Laser ratio data standardization

Figure 2 shows survey Raman spectra of the hcmopolymers, poly(methyl methacrylate)(PMMA.), poly(3-oximino-2-hutannone methacrylate)(pom), and poly(methacrylonitrile)(PMAN), and one terpolymer(P(M-0M-CN)) with a S/N ratio of about 10 1. Each of the polymers has a band specific to that polymer 8l2 dcm-1 (vg (C-O-C) for IMMA), 1622 hem" (Vg(C=N) for POM), and 2237 dcm l(vg(CHN) for PMAN). Additionally, there is an asymmetric C-H bending mode at 1 53 Acm l, common to all three homopolymers, which serves as an internal standard. These bands are indicated by arrows in Figure 2. A broad fluorescence background is evident, but it can be reduced to acceptable levels by exposure to high laser power for 10-30 minutes, depending on the sample. Residual background fluorescence may be due to the oximino chromophore itself. Figure 3 depicts an example of actual data for a 75 15 10 terpolymer with a S/N ratio of about 50 1.

Figure 7. Laser-induced absorbance changes at 561 nm as a function of time in detergent solubilised bovine rhodopsin (X) and isorhodopsin (9) at room temperature. Bathorhodopsin is the only intermediate during the bleaching of bovine rhodopsin known to absorb strongly at 561 nm. The energy of the 530-nm pump pulse was about 10 4J the energy of the 561-nm probe pulse was about 10 7J. The beam sizes were about 1 mm2 for the pump and 0.5 mm2 for the probe. The samples (about 1.5 mL) were held in 0.5-cm cuvettes. The concentrations were about 4 A cm I at the absorption peaks near 500 nm the ratios A Ajjj were about 0.3 and ratios ASsa. As<)o were about 0.7 for rhodopsin and 0.5 for isorhodopsin. Each data point shown is the average of six (rhodopsin) and nine (isorhodopsin) laser shots. Typical mean standard deviations are 0.03. The zero time is located using a 0.5 cm CS2 Kerr optical shutter at the sample site. The half width at half maximum for the CS2 shutter prompt response curve is about 6 ps.

MALDI-TOF was done on a Kratos Kompact Maldi III mass spectrometer fitted with a standard 337 nm nitrogen laser and operated in the linear mode at an accelerating voltage of 20 kV. The matrix used was a-cyano-4-hydroxyciimamic acid (33mM in acetonitrile/methanol, premade from BRS) at a ratio of 1 1 with purified peptide samples. MALDI sample slides were loaded with O.S-1.0 uL of matrix/sample mixture (estimated 1-10 pmol peptide). The data was reprocessed using the Kratos software provided with the instrument. Theoretical masses were determined by utilizing a spreadsheet in which individual peptide masses were added to all possible caibohycfrate forms these masses were then compared to the observed masses to identify structures consistent with the mass results obtained. 

---