labs

Question 1

define spectrophotometry

Answer 1

used to measure conc of specific substances w/i body fluids

Question 2

how does spectrophotometry work

Answer 2

EM radiation of particular freq passed through sample

under ordinary conditions, atoms exist in ground energy state and via incident radiation, e-s in outer shell jump to higher energy level. Each jump is unique and requires absorption of specific amount of energy

Transmitted radiation analyzed in terms of intensity at different frequencies by showing the relationship between wavelength of radiation stored and stimulated transmission on absorption spectrum

Question 3

why is acid erythrogram used

Answer 3

to determine historical changes in number of cells that are hemolyzed under influence of weak acid like HCl

Question 4

how to study acid hemolysis

Answer 4

the H+ from HCl will penetrate PM of erythrocytes via protonation of Hb and protein membranes = osmotic destruction
We use spectophotometry to find extinction at time intervals to find change in extinction and % hemolysisi

Question 5

acid erythrogram of a healthy individual

Answer 5

left = older cells that 1st undergo hemolysis since they have low diameter/area/vol
right= young cells that undergo lysis last and have high sphericity index

Question 6

define hemosomes

Answer 6

liposomes containing entrapped Hb

Question 7

describe liposomes

Answer 7

stable, closed systems, used to administer different drugs:

unilamellar liposomes fuse their membranes w/ target cell membrane and introduce contents directly into cytosol

multilamellar liposomes fuse w/ plasmalemma of target cells so liposomes inject content w/o outermost bilayer

substances can be taken up by adsorption as liposomes attach to cell surfaces via electrostatic attraction

Question 8

preparation of hemosomes

Answer 8

1. lipids dissolved and mixed in solvent of chloroform:methanol to a homogenous mixture of lipids
2. solvent is removed by evaporation via dry nitrogen steam for small vol of solvent, for larger vol, at reduced pressure via rotary evaporation = this forms thin lipid film on sides of flask
3. lipid film dried to remove residual solvent in vacuum pump overnight
4. dry lipid film hydrated by adding buffer w/ water
5. spin flask in warm water bath to hydrate lipid, temp must be above phase transition temp
6. 1 hr hydration time = vigorous shaking, mixing, stirring = lipids self organized into closed vesicles as small amounts of solvent is entrapped

Question 9

define molecular chromatography

Answer 9

separates and purifies components according to size and volume in mixture of molecules containing 2 phases, immobile and mobile phase

separation is based on ability of sample molecules to penetrate highly porous bead-like structure of stationary phase

Question 10

define thin layer chromatography

Answer 10

used to separate non-volatile/dissolved substances by their migration.

stationary phase as silca/alumina gel applies to glass in thin later, gypsum (inert binder) added to maintain phase

mobile phase = suitable mix of solvents like acetic acid or alcohol

Question 11

Kd

Answer 11

distribution coefficient sample conc in phase 1/sample cone in phase 2 = V1/V2

Question 12

Rf

Answer 12

Retardation factor, distance run by solute/distance run by solvent front

Question 13

electrophoresis

Answer 13

movement of charged particles relative to liquid it’s suspended in under influence of electric field

Question 14

microelectrophoresis

Answer 14

a type of electrophoresis using zeta potential since it is more accessible than surface and stern potential

Question 15

zeta potential

Answer 15

potential at surface of shear plane between particle w/ ion cloud and surrounding medium

Question 16

hemodialysis

Answer 16

life-sustaining procedure for patients w/ end-stage renal disease

extra-corpeal process, filters waste products by removing extra fluid from blood, restores proper electrolyte balance and eliminates edema from body

Question 17

hemodialysis circuit

Answer 17

hemodialyses, dialysate fluid, permanent vascular access

artificial membrane tube immersed in large vol of dialysate, blood pumped through tubing and back into vein. The membrane only allows blood solutes to pass out, no proteins or cells

Question 18

hemodialysis fluid flows via

Answer 18

countercurrent flow, opposite to blood flow, to maintain conc gradient of solutes for max efficiency

Question 19

ultrafiltration

Answer 19

hemodialysis fluid removal from blood by altering hydrostatic pressure of dialysate fluid

Question 20

t0.5

Answer 20

time from which dialysis onset at which initial sol conc of urea decreases 2 fold, prescribes the adequate does of dialysis

Question 21

describe Cu

Answer 21

trace element, co-factor for enzymes EG cytochrome oxidase, Cu-Zn-superoxide dismutase

Question 22

is Cu good or bad for the body

Answer 22

can be bad, since an XS of Cu leads to Wilson’s disease and pathogenesis of Alzheimer’s

Cuprous ion participates in formation of ROS = damages lipids, NA, proteins and carbohydrates

Cu+ also leads to oxidative injury by and produces cupric ions and .OH to induce oxidaton of LDL/HDL by Cu+ = promotes atherosclerosis

Question 23

how can we study Cu generation in membranes

Answer 23

during oxidative injury cuprous ions + hydrogen peroxide produce cupric ions and .OH.

We can add adrenaline as it can measure superoxide anion radical formation = oxidizes to adrenochrome = colour = so we use spectrophotometry

Question 24

how to determine catalytic activity of Fe and Pb in lipid peroxidation of guinea pig liver

Answer 24

Fe involved in FR mediated oxidations = Haber-Weiss reactions that will produce SOR involved in LP

MDA is end product of LP, adding TBA/Thiobarbituric acid = will produce a colour= measure the extinction using spectophotometry

Question 25

define optical activity

Answer 25

ability of substances to rotate plane of polarized light passing through

Question 26

how to study acid-catalysed sucrose hydrolysis

Answer 26

sucrose will be hydrolysed to sucrase, since all sugars are optically active, we measure this reaction using polarimetry to accurately study sucrose inversion by observing angle of rotation of polarised light at regular time intervals