Ljunglof and co-workers found that most adsorption takes place on the surface of the particles. After calculating the available surface area in a packed bed (with particles varying from 45 to 165 um in diameter) and in a sieved fraction (with particles 50-60 um in diameter) , the researchers determined that the smaller particles in the sieved fraction would provide more surface area and, thus, would allow more adsorption. Comparing a sieved fraction with a packed bed confirmed that the sieved fraction's increase in break-
Electrospray of large DNA The high mass accuracy and short analysis times inherent in electrospray ionization (ESI) MS make it an attractive method for analyzing nucleic acids. However, characterizing large polymerase chain reaction (PCR) products is a challenge because of primers, buffers, and detergents left after the reaction. Now, David C. Muddiman and colleagues at Virginia Commonwealth University show that double-stranded, 500 base-pair (bp) PCR products can be analyzed using ESI in conjunction with Fourier transform ion cyclotron resonance MS The researchers amplified a 500-bp region of DNA from the genome of the bacteriophage lamda. Five separate PCR reactions were pooled, and the combined sample was desalted and purified using the standard ethanol precipitation technique. The product was infused at a concentration of 0.8 pmol/uL, a flow rate of 200 nL/min, and a shutter pulse width of 1 s. Five acquisitions of —2.7 fmol of product were performed and averaged, resulting in a mass spectrum where adjacent charge states could be resolved from [M - 235H+]235" to [M-172H+]172". Based on these results and previous work, Muddiman and co-workers argue that the ability to obtain an ESI mass spectrum of a relatively large PCR product depends primarily on the removal of salt. They note that the spectrum shows no evidence of cation adduction, which would disperse the molecular ion signal over a wide range of m/z values, and no evidence of systematic shifts to higher m/z due to the production of Na+ adducts. The experi-
through capacity—the amount of plasmid DNA adsorbed when the effluent concentration reaches 5% of the initial concentration—corresponded to the increase in surface area. (J. Chromatogr., A 1999, 844, 129-35) Confocal images of plasmid DNA bound to the surfaces of adsorbent particles and visualized with YOYO-1 dye. (Adapted with permission. Copyright 1999 Elsevier Science..
mental mass was determined to be 309,406±27 Da; the researchers suggest that the uncertainty is due to the presence of blunt-ended, mono-adenylated, and diadenylated PCR products, which vary from
308,859.35 to 309,485.77 Da. Thus, they say the apparent error actually indicates detection of three unresolved products. {Rapid Commun. Mass Spectrom. 19919 13,1201-04)
K + chemosensor
ond compound, which contained only a The levels of sodium in biological syssingle 15-crown-5 group, to determine the tems are much higher than those of poimportance of having two crown groups tassium, making detection of K+ difficult on the sensor molecule. The singlebecause of competition with Na+. Russell crown sensor molecule exhibited no H. Schmehl, Chao-Jun Li, and co-workers change in luminescence intensity upon at Tulane University have developed a addition of K+ and Na+. possible solution to the problem—a The formation of K+ sandwich com+ highly selective K fluorescent chemoplexes is believed to involve at least two sensor from a bis-15-crown-5 derivative. double-crown sensor molecules; however, the definitive structure of the comThe sensor relies on the fluorescence plex has yet to be determined. Efforts are behavior of a distyryl benzene derivative under way to further characterize the attached to two 15-crown-5 groups. The complex. (J. Am. .oc. Chem. 1919,121, sensor molecule forms intermolecular 5599-600) sandwich complexes with K+, resulting in dramatic increases in fluorescence intensity with increasing K+ concentrations. In contrast, complexes with Na+ do not produce bridged sandwiches and little change in fluorescence is observed The researchers examined the fluorescence behavior of a sec(1) Double- and (2) single-crown sensor molecules.
Analytical Chemistry News & &eatures, September 1, 1991 5 8 7 A
