Identification of unknowns

Although there are millions of organic compounds that an organic chemist might be called on to identify, the scope of this experiment is necessarily limited. In this textbook, about 500 compounds are included in the tables of possible unknowns given for the experiment (see Appendix 1). Your instructor may wish to expand the list of possible unknowns, however. In such a case, you will have to consult more extensive tables, such as those found in the work compiled by Rappoport (see References). In addition, the experiment is restricted to include only seven important functional groups  

Even though this list of functional groups omits some of the important types of compounds (alkyl halides, alkenes, alkynes, aromatics, ethers, amides, mercaptans, nitriles, acid chlorides, acid anhydrides, nitro compounds, and so on), the methods introduced here can be applied equally well to other classes of compounds. The list is sufficiently broad to illustrate all the principles involved in identifying an unknown compound. 

In addition, although many of the functional groups listed as being excluded will not appear as the major functional group in a compound, several of them will frequently appear as secondary, or subsidiary, functional groups. Three examples of this are presented here. 

The groups included that have subsidiary status are 

The experiment presents all of the chief chemical and spectroscopic methods of determining the main functional groups, and it includes methods for verifying the presence of the subsidiary functional groups as well. It will usually not be necessary to determine the presence of the subsidiary functional groups to identify the unknown compound correctly Every piece of information helps the identification, however, and if these groups can be detected easily, you should not hesitate to determine them. Finally, complex bifunctional compounds are generally avoided in this experiment only a few are included. 

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When the gas chromatograph is attached to a mass spectrometer, a very powerful analytical tool (gas chromatography-mass spectrometry, GC-MS) is produced. Vapour gas chromatography allows the analyses of mixtures but does not allow the definitive identification of unknown substances whereas mass spectrometry is good for the identification of a single compound but is less than ideal for the identification of mixtures of... 

Identification of unknown compounds in solutions, liquids, and crystalline materials characterization of structural order, and phase transitions... 

TLC measurements used for the identification of unknown solutes depends on two basic pai ameters. Firstly, the distance traveled by the solvent front, measured from the sampling point or sampling boundary, and secondly, on the distance traveled by the spot from the sampling point or sampling boundary. These are the sole... 

J. Ogorka, G. Schwinger, G. Bmat and V. Seidel, On-line coupled reversed-phase high-performance liquid cliromatography-gas chromatography-mass specti ometi y , A powerful tool for the identification of unknown impurities in pharmaceutical products , J. Chromatogr. 626 87-96 (1992). 

H. Bagheri, J. Slobodnik, R. M. Marce Recasens, R. T. Ghijsen and U. A. Th Brinkman, Liquid cliromatography-particle beam mass specti ometiy for identification of unknown pollutants in water , Chmmatogmphia 37 159-167 (1993). 

Identification of unknowns, as well as confirming the presence of known AAs, is more reliable if accurate mass measurement... 

Identification of unknowns using GC/MS is greatly simplified if accurate mass measurements are made of all the ions in a spectrum so that reasonable elemental compositions of each ion are available. Unfortunately, obtaining a mass measurement that is accurate enough to significantly limit the number of possible elemental compositions requires expensive instrumentation such as a double-focusing magnetic sector or fourier transform ICR MS. 

The Use of PLC for Isolation and Identification of Unknown Compounds from the Frankincense Resin (Olibanum) Strategies for Finding Marker Substances... 

Study type Required pre- or post-registration Use of radiolabel Identification of unknowns Need for mass balance Importance of low detection limits Importance of adjustable sampling frequency... 

The computational prediction of vibrational spectra is among the important areas of application for modem quantum chemical methods because it allows the interpretation of experimental spectra and can be very instrumental for the identification of unknown species. A vibrational spectrum consists of two characteristics, the frequency of the incident light at which the absorption occurs and how much of the radiation is absorbed. The first quantity can be obtained computationally by calculating the harmonic vibrational frequencies of a molecule. As outlined in Chapter 8 density functional methods do a rather good job in that area. To complete the picture, one must also consider the second quantity, i. e., accurate computational predictions of the corresponding intensities have to be provided. 

NMR provides one of the most powerful techniques for identification of unknown compounds based on high-resolution proton spectra (chemical shift type integration relative numbers) or 13C information (number of nonequivalent carbon atoms types of carbon number of protons at each C atom). Structural information may be obtained in subsequent steps from chemical shifts in single-pulse NMR experiments, homo- and heteronuclear spin-spin connectivities and corresponding coupling constants, from relaxation data such as NOEs, 7) s 7is, or from even more sophisticated 2D techniques. In most cases the presence of a NOE enhancement is all that is required to establish the stereochemistry at a particular centre [167]. For a proper description of the microstructure of a macromolecule NMR spectroscopy has now overtaken IR spectroscopy as the analytical tool in general use. 

Applications Desorption chemical ionisation has proven potential in the analysis of thermally labile, nonvolatile and polar compounds [40,67,68], for the identification of unknown polymers and the study of the thermal degradation mechanisms of polymers. Considering the overall ease of DCI operation, the capability of analysing nonvolatile compounds, and the selectivity provided by choosing different reagent gases, DCI has found surprisingly few practitioners in the analysis of polymer additives. 

The main characteristics of electrospray ionisation are given in Table 6.21. Electrospray ionisation is a suitable technique for producing accurate molecular mass information on a wide range of low-MW samples. ESI-MS is particularly appealing for polar, high-MW samples (more than ca. 1000 Da), where the multiply charged ions formed have m/z values within the range of the spectrometer. However, ESI presents some problems in the identification of unknowns ... 

Applications With the current use of soft ionisation techniques in LC-MS, i.e. ESI and APCI, the application of MS/MS is almost obligatory for confirmatory purposes. However, an alternative mass-spectrometric strategy may be based on the use of oaToF-MS, which enables accurate mass determination at 5 ppm. This allows calculation of the elemental composition of an unknown analyte. In combination with retention time data, UV spectra and the isotope pattern in the mass spectrum, this should permit straightforward identification of unknown analytes. Hogenboom et al. [132] used such an approach for identification and confirmation of analytes by means of on-line SPE-LC-ESI-oaToFMS. Off-line SPE-LC-APCI-MS has been used to determine fluorescence whitening agents (FWAs) in surface waters of a Catalan industrialised area [138]. Similarly, Alonso et al. [139] used off-line SPE-LC-DAD-ISP-MS for the analysis of industrial textile waters. SPE functions here mainly as a preconcentration device. 

Separation and positive identification of unknown components (fingerprinting capability)... 

Positive identification of unknown compounds (library searchable EI/CI spectra)... 

Various LC-PB-MS and LC-APCI-MS comparisons have been reported on polymer additive extracts [540, 563,629,630]. The complementary character of the El and APCI modes was confirmed. Yu et al. [630] compared LC-PB-MS and RPLC-UV-APCI-MS for detection and identification of unknown additives (in the 252 to 696 Da range) in an acetonitrile extract from PP (containing Irganox 1076, Naugard XL-1 and a degradation product, NC-4, 3-(3,5-di-f-butyl-4-hydroxyphenyl) propanoic acid, 7,9-di-f-butyl-l-oxaspiro [4,5] deca-6,9-diene-2,8-dione and octadecanol-1). Comparison was based on El data (identification of chemical structure), APCI (MW information CID spectrum with limited fragmentation) and PDA (210 nm). The components were identified by El and confirmed by APCI- (with better sensitivity and linearity) MS and PDA showed... 

Whereas the components of (known) test mixtures can be attributed on the basis of APCI+/, spectra, it is quite doubtful that this is equally feasible for unknown (real-life) extracts. Data acquisition conditions of LC-APCI-MS need to be optimised for existing universal LC separation protocols. User-specific databases of reference spectra need to be generated, and knowledge about the fragmentation rules of APCI-MS needs to be developed for the identification of unknown additives in polymers. Method development requires validation by comparison with established analytical tools. Extension to a quantitative method appears feasible. Despite the current wide spread of LC-API-MS equipment, relatively few industrial users, such as ICI, Sumitomo, Ford, GE, Solvay and DSM, appear to be somehow committed to this technique for (routine) polymer/additive analysis. 

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