Chem 241 Final Exam PDF

Preview

Summary

Study Guide for Final Exam...

Description

1

Chapter 22 ● adsorption - separation based on solute adsorption on surface of stationary phase ● partition - separation based on solute dissolving into stationary liquid ● ion-exchange - separation based on charge ● size exclusion - separation based on molecular size ● affinity - separation based on antibody-antigen or enzyme-substrate interactions larger k’ means greater retention times. more solute is interacting with the stationary phase. Higher R means better separation. Two factors affect how well two components are separated: 1. difference in retention time. Increase y to make peaks further apart/closer to each other. 2. peak widths. Increase N/decrease H to make peaks wider/narrower. Theoretical plates (N) - number indicating how good a column is for separation. With a longer column, you have more plates, better resolution between two peaks. HETP = L/N. As HETP decreases, N increases. ^height of plate?yea theoretical plate want small H value and large N value to get best resolution. usually from a long, narrow, open tubular column Van Deemter Equation: HETP = A + (B/v) + Cv where v is the linear flow rate. Causes of band broadening: ● Eddy diffusion (A) - multiple paths a solute can take from top of column to bottom of column. reduced in HPLC by using small solute particles ● Longitudinal diffusion (B) - solute concentration is lowest at the edges of a band; solute diffuses to the edges ● Resistance to mass transfer ( C ) - finite time necessary to equilibriate ● Trade off between decreasing flow rate to increase resolution with C term or increasing flow rate to increase resolution with B term. Ideal flow rate is at Hmin. overloaded column - too much solute applied to the column such that the

2

stationary phase sites are now masked by solute. New solute does not see any stationary phase, just other solute which leads to poor separation. Tailing - solute interacts with stationary phase too strongly. Solute molecules aren’t being released at same K. they slowly come off. Get an abrupt rise of [S]s at the start of the band. Long tail of gradually decreasing solute after the peak. A common form of stationary phase is silica. when the silanol groups lose a proton, they are charged and that leads to tailing. Cap silanol groups to reduce tailing. The difference between K and D is that K has total concentrations of neutral species in each phase and D includes charged species too for molecules that have different chemical forms. K→ neutral D→ all phase 2 isn’t polar, do its only the [HA] of the organic phase. phase one is polar and undergoes a weak acid dissociation.

Chapter 23 (Gas Chromatography) LASER stands for Light Amplification by Stimulated Emission of Radiation retention index - relates the retention time of compound to those of linear alkanes (GC). Used to standardize the elution time of your solute compared to others. thermal conductivity - all analytes flame ionization - most hydrocarbons electron capture - halogens, CN, NO2, conjugated carbonyls, organometallics. nitrogen-phosphorus - compounds containing N and P photoionization - aromatic and unsaturated compounds sulfur chemiluminescence - S atomic emission- most elements (selected by wavelength) mass spectrometer - all analytes flame photometric - P, S, Pb, Sn, or other selected elements.

3

in case he asks us to draw it. where is this from??? not seeing it in hte book (they just have a shitty one with a box that says gas in, gas out, and injector/detector box) ^no clue, its been up on here for a few days now though Refractive index detector is useless in gradient elution. It can only be used for isocratic elutions. Packed column ● lower resolution ● small, uniform particle size decreases Eddy diffusion (requires higher pressures) Open tubular (has no A term) 8-9 silica dissolves (not cool) pH < 2 silica hydrolyzes (no charge which means no electroosmosis) 1. normal phase - polar stationary phase and less polar solvent (if you go to more polar solvent, increases eluent strength)... the least polar compound would be eluted first and the most polar compound last (the compound that is most similar in polarity to the stationary phase is eluted last) run a gradient elution usually from less→ more polar solvent to get longer-eluting solutes off the column. opposite for reverse phase chromatography 2. reverse phase - nonpolar stationary phase and solvent is more polar (if you move to less polar solvent, eluent strength increases) HPLC has a reduced A term on the VD plot (as opposed to regular LC) because HPLC uses extremely small particles that decrease the magnitude

5

of irregular flow paths -large particles only good for extremely concentrated solutes, but have poorer resolution (HIGHER A and C terms) Chapter 25 A greater charge leads to faster migration. electrophoresis - movement of charged molecules through an electric field. electroosmosis - the bulk flow of fluid in a capillary caused by migration of the dominant ion in the diffuse part of the double layer toward the anode or cathode. Sephadex - cross-linked Dextran used to separate molecules by size (SEC) Electrophoresis is based on differences in migration of charged ions in an electric field in solution. Migration rate depends on size of solute and charge of solute. Greater charges migrate faster. Large molecules are slower due to friction. Deprotonated silanol groups are what causes charge buildup at the walls of a glass capillary. Only have a B term for CE. if you coat the walls of the capillary with some compound (usually an amine) that will bond to the Si-OH groups and cause them to be positively charged, then negative charge will accumulate along capillary walls and electroosmotic flow will be towards the anode instead of the cathode. Electrophoresis can’t be reversed, cations always move towards the cathode and anions always move towards the anode. but you should consider magnitude of charge and friction coefficient when figuring out which will get there first. Molecular Exclusion ● larger molecules don’t fit in pores, so they elute faster. ● Large solutes, Vr = V0 and K = 0 ● small solutes, Vr = Vm, K = 1 ● intermediate: 0 < K < 1 ● K > 1 - analyte has adsorbed to sites on column (stationary phase). ● Vm = total volume of column ~ pi*r-squared*L

6

Ion-Exchange Chromatography - separation based on attraction between solute ions and charged sites bounds to the stationary phase. - anion exchangers: positively charged stationary phase - cation exchangers: negatively charged stationary phase Depolarized Water (i think)- purified with water to remove cations and anions and replace them with H+ and OH-. does not remove organic impurities, etc.

Chapter 17 Franck-Condon principle - electronic transitions are fast relative to nuclear motion so each atom has nearly the same position and momentum before and after a transition. S0 → S1 absorption S1 → T1 intersystem crossing S1 → S0 fluorescence S1→ S0 with no light emitted internal conversion T1 → S0 phosphorescence Fluorescence lifetimes are shorter than phosphorescence lifetimes because T1 to S0 transitions require a change in spin quantum number.

7

● microwave radiation - causes molecules to rotate (Think about a microwave. Its plate ROTATES while heating your food.) ● infrared radiation - causes molecules to vibrate, heat emitted during vibrational relaxation ● UV-Vis radiation - causes valence electron excitation ● X-rays - break chemical bonds and ionize molecules ● gamma ray, x-ray, UV, Vis, IR, microwave, radio. listed in order of decreasing energy (gamma has most energy).

Beer’s law - the amount of light absorbed by a sample is dependent on the path length, concentration of sample, and molar absorptivity (constant at a compound for at a given frequency or wavelength) Not accurate to measure absorbances in the 0.3-2 range because if absorbance is too high, then too little light reaches the detector for accurate measurement. If absorbance is too low, there is too little difference between sample and reference for accurate measurement. fluorescence - emission of a photon during a transition between states with the same spin quantum numbers phosphorescence - emission of a photon during a transition between states with different spin quantum numbers. Longer lifetimes b/c spin forbidden luminescence - light given off after a molecule absorbs light

8

chemiluminescence - light given off by a molecule created in an excited state in a chemical reaction. emission spectrum - constant excitation wavelength and variable emission wavelength excitation spectrum - variable excitation wavelength and constant emission wavelength Excitation spectrum looks like an absorption spectrum because the greater absorbance at the excitation wavelength, more molecules are promoted to excited state and more emission will be observed. Chapter 18 only 18-1 Chapter 19 LASER - light amplification by stimulated emission of radiation. Advantages of a double-beam spectrophotometer: 1. easy to scan multiple wavelengths 2. kinetics experiments - monitor absorbance as a function of time 3. reduced source and detector drift because of beam chopper Deuterium arc lamp - UV light Tungsten lamp - visible and near -IR light in fiber optics, light is carried by total internal reflectance. light can be: transmitted, reflected, absorbed, or scattered. Snell’s Law: n1sin(theta 1) = n2sin(theta 2) n is the refractive index, a measure of how much rays of light are bent by a medium. population inversion - greater population of particles in higher-energy excited state than a lower energy (ground) state -once one excited molecule emits a photon it bounces all around the tube (transmits nearly all light) and causes every excited molecule that it strikes to release a photon with the same wavelength/energy. this is why lasers produce beams of light that are monochromatic, polarizable (only deviate on one plane), spatially and temporally coherent, and very intense for a certain wavelength. Monochromators are wavelength selectors. R = nN

9

resolution - ability to separate two closely spaced peaks dispersion - measure of the ability to separate wavelengths differing by delta wavelength through the difference in the reflecting angle. smaller slit width increases resolution and decreases amount of light reaching the detector. larger slit width gives higher--you want a higher-- signal-noise ratio and if too much light is let in detector may become saturated. photodiode array - records entire spectrum, has rows of p-type silicon on a substrate of n-type of silicon to create pn junction diodes, uses a reverse bias to draw e- and holes away from junction, CCD- most sensitive detector. rows for n type Si substrate capped with p type Si to create pixels, each pixel can hold up to 10^5 electrons, electrons that are promoted to the conduction band from the valence band collect beneath the positive electrode then are moved up and to the right and then counted. Pregnancy Tests - early detection is important. results in lifestyle changes ● Ancient Egyptians used urinalysis. Water bags of wheat and barley. Germination meant pregnancy and grain type indicated gender.987878765654iuy ● medieval urology - analysis by sight and smell ● Hippocrates - honey and water before bed. Pain and cramps meant you were pregnant. ● IU quantifies a substance based on biological activity. used for vitamins, hormones, drugs. cannot be converted to a mass. use it because all hormones that indicate pregnancy are detectable or measurable) with it. ● Pregnancy in a nutshell: FSH → LH → progesterone → fertilization, implantation of fertilized egg. Trophoblast cells secrete hCG to continue stimulation of corpus luteal progesterone. hCG is a dimeric glycoprotein. It is produced by trophoblast cells in the placenta during pregnancy. ● hCG= human chorionic gonadotropin ● OTC Kits are QUALITATIVE, not quantitative

10

● ● ● ● ● ●

● ●

● ● ●

‘sandwich’ assay: Ab2-hGC-Ab1* ELISA - enzyme-linked immunosorbent assay epitope - particular feature on another molecule hCG doubles every 2-3 days until 8 weeks. the progesterone that it stimulates the production of is the cause of “morning sickness.” labeled monoclonal Ab binds hCG (enzyme label is QUALITATIVE ANALYSIS) polyclonal Ab immobilized on support binds complex ○ sample wick - Ab1* binds all hCG in sample. Ab1* has a qualitative enzyme label. ○ test line - binds all hCG (in hcG-Ab1* complex) color change is a result of Ab1* proximity to substrate in support. ● “sandwich assay” created once all hCG has been bound to Ab1* and Ab2 ○ control line - Ab3 binds unbound Ab1* but Ab1* presence still causes the color change ○ color change always results from Ab1* proximity to substrate in support on the test line, and the control line. low levels of hcG could be miscarriage while high levels could mean multiple births. may get false positive- hCG produced by some trophoblast cells in response to disease. or, egg implantation begins but later fails. or take steroids/hormones with it. false positives are harmless, false negatives don’t result in lifestyle change and can be harmful to mother and child. false negatives are result of improper procedure or premature testing. myths- longer reading times enhance sensitivity, basal hCG levels can trigger false positive, blood tests more sensitive, darker lines indicate hCG doubling.

Professor Schoenfisch’s Research- glucose sensors. Remember Nitric Oxide - The body produces NO in response to pathogens. he talked

11

about biosensors are implanted into the body to directly measure glucose levels in the blood but it is treated as a foreign body so inflammation begins. coating the biosensor with NO reduces inflammation, up until the coated NO runs out... sepsis - disease that affects people who go to hospital because they have some other problem. Catheters promote bacterial adhesion. Sepsis is the 10th leading cause of death in the US and you have a 1 in 3 chance of dying if you develop it. one of his goals is to find a way to measure NO release to better predict sepsis DOES LOTS AND LOTS OF RESEARCH EXPERIMENTS WITH ETHANOL ● works with subcutaneous (beneath skin) glucose sensors ● developing internal electrochemical sensor is far more challenging than developing an external one. must be small, fast, and BIOCOMPATIBLE (doesn’t lose sensitivity as scar/scabs form, body doesn’t try to expel/isolate it, etc). ● why?- able to catch abrupt changes in important parameters (as opposed to just randomly checking during the day, because you may miss something important). ● helps diabetics, physicians, etc. all about REAL TIME RESULTS (glucose levels for diabetics, sepsis in hospitals) ● problems- formation of thrombus (clot/platelets) or blood vessel constriction over surface of sensor. ● solution- create sensor with biofilm that will release NO in order to prevent platelet adhesion/vasodilate (widen) surrounding blood vessels ● first attempt led to leaching of a toxic molecule, was unsuccessful ● now working with sol-gels, attempting to find one that will release NO to increase 6yubiocompatibility ● NO reduces inflammation, scarring, biofouling, infection (natural immune response) ● sensors may help detect sepsis faster ● also attempting to find a better way to measure NO levels, which fluctuate based on the bodies immune response, to better predict sepsis...