- 4% ASPECTS OF ANALY TI BIOCHEMISTRY & SEPARATION TECHNIQUES The methods employed for cell fraction (A) Sonic homogeni: s enization and (B) Mast lala (C) Therm; (D) (E) on include ration hy pressure | nil disintegration | ding ina mortar | Centrifugation “| The follow 2 mes ‘c useful in tissue disintegration and cell fractionation (A) Sucrose medium \ (B) — Caesium (C) Sephadex (D) Pol (BE) il epichlohydrin “| During differential centrifug: emerge are (A) Lysosomal fraction at 1 (B) Mitochondrial fraction at 6003 ic) Soluble tion at S.000g (D) Nuclear fraction at 6003, |. the fractions and speed at which they ‘The appli (Ay Dete' (B) «cy nant purification of eZ «) of enzyme preparations (ik) Frac ont paper chromatography between two immiscible nyg statement asec! on pa Consider the follov (Ay Itseparates out solutes liquids and adsorption to paper. | (8) Comparison of the retention factor of unknown samples with known is employed for identifying the samples. | (C) The paper containing | the separated samples is called a chromatogram. ~! | (D) The chromatographic tank must be saturated with the solvent system for the run. | - (E) _ Ninhydrin solution can be employed for identifying amino acids, 307 ‘Scanned with CamScennerA Mark true or false (A) (B) (c) (D) «) In concentrated HCL solutions, Ketoses are dehydrated faster than aldoses to furfurals \ Most furfurals form colored complexes with resorcinol reagent. | Seliwanoff's reaction is used to Identify hexoses { Bial's test Is specific for pentoses — Molisch’s test Is # general one for carbohydrates” | Examine the following statements (A) (B) (c) (D) () ‘A colorimeter permits the passage of a broad range of wavelengths, while a spectrophotometer splits the light and allows waves of specific wavelengths to enter a solution | A spectrophotometer is a much refined version of a colorimeter | Using a colorimeter, measurements can be made in the uv range | A spectrophotometer is better than a colorimeter for obtaining 7 absorption spectrum A colorimeter is a more sensitive instrument than a spectrophotometer Examine the following statements (A) (B) (c) @) () Beer's law states that the amount of light absorbed by a solution is inversely proportional to its concentration Lambert’s law states that the amount of light absorbed is directly proportional to the thickness of the solution, Monochromatic light means light of uniform wavelength. Molar extinction coefficient Impurities decrease the value of molar extinction coefficient is a constant for a given compound. Still on colorimetry/spectrophotometry (A) Beer-lambert’s law is valid at all concentrations of absorbing species | (B) There is deviation from linearity at high concentrations (C)__ The absorption of light by 2 compounds at the same wavelength is additive, provided there is no reaction between them, | (D) Absorbance is the same as optical density (E) A cuvette makes for uniform thickness ¢ Fluorimetry a Can be used to estimate concentration (T]_ “| ( Measures the diffe ii ae eaifernce in energy levels between excited and ground © Maes Ps of an instrument called Muorimeter | (D) Differs from spectrophotometry in that the fluoresce ig i measured at right angles to the incident Ught {1} sce ligheleR © Resembles spectrophotometry in having a prism filter for adjusting wavelength 308 ‘Scanned with CamScanner1. The following are com, (AYA Tight souree ns OF 8 colorimeter (B) A column packed (C)A cuvette weit al (D)Lenses (E) Galvanometer 12. ane — the following with regard to paper chromatography 1s based on partition between chemical; i water in mobile pha: — ss (B) tis based on the principle of adsorption (C) Ihas a variant called 2-dimensional paper chromatography for ” resolving components with close R, values If 2 components have the same R, value in the same solvent system they are most likely the same. (E) tis a miniaturized form of counter-current distribution. 13 The following separation techniques are based on adsorption (A) Thin layer chromatography (B) _Ion-exchange (C)__ Electrophoresis (D) — Gas-liquid chromatography (E) Column chromatography is 14 Examples of adsorbents are (A) Cellulose (B) Alumina (©) Activated charcoal ) cic acid (T (E) Magnesium silicate [T 15 Features of gas-liquid chromatography include (A) Along; coiled tubular column coated with adsorbent (B) An inert carrier gas © A refrigerated chamber (D) Anoven (E) A flame ionization detector 16 Inelectrophoresis, the rate of migration is influenced by (A) pil (B) Total charge (C) Size (D) Molecular weight (E) Purity of sample ‘Scanned with CamScanner\7 18, Ww 20. Solution methods for the fractions ia (A) De-proteinization, by) shaking with) a phenol solution and then centrifugation (B) Use of detergents ar high salt concentrations to dissociate protein fram nucleic acid. Ultra centrifugation techniques, Use of electrophoresis Precipitation with ethanol Still on nucleic acids and chromatograph (A) Paper and thin-layer ebiror oligonucleotides. (1) The techniques of paper ayer chrom: largely replaced hy more pow chniques of HE (C) Large nuc! Js are often sep eXchange chromatogr (D) Hydroxyapatite binds: stranded DNA. (8) mRNAs can be ise shy are useful in fractionating e been With respect to the electrophoretic sep (A) Nucleic acids can be separated by polyacrylamide gel electrophores (PAGE) bécause their electrophoretic mobiliteis vary inversely with their molecu (B) DNAs of more than a few thous: cross-lined polyacrylamide gel. (C) Using gels with appropriately low agarose contents can fractionate te a weakly «) the larger chromosomal DNA of ptents. e) ield gel electrophoresis, Deww (Ay ine orange, ethidium ion, ete. (B) Molise test. «cy n blotting technique (D) Western blotting technique (BE) wrn blotting technique 4s in electrophoresis include charcoal ‘Scanned with CamScanner26. 27. About electr tA) Fates proteins ha stil A their ismetes i Sess tate peleetrie paints « % : can molecu (Dy ‘The medi sian U2). gc Medium is inypresnated with savdiuan duddeeyt sulphate Y Amphalytes are aseit in eres a ating plL gradients ti gel Filtration, (M1 Motecutes tre separated an has wy aarger molecules chute first, toll (CY AL motecuites, irrespective of el iD) is reluted to m of polarity ved! hy smaller al in the column lecular wig! ) 1 (EY Very large motecutes are eluted at the void volume Still on. iy 1B) lumina iy min (C) ‘The column is packed with septadey gel (D) His useful for separation of monosacch: (i) Tis used for fractionation of proteins On dialysis (A) Tt separates ms ay desall (cy) We (Dy Su membrane (E) The process is fact ecules from smatter molecules protein precipi rate mixtures of sniall jans ecules readily diffuse through the se rye ones are retained ted by stirring rer persulphate a ed sieving gel results if bisserylam dervlamide in the presence of ammonium persulphais (C) The degree of erosselinking is independent of nerylumide {b) Polyacrylamide gel eleetrophorests can be used far molecular weight determina (k) Incorporation of S le is mixed with 5 enhances detection of protein subunits Affinity chrom: : A) Isaspecial type of electrofocussing oH Exploits the naturaf attractlon/interaction between biomolecules {C)__ Depends on grass physical and chemical properties of blomolecutes ()) Isa type of adsorption chromatography (E) Makes use of agarose gel ography ae ‘Scanned with CamScenner28, Still on affinity chromatography (A) Typical interactions of interest here include antigen-a enzyme-substrate; and hormone-receptor bodys (B) Ligand attachment to the gel is via electrostatic forces: ; (C) Ligand attachment is achieved through simple chemical reactions @) ly specific and fast (E) Separation can be performed on paper 29. More on affinity chromatography, the adsorbents used include (A) Con A-sepharose (B) S*AMP-sepharose (C) CNBr-activated sepharose (D) Poly U-sepharase (E) — Epoxy-activated sepharose 30. Still on affinity chromatography (A) Typical ons of interest here include antigen-antibody; enzym nd hormone-receptor (B) Ligand attachment to the rostatic forces (C) Ligand attachment i simple chemical reactions (D) — Itishighly specific and fast (E) Separation can be perf ‘Scanned with CamScanner