Biophysical Research Communications 306 (2003) 408–415.
The purpose of this experiment was to study the reactions of amino acids and aspartame. Several solutions were prepared and used in TLC analysis. A permanganate test and a ceric nitrate test were also performed. A summary of the results is shown below. TLC Analysis/Rf Values • Plate I o Solution 1: Aspartame Rf: 0.571 o Solution 2: Phenylalanine Rf: 0.571 o Solution 3: Aspartic Acid Rf: 0.217 o Solution 4: Base-hydrolyzed Aspartame Rf:
Introduction: Purifying proteins is an important part of biology because it can help identify the function of that protein. Once a protein’s function has been identified, it can be manipulated to see how the function would change if the protein was changed. A common way to purify a protein is through Ion Exchange Chromatography, which is where charged proteins will bind to the beads in the column to purify it from the solution (Berg JM, 2002). The purpose of this experiment is to use Ion Exchange Chromatography to purify cellulase.
Due to the nature of amino acids, a titration curve can be employed to identify
Nutrition and physical activity are important aspects as they improve health and reduce chronic diseases. In June of this year my family physician placed me on a doctor assisted weight loss program that enabled me to eat my favorite foods but in appropriate proportions and in heathier ways. The best part about Supertracker is that it gives me wonderful tools to support me in understanding and meeting my goals. Once I created a profile through Supertracker I was provided a personal plan to follow including the amount of grains, vegetables, fruits, and protein I needed to be successful. For three consecutive days, I entered my food and activity intake into Supertracker and was able to analyze the excesses and deficiencies
Gel electrophoresis is used in a variety of settings, particularly in molecular biology. Besides being used to separate nucleic acids, such as DNA and RNA, gel electrophoresis is also employed to divide proteins (Gel Electrophoresis). According to research, electrophoresis is applied for the following reasons, "To get a DNA fingerprint for forensic pur...
LAB REPORT 1st Experiment done in class Introduction: Agarose gel electrophoresis separates molecules by their size, shape, and charge. Biomolecules such as DNA, RNA and proteins, are some examples. Buffered samples such as glycerol and glucose are loaded into a gel. An electrical current is placed across the gel.
= Before conducting the experiment I would conduct a simple test for the protein by placing a sample of the albumen into a test tube and add biurett reagent. This contains copper (II) sulphate and sodium hydroxide.
The enzyme used in this lab is catalase. Catalase has a molecular weight of approximately 240,000 daltons and contains four polypeptide chains, each composed of more than 500 amino acids. This enzyme occurs universally in aerobic organisms. One function of catalase within cells is to prevent the accu...
An alpha amino acid is made up of a central carbon atom, or the alpha carbon, which is linked to an amino group, a carboxylic acid group, a hydrogen atom, and a distinct R group, called the side chain. There are twenty different kinds of side chains that vary in shape, hydrogen-bonding capacity, chemical reactivity, charge, size, and hydrophobic character that are typically found in proteins. All proteins in all species are made up of the same set of twenty amino acids, with a few exceptions. In order to classify amino acids, the molecules are assorted in four groups on the basis of the general characteristics of their R groups. The four groups are hydrophobic amino acids with nonpolar R groups, polar amino acids with neutral R groups but the charge is not evenly distributed, positively charged amino acids with R groups that have a positive charge at physiological pH, and lastly, negatively charged amino acids with R groups that have a negative charge at physiological pH. The simplest amino acid is glycine because it has only a single hydrogen atom as its side chain. Alanine is the next simplest amino acid because it has a methyl group as its side chain. Seven of the twenty amino acids have side chains that are readily ionizable and they are able to accept or donate protons to facilitate reactions and form ionic bonds. Amino acids are typically abbreviated to a three-letter, which are typically the first three letters
The movie Food Inc. was very engrossing. I honestly believed that all foods in the grocery store came from different places. It made me very sad to see how the major food companies would deport illegal immigrants even though, they were the ones who provided transportation and ways for the immigrants to come to America illegally. 95% of my family are all immigrants. My family comes from Belize which is not very from Mexico; So that part of the movie struck me the hardest. I imagined about what would’ve happened if they decided to take one of my family members. The movie opened my eyes to many evil things I did not know about.
Defying Robinson Appleby 's agreement to never teach a Negro how to read, Mamed, the overseer with a raised cane, proposes Aminata an offer which she cannot decline. Being the overseer in Appleby 's plantation, Mamed 's duty is to ensure that the plantation is properly functioning while Master Appleby is absent. In fear of being disciplined for divulging the prohibited prayer, Aminata recognizes Mamed 's humanity as he murmurs, “Allahu Akbar”. With Mamed 's offer to Aminata to tutor her how to read and write, perhaps if “Georgia was teaching [Aminata] how to survive in the land of the buckra, but maybe Mamed could teach [her] how to get out” (Hill, 216). Mamed 's teachings start with the pronunciation of Aminata 's name to eventually forming a sentence with basic, common words to the rules and procedures established in Appleby 's hacienda. As Aminata 's fear of Mamed eventually subsides, Aminata apprehends the idea of Mamed being a different man, “like a man who was willing to teach” (Hill, 218).