Mass spectrometers 240 INDEX

Specifications for modem detectors in HPLC are given by Hanai [538] and comprise spectroscopic detectors (UV, F, FUR, Raman, RID, ICP, AAS, AES), electrochemical detectors (polarography, coulometry, (pulsed) amperometry, conductivity), mass spectromet-ric and other devices (FID, ECD, ELSD, ESR, NMR). None of these detectors meets all the requirement criteria of Table 4.40. The four most commonly used HPLC detectors are UV (80%), electrochemical, fluorescence and refractive index detectors. As these detectors are several orders of magnitude less sensitive than their GC counterparts, sensor contamination is not so severe, and... 

The sensors in this section can also be utilized to detect chemicals in liquid through the bulk solution refractive index change induced by the presence of target chemicals. Since no recognition molecules are used, this type of chemical sensing may usually have low specificity. However, these sensors may perform excellently in conjunction with other technologies such as capillary electrophoresis, mass spectrometer, and liquid chromatography in chemical detection. 

A simple convention for description of peptide fragments formed in the mass spectrometer was proposed by Roepstorff and Fohlman in 1984 [1] and further modified by Johnson in 1987 [2]. Fragment ions are described by single lower-case letters with additional indexes (Fig. 6.5). If we consider fragmentation of a peptide backbone only, six ion series can be formed due to fragmentation at ... 

The development of mass spectroscopic techniques such as matrix assisted laser desorption (MALDI) and electrospray mass spectrometry has allowed the absolute determination of dendrimer perfection [7,8], For divergent dendrimers such as PAMAM and PPI, single flaws in the chemical structure can be measured as a function of generation to genealogically define an unreacted site of or a side reaction producing a loop at a particular generation level. Mass spectromet-ric results on dendrimers, not only demonstrate the extreme sensitivity of the technique, but also demonstrate the uniformity of the molecular mass. The polydispersity index of Mw/Mn for a G6 PAMAM dendrimer can be 1.0006 which is substantially narrower than that of living polymers of the same molecular mass [7],... 

Manufacturers publish their product s performance characteristics as specifications, which are often used by the customer for comparison during the selection process. Table 1 shows the specifications of an Agilent 1100 Series Quaternary Pump, which is quite representative of other high-end analytical pumps. Note pulsation is particularly detrimental to the performance of flow-sensitive detectors (e.g., mass spectrometer, refractive index detector). Differences in dwell volumes and composition accuracy between HPLC systems might cause problems during method transfers. 

Methylation analysis was run by the method of Hakomori ( ), followed by hydrolysis with trifluoracetic acid, sodium borohydride reduction, and acetylation. 6LC was performed on a Hewlett-Packard 5970, used as an inlet for a mass spectrometer. Molecular weight was determined on a Sephacryl S-500 column (2.6 x 70 cm), using deionized water as solvent, upward flow, 2.75 ml/min, and detection by refractive index monitor. Model R-401 (Waters Associates). 

Since the development of HPLC as a separation technique, considerable effort has been spent on the design and improvement of suitable detectors. The detector is perhaps the second-most important component of an HPLC system, after the column that performs the actual separation it would be pointless to perform any separation without some means of identifying the separated components. To this end, a number of analytical techniques have been employed to examine either samples taken from a fraction collector or the column effluent itself. Although many different physical principles have been examined for their potential as chromatography detectors, only four main types of detectors have obtained almost universal application, namely, ultraviolet (UV) absorbance, refractive index (RI), fluorescence, and conductivity detectors. Today, these detectors are used in about 80% of all separations. Newer varieties of detector such as the laser-induced fluorescence (LIE), electrochemical (EC), evaporative light scattering (ELS), and mass spectrometer (MS) detectors have been developed to meet the demands set by either specialized analyses or by miniaturization. 

Fig. 4.5.9 HPLC and mass-spectromet-ric analysis of transferrin-linked oligosaccharides. Transferrin is purified from sera of a control and the index CDG-IId patient. Oligosaccharides are released by N-glycosidase F digestion and subsequently analysed by HPLC. Peak fractions of the control and the patient are further investigated by mass spectrometry and are compared to oligosaccharide standards. Values above the HPLC peaks indicate the detected masses...

There are many types of HPLC detectors available today with the most popular ones including UV and UV-photodiode array (PDA), fluorescence, refractive index, evaporative light scattering (ELSD), charged aerosol (CAD), and the mass spectrometer. Of these, the most commonly used detector for pharmaceutical analytical methods is the UV detector since a majority of pharmaceutical compounds have some type of chromophore. Multiple detectors in series can also be utilized in order to obtain more information per chromatographic run. For example, a PDA detector can... 

A refractive index detector can see both of these types of compounds. A variable UV detector at 206 nm or 195 nm will work for both at reasonable concentrations. I would recommend either a CAD (see Chapter 5) or a mass spectrometer (see Chapter 15) if you need to do gradients or high-sensitivity detection. Both are expensive but give good results. The MS will give you molecular weight data for your separated peaks as well. 

Several languages are focusing on more specialized areas of physical measurements. MatML (http //www.matml.org/) has been developed for the exchange of materials information. The HUPO proteomics standards initiative (http //www.psidev.info/) is developing several standards including mzML (http //www.psidev.info/index.php q=node/257), a standard for encoding raw data from mass spectrometers that builds on the previous formats DataML (http //www.psidev.info/index.php q=node/80) and mzML (Pedrioli et al. 2004). Another HUPO standard, PSI MI XML, provides syntax for the description of molecular interactions (http //www.psidev.info/index.php q=node/31). 

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