pdf BIOCHEM-Post-Lab-Discussion-_E1-and-E2.pdf

BIOCHEMISTRY POST LAB DISCUSSIONS  MC2 BIOCHEMISTRY Ø Experiment 1: The Bunsen Burner & Common Laboratory Operations Objective/s: At the end of the experiment, students will be able to: Demonstrate the proper techniques for the commonly encountered laborato...

BIOCHEMISTRY POST LAB DISCUSSIONS  MC2 BIOCHEMISTRY Ø Experiment 1: The Bunsen Burner & Common Laboratory Operations Objective/s: At the end of the experiment, students will be able to: Demonstrate the proper techniques for the commonly encountered laboratory operations, such as:  Operation of the Bunsen burner  Transferring solid and liquid chemicals into another container  Heating of flammable and nonflammable liquids  Filtration  Evaporation  Solubility testing  Ø Experiment No. 1 The Bunsen Burner, parts and functions Base – supports the entire burner assembly Gas Valve – adjusts the height of the flame Chimney or Barrel – is where the air and fuel mixes for combustion Gas Inlet (gas intake) – where the fuel Butane gas) enters Air Hole – is an opening where air enters when the collar is adjusted Collar – is the movable part that allows the air to enter the airhole  Ø The Bunsen Burner, parts and functions Oxidizing flame – Luminous flame butane is The yellow flame signifies an completely incomplete combustion where not all oxidized to CO gas and water of the fuel butane are completely vapor converted to CO2 and H2O instead unburned C is visible by the black substance deposit and the emission of Carbon Monoxide (CO). The equation for incomplete combustion is: Non-luminous flame the desired type of flame in the laboratory C4H4 + 2 O2  CO2(g) + 2H2O(g) + 2CO(g) + The nature of the flame is blue which indicates complete C(s) combustion of fuel It signifies that full combustion takes place in this flame The equation for complete combustion is: 2 C4H4 + 10 O2  8 CO2(g) + 4 H2O(g)  Ø Weighing and Transferring Solid Chemicals Top Loading Balance Triple Beam Balance Analytical Balance This is intended to achieve This is more tedious and prone to Accuracy is highest than the 2 less error in reading error due to handling and reading. other balance. compared to triple beam. Air drafts and contamination The reading’s accuracy is The pointer is prone to erroneous from the surrounding are higher because the read out reading due to movement and air negligible due to its cover is around the 10 drafts Read out is up to the 10th thousandth. Longer time to determine mass. digits. Ø Weighing and Transferring Solid Chemicals Ø Measuring volume of chemicals Incorrect reading the volume of liquids result to errors. We call this error as parallax.  Ø Calculating for errors in measurement Incorrect reading the volume of liquids result to errors. We call this error as parallax. Errors are also attributed to many different cause which include: incorrect measuring device, other human errors such as handling, dilapidated markings, or defective device. On inappropriate device, errors are calculated against a measured value using low precision instrument versus a measured value against a high precision instrument. Normally, the high precision instrument gives the true value because it has the highest accuracy and repeatability. Moreover, high precision instruments brings forth a calibration certificate to prove it has been pre-calibrated against a known standard. Error, as % error = measured valueuncalibrated device – true value high precision device X 100 true value high precision device Sample Problem: A 150-ml alcohol is measured in a beaker and transferred to a 200-ml graduated cylinder. Upon all liquid is transferred to the graduated cylinder without spillage, the reading at the lower meniscus revealed that it is only 148.5 ml. Determine the error incurred by the first instrument (beaker) reading in beaker – reading in graduated cylinder 𝟏𝟓𝟎 −𝟏𝟒𝟖.𝟓 % error = X 100 = 𝑿 𝟏𝟎𝟎 = 1.01 % 𝒓𝒆𝒂𝒅𝒊𝒏𝒈 𝒊𝒏 𝒈𝒓𝒂𝒅𝒖𝒂𝒕𝒆𝒅 𝒄𝒚𝒍𝒊𝒏𝒅𝒆𝒓 𝟏𝟒𝟖.𝟓  Ø Transferring liquid chemicals to another container Note: Beakers are often used for transfer of liquids it contains a cleft (indentation) in its mouth that shall be placed On the rim of the receiving vessel (see left picture)  Ø Heating Liquids Hotplate – Heating Non-flammable chemicals Heating flammable chemicals is better with this type of instrument since it does not involve a flame. It is much safer to use this type even direct heating or water bath system.  Ø Precipitation and Filtration Precipitation is a chemical reaction between two aqueous solutions resulting to a formation of an insoluble solid at the bottom of a liquid. The insoluble solid is termed as precipitate and the liquid above it is called supernatant liquid or supernate.  Ø Precipitation and Filtration The precipitated insoluble solid can be separated from its liquid component through FILTRATION. The liquid is separated from the insoluble solid by passing it through a porous medium called the filter paper placed in the funnel. The insoluble solid that is left in the filter media is called the residue while the liquid that passes through the filter paper is called filtrate. The liquid filtrate is transferred to the evaporating dish and allowed to evaporate until dryness. The liquid that has evaporated is normally water vapor and what is left in the evaporating dish is the residue of the pure substance which in this expt. is KNO3 In precipitation, the glass rod may be used to allow the liquid to flow towards the filter paper without spilling the contents.  Ø Precipitation and Filtration The filter paper is the medium used to filter the insoluble solid and allowed the liquid to pass through. It is placed through the funnel as properly folded in quarters to ensure efficient filtration. A tear at one-corner of the filter paper is also necessary if the filter paper folded in quarter is smaller than the funnel to allow the air entry for faster flow of the liquid. Ø Evaporation Evaporation – is the process of separating a soluble mixture (homogeneous solution) from its components by heating. The evaporating dish is a ceramic apparatus used for evaporation processes. The liquid evaporates and the solid retained in the dish is a pure substance called the residue. Example a Potassium Nitrate aqueous solution is separated into components of water vapor and potassium nitrate powder.  Ø Schematic of precipitation, filtration & evaporation KNO3 (s) PbI2(s) KNO3 (aq) 𝑷𝒃 𝑵𝑶𝟑 𝟐 + 𝑲𝑰 → 𝑷𝒃𝑰𝟐 + 𝑲𝑵𝑶𝟑 Ø Liquid-Liquid Extraction Extraction – is the process of separating mixtures having different densities. The apparatus used for this operation is the separatory funnel. The higher density liquid comes out first than the lower density liquid. It is equipped with a stopcock to control the flow of the liquid.  Ø Solubility Solubility of a solute (a dissolved substance) in a solvent (one that dissolves the solute) is the most common principle or technique applied to laboratory operations such as crystallization, extraction or chromatography. A solid solute dissolved in a liquid solvent is termed soluble. If it does not dissolve, it is insoluble. Likewise, a liquid solute dissolved in a liquid solvent is miscible, otherwise, it is immiscible. The principle of like dissolve like is applied in solubility. A polar solute is dissolved in polar solvent, while nonpolar solute is dissolved in nonpolar solvent. Most ionic substances will dissolve in water (being a polar substance) ionic compounds with water normally dissociates into their ions while water creates a dipole, hence the interaction of positive and negative charges allows hydrogen bonding to happen. Organic compounds are normally like hydrocarbons are nonpolar substances hence they can only dissolved with nonpolar solvents.  Ø Solubility Organic Compound Water (polar) Ethanol (intermediate Hexane (nonpolar polar) solvent) Benzoic acid (nonpolar) insoluble soluble Soluble Citric acid (nonpolar) highly soluble soluble soluble Organic Compound Mixture Solubility at low Temp (25oC) Water (polar) and Ethanol (polar) Miscible Water (polar) and Hexane (nonpolar) Immiscible Water (polar) and Diethylether (med polar) Immiscible Miscible –solubility at room temp is 65 Ethanol (polar) and Hexane (nonpolar) g/100 ml which indicates that they are miscible to each other up to this range Hexane (nonpolar) and Diethylether (med polar) Miscible  BIOCHEMISTRY Ø Experiment 2 Qualitative Test for the Functional Groups in Organic Compounds Objective: To identify the common functional groups based on their general structural features  BIOCHEMISTRY Ø Qualitative Test for the Functional Groups in Organic Compounds BIOCHEMISTRY  BIOCHEMISTRY Ø Various Tests for the Identification of Functional Groups Functional Group Tests Reagents Results/ Product Formed Present Indicator (+) Alkenes Tests for KMnO4 Brown ppt: MnO2 Unsaturation: CH2 OH-CH2OH Alkynes Von Baeyer’s Test Normal CH3C(=O)-C(=O)CH3 diketone Basic 2 molecules of Formic Acid  BIOCHEMISTRY Ø Various Tests for the Identification of Functional Groups Test for Unsaturation – Von Bayer’s Test (addition reaction) Alkenes CH2=CH2 + KMnO4 → CH2OH – CH2OH + MnO2 + KOH + H2O - (vicinal diol) (Ethene) Ethene 1,2- ethane diol (brown ppt, reduced product) ethylene glycol-oxidized product) Cyclohexene + KMnO4 → + MnO2 + KOH cyclohexane glycol (brown ppt, red prod) Alkynes CH≡CH + KMnO4 𝑯𝟐𝑶 HCOOH + HCOOH + MnO2 + KOH (Acetylene) Acetylene 2 moles of formic acid, ox prod) (brown ppt, red. prod.)  Ø Various Tests for the Identification of Functional Groups Cyclohexane cyclohexene (-) result (+) result Tests for Unsaturation – Halogenation or Addition Reaction Functional Group Reagents Results/ Indicator (+) Product Formed Present Alkenes – Br2 Brown to colorless cyclohexene Alkynes- Br2 Brown to colorless Ethyne/acetylene Note: Alkanes will only react in the presence of a catalyst, UV light or sunlight, HENCE THE REACTION IS A SUBSTITUTION REACTION.  BIOCHEMISTRY Ø Various Tests for the Identification of Functional Groups Halogenation (Substitution) Reaction for Alkanes and Aromatic HC Alkanes Aromatic (Toluene) NOTE: If all hydrogens are replaced by a halogen (Br, Cl, etc), the by-product is no longer hydrogen halide but only Hydrogen  BIOCHEMISTRY Test for Alcohols and Phenols Comp’d Structure Bordwell-Wellman’s Lucas Test FeCl3 Test Reagent >>>>>>>>> K2Cr2O7 /conc H2SO4 ZnCl2 in conc HCl FeCl3 soln Ethanol CH3CH2-OH Turns blue green soln No turbidity; clear No test Isopropyl CH3-CH(OH)-CH3 Turns blue green soln Turbid after 5 min No test Alcohol T-Butyl (CH3)3-C-OH Remains orange Immediate turbidity No test Alcohol Phenol C6H5OH No test done No test done Violet complex  BIOCHEMISTRY Ø Rate of Oxidation tests for 1o, 2o and 3o alcohols Bordwell-Wellman’s Test Ethanol 1o + K2Cr2O7 + H+  + Cr2O3 + K2SO4 Cr2O72- Cr3+ Isopropyl Alcohol 2o + K2 Cr2O7 + H+  + Cr2O3 + K2SO4 Cr2O72- Cr3+  BIOCHEMISTRY Ø Rate of Oxidation tests for 1o, 2o and 3o alcohols Bordwell-Wellman’s Test T-BUTYL ALCOHOL 3o + K2Cr2O7 + H+  NO REACTION (color remains orange/ yellow) Phenol Ø Lucas tests for 1o, 2o and 3o alcohols  BIOCHEMISTRY Ø Ferric Chloride test for the presence of Phenol  BIOCHEMISTRY Ø Test for Aldehydes and Ketones Compound 2,4-DNP Test Tollen’s Test Acetaldehyde Formed a yellow precipitate Reduced product is Metallic 2,4-DNP Hydrazone silver Oxidized product is Ethanoic/Acetic Acid Acetone Formed a yellow precipitate NO reaction 2,4-DNP Hydrazone Ethanol No reaction NO reaction  BIOCHEMISTRY Ø Test for Aldehydes and Ketones – 2,4-DNPH test Acetaldehyde acetaldehyde-2,4-dinitrophenylhydrazone (yellow to red-orange precipitate)  BIOCHEMISTRY Ø Test for Aldehydes and Ketones – 2,4-DNPH test 2-propanone 2-propyl-2,4-dinitrophenylhydrazone (yellow to red-orange precipitate)  BIOCHEMISTRY Ø Test for Aldehydes and Ketones – Tollen’s Test  BIOCHEMISTRY Ø Test for Carboxylic Acid – NaHCO3 test  GOOD LUCK TO THE LONG QUIZ and the PRACTICAL MIDTERM EXAM. “Falling down is not a FAILURE. Failure comes when you stay where you have FALLEN” - Socrates So,