MS Interpretation

Two approaches are often used to improve the detection limit, including selected ion monitoring (SIM) and multiple reaction monitoring (MRM). In LC/MS studies, it is often desirable to increase detection sensitivity by hmit-ing the mass analyzer scan to just one ion— that is, SIM. In this mode, a single ion of interest is monitored continuously by a mass spectrometer and no other ions are detected. This results in signihcant improvement of signal-to-noise ratio. SIM trades specihcity for sensitivity. In general, the sensitivity in SIM is increased by a factor of 100 to 1000 over full-scan mass spectra. This can be quite useful in detection and quantihcation of specihc compounds at low levels. 

Another related technique, MRM, involves a MS/MS experiment that is often performed in a triple quadrupole mass spectrometer. Typically, Q1 is set to pass the parent ion and Q2 is used as a colhsion cell to fragment the parent ion. The third quadrupole is to transmit only diagnostic product ion from the parent ion. The advantage is the increased specihcity from MS/MS experiment. MRM provides the highest-duty cycle scan in a triple quadrupole and is widely used in quantitating multiple analytes in a complex matrix (examples are given in the section on pharmacokinetic studies of drugs). 

Mass spectrum interpretation is essential to solve one or more of the following problems establishment of molecular weight and of empirical formula detection of functional groups and other substituents determination of overall structural skeleton elucidation of precise structure and possibly of certain stereochemical features. As detailed in the previous sections, ESI and APCI are two of the most effective interfaces for LC/MS that have been developed. Thus, the focus of the discussion will be on interpretation of mass spectra obtained by ESI or APCI. 

The qualitative applications of mass spectrometry are based on the determination of the mass-to-charge m/z) ratio of an analyte. ESI and APCI MS are well known as a soft ionization technique. This means that relatively small amount of energy are needed to ionize the analyte. The efficiency of ion formation depends on a molecule s ability to associate and carry a charge. In a positive ion experiment, this ionization process can be defined by the following simple protonation reaction  

1 Pseudomolecular Ions. In contrast to the traditional MS, the highest mass peaks in ESI/APCI spectra are not always the molecular ion of interest. Instead, pseudomolecular ions, or noncovalent complex ions, are commonly observed. The pseudomolecular ions are generally formed by the analyte-adduct interaction in the solution system that is preserved as a result of the soft ionization of the ESI/APCI process. These ions are also formed by analyte-adduct gas-phase collisions in the spray chamber [49]. The exact mechanisms of how the analyte adducts are formed in ESI/APCI still remain unresolved at this point. More often than not, the adduct ion formation is a major cause for the low detection limit for ESEAPCI MS. However, these associative processes have also created interest in the study of drug-protein/ drug-oligonucleotide gas-phase complexes that benefit from the ability of ESI/APCI MS analysis. 

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The book is divided into four parts. Part I, The Fundamentals of GC/MS, includes practical discussions on GC/MS, interpretation of mass spectra, and quantitative GC/MS. Part II, GC Conditions, Derivatization, and Mass Spectral Interpretation of Specific Compound Types, contains chapters for a variety of compounds, such as acids, amines, and common contaminants. Also included are GC conditions, methods for derivatization, and discussions of mass spectral interpretation with examples. Part III, Ions for Determining Unknown Structures, is a correlation of observed masses and neutral losses with suggested structures as an aid to mass spectral interpretation. Part IV, Appendices, contains procedures for derivatization, tips on GC operation, troubleshooting for GC and MS, and other information which are useful to the GC/MS user. Parts I to III also contain references that either provide additional information on a subject or provide information about subjects not covered in this book. 

The integrated ion current for each analyte ion listet in Table 5 must be at least 2.5 times background noise and must not have saturated the detector. The internal standard ions must be at least 10.0 times background noise and must not have saturated the detector. However, if the M-[COCl]+ ion does not meet the 2.5 times S/N requirement but meets all the other criteria listed in Section 11 and in the judgement of the GC/MS Interpretation Specialist die peak is a PCDD/PCDF, the peak may be reported as positive and the data flagged on Form I. 

Gas chromatography coupled to mass spectrometry is a widely used analytical method [31] but does require prior reduction of the involatile N-oxides. Additionally, there are problems associated with some alkaloids that require prederivatization to enhance the GC characteristics. These requirements for prederivatization can adversely affect the GG-MS interpretation, especially at trace levels of alkaloids [32]. Gombined SPE-LC-MS approaches have provided methods for qualitative profiling and quantitative analysis of pyrrolizidine alkaloids and their N-oxides. 

The masses shown in Table 6.1.1 and other similar tables in this book represent only the mass obtained from the most abundant isotopes rounded to the unit. This mass is typically used for MS interpretations [58]. 

Directive 2001/20 has also led to a set of detailed Guidance documents, to help MS interpret the Directive and implement appropriate ethical committee oversight. The Detailed. Guidance on the Application Format and Documentation to be Submitted in an Application for an Ethics Committee Opinion on a Clinical Trial for a Medicinal Products for Human Use (April 2003) is self-explanatory. This document includes a table that collates each special requirements of MS, and it is designed to be used with the national Guidances, where they exist. 

In order to avoid problems with sample inhomogeneity, the entire oil sample from each sample of shale was dissolved in 1.5 to 2.5 mL of CS2 (about 1 g oil to 1.5 mL solvent). One pL of this solution was injected into a Hewlett-Packard Model 5880 Gas Chromatograph equipped with capillary inlet and a 50 m x 0.25 mm Quadrex "007" methyl silicone column. Injection on the column is made with a split ratio of approximately 1 to 100. The column temperature started at 60°C and increased at 4°C/min to 280°C where it remained for a total run time of 90 min. The carrier gas was helium at a pressure of 0.27 MPa flowing at a rate of 1 cm /min. The injector temperature was 325°C and the flame ionization detector (FID) temperature was 350°C. Data reduction was done using a Hewlett-Packard Model 3354 Laboratory Automation System with a standard loop interface. Identification of various components was based on GC/MS interpretation described previously (4). For multiple runs on the same shale, the relative standard deviations of the biomarker ratios were about 10%. 

Introduction to NRPS and PKS Biosynthesis New Tools in the Characterization of Multidomain and Phosphopantetheinylated Proteins A Brief Introduction to FT-ICR-MS Interpretation of FT-ICR-MS LC-FT-ICR-MS Analysis of NRPS and PKS Proteins The Phosphopantetheinyl Ejection Assay How Is the PEA Accomplished ... 

The MST/EPA/NIH Mass Spectral Library 1998 database ( www.nist.gov/ srd/analv.htm) is the product of a muftiyear, comprehensive evaluation and expansion of the world s most widely used mass spectral reference library, and is sold in ASCII or Windows versions. It contains 108,000 compounds with electron ionization spectra, chemical structures, and molecular weights. It is available with the NIST MS Search Program for GC/MS deconvolution, MS interpretation, and chemical substructure analysis. The NIST chemistry WebBopk ( http //webbook.nist.gov) is a. free online system that contains the mass spectra of over 12,000 compounds (this Standard Reference Data Program also has IR and UV-Vis spectra). 

A quadrupole type gc-ms (16) was used for component identification In nearly all Instances. A few components were Isolated by preparative gc for infrared and nuclear magnetic resonance spectrometrlc examination. The retention behaviors of all tentatively-identified constituents were checked, using authentic samples, in order to verify the ms interpretations. 

Insect cuticular hydrocarbons are commonly identified on the basis of retention indices (Nelson Blomquist, 1995). Pomonis et al. (1989) determined the Kovats retention indices of monomethyl-jrentacosanes, some internally branched dimethylalkanes and 2,x-dimethylheptacosanes on a cross-linked methyl silicone fused silica capillary column (Hewlett-Packard). Carlson et al. (1998) described a protocol for the identification of methyl-branched hydrocarbons in insect cuticular waxes. In this protocol, programmed-temperature retention indices are assigned to peaks, then the patterns in GC peaks that probably contain homologues are marked to assist subsequent GC-MS interpretation. The authors also included data from the literature covering most of the insect methylalkanes. 

The NIST library is available with a new version of the NIST Mass Spectral (MS) Program (v.2.0g) and the enhanced versions of MS Interpreter and AMDIS, the mass spectral interpretation tools with thermodynamics-based interpretation of fragmentation and chemical substructure analysis. The binary format has not changed from the 2002 version, although several new files have been added that associate equivalent compounds and link individual compounds to the RI library. Raw data files are provided in both an SDFile format (structure and data together) as well in earlier formats. The SDFile format holds the chemical structure as a MOLFile and the data in a simple ASCII format. The NIST MS Search Program is also part of many commercial instrumental GC-MS software suites. 

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