Two dimensional TLC

Naturally, several other possibilities can be used to increase the number of dimensions. Between the first and second developments, or sample, the characteristics of the chromatographic plate or the properties of the sample can also be modified. Although interfacing of on-line OPLC with one- or two-dimensional TLC is not particularly difficult, it is not yet widely practiced. It must be concluded that full exploitation of the versatility of MD-PC is at an early state of development as a consequence several significant changes in practice might be expected in the next few years (10). 

Two-dimensional TLC on silica gel G has been used to identify alcohol ether sulfates in liquid laundry detergents. The spots of the chromatograms were examined by UV, IR, and NMR spectroscopy and the spectra compared with those of standard surfactants [283]. 

Propyl ether is the solvent of choice, and two-dimensional TLC with this and a chlorinated solvent mixture gives an exclusive sultone area as well as added sensitivity. 

Pyridine, pyrrole, indole, quinoline and isoquinoline alkaloids Silica gel G Detection by two-dimensional TLC [11]... 

Paper chromatography (PC) and thin layer chromatography (TLC) have been used since the 1940s. Preparative PC on Whatman 3 paper, analytical PC on Whatman 1 paper, and analytical TLC on microcrystalline cellulose, silica gel, or polyamide have been applied with a variety of solvents and the behaviors of anthocyanins have been similar in all media. Two-dimensional TLC allows the separation of several compounds and has been nsed to clarify the anthocyanin compositions of different commodities. ... 

In addition to the aforementioned methods, TLC in combination with other instrumental techniques have also been used for quantification of inorganic species. For example, two-dimensional TLC coupled with HPLC has been utilized for the separation and quantification of REEs in nuclear fuel fission products using silaiuzed silica gel as layer material [60]. In another interesting method, REEs in geological samples have been determined by ICP-AAS after their preconcentration by TLC on Fixion plates [32]. TLC in combination with neutron activation has been used to determine REE in rock samples on Eixion 50 x 8 layers with the sensitivity limit of 0.5 to 10 pg/g for 10- to 30-mg samples [41]. A combination of TLC and A AS has been utilized for the isolation and determination of zinc in forensic samples [27]. 

Common pharmaceutical products of olibanum and salai guggul are tablets prepared from dried extracts of boswellic adds, which are obtained by processes involving treatment of the resins with alkali and acid. The stress involved in this treatment is expected to lead to alteration of some triterpenes as, e.g., the conversion of the unstable 3-(9-acetyl-ll-hydroxy-[3-boswellic acid (compound 12) to the stable compound 3-(9-acetyl-9,ll-dehydro-[3-boswellic acid (compound 13). Two-dimensional TLC is an excellent means of observing this conversion [5]. For verification of this process, the substances have to be isolated by PLC and identified by GC-MS. 

For two-dimensional TLC under capillary flow controlled. conditions it should be possible to achieve a spot capacity, in theory, on the order of 100 to 250, but difficult to reach 400 and nearly impossible to exceed 500 [52,140]. Theoretical calculations indicate that by forced-flow development it should be relatively > easy to generate spot capacities well in excess of 500 with an upper bound of several thousand, depending on the choice of operating conditions. -fE... 

The Achilles heel of two-dimensional TLC is the difficulty recording and quantifying chromatograms. 

The enantiomeric purity of protected amino acids used in peptide synthesis can be determined by pre-column partial deprotection followed by derivatization with Marfey s reagent (116). The Marfey diastereoisomers can be easily resolved and determined by RP-HPLC using an ODS-Hypersil column288. Fifteen amino acids collected from mammalian tissues were derivatized with Marfey s reagent and subjected to two-dimensional TLC. Each individual spot (enantiomeric mixture of a diasteroisomer) was then resolved by RP-HPLC. Except for tyrosine (46) and histidine (117), subnanomole quantities of enantiomers could be analyzed289,290. 

A. Pyka and M. Dolowy, Separation of selected bile acids by TLC. II. One-dimensional and two-dimensional TLC. J. Liq. Chrom. Relat. Technol. 27 (2004) 2031-2038. 

The toxins are easily detected, after separation of the compounds from plasma and renal tissue on silica thin layers, by their fluorescence in UV light orellanine is visible as navy blue, orellinine as dark blue, and orelline as light blue (Horn et al., 1997). Beside thin-layer chromatography (TLQ, use of HPLC for the analysis of orellanine, e.g., in mushroom extracts, has also been reported. Quantitative analysis of orellanine in plasma samples, or in (rat) urine samples, was performed by extraction of orellanine on XAD-4 resin, two-dimensional TLC on cellulose, and spectrophotometric evaluation of the orelline produced on the TLC plates after UV-induced decomposition of the orellanine. 

Phosphorus, fatty acids, carbohydrates, glycerol, and amino acids were analyzed by the method described in our previous paper [8] and references cited therein. SDS-PAGE [8], TLC [9], HPLC [9], determination of phos-phomonoester [8], reducing sugar analysis [13], methylation analysis [14], and hexose analysis [15] were performed as described in the respective literature. Two dimensional TLC was performed on silica-gel plate (Merck Silicagel 60 F254 No. 5715) using the solvent systems, chloroform-methanol-acetic acid (65/10/1, v/v/v) for the first development and chloroform-methanol-25% ammonia solution (65/10/1) for the second. 

TLC. Aliquots of samples and standards were run on silica-coated thin layer chromatography (TLC) plates in either n-butanol/acetic acid/water (4 1 1 by vol. (BUOH/HAC/H2O)) or n-propanol/concentrated ammo-nia/water (8 1 11, pre-equilibration). These eluents were previously described (Keller et ah, 1984) for the two-dimensional TLC-separation of elastin cross-links. 

Silufol Five mobile phases, and combination for two dimensional TLC. Dragendorff, iodoplatinate and ninhydrin reagents. In eye lotions and infusion solutions. 109... 

More recently sulfation with H2S04/DCC has been proposed for the preparation of tyrosine 0-[35S]sulfated peptides, e.g. of CCK-8 in nanomolar amounts, since analytical HPLC or two-dimensional TLC can be applied for the separation of the sulfated peptide from the un-sulfated starting material as well as from side products. 84 The low yields of sulfation reported (max 40% even upon optimization with excess DCC and H2S04) most probably derive from the difficulties encountered by working under strictly anhydrous conditions with lyophilized peptide samples. 

Two-dimensional TLC uses the same liquid chromatographic procedure twice to separate spots that are unresolved by only one process. After a sample is run in one solvent, the TLC plate is removed, dried, rotated 90 degrees, and run in another solvent. Any of the spots from the first run that contain mixtures can now be separated. The finished chromatogram is a two-dimensional array of spots. 

Cochromatography is mandatory (for confirmatory purposes) Two-dimensional TLC is mandatory (for confirmatory purposes)... 

Figure 5.3—Principle of two-dimensional TLC. The use of distinctly different mobile phases is recommended for a better separation, a) Deposition of three standards and an unknown b) Migration with the first solvent c) drying and rotation of the TLC plate d) migration with the second solvent. Conclusion . v is probably a, but the impurity is not b or c.

Fig.2.2. Separation of some amino acids by two-dimensional TLC on cellulose MN-300. Solvent systems (1) n-butanol-acetone-diethylamine-water (10 10 2 5) (2) isopropanol-formic acid-water (20 1 5).

Methylcarbamate insecticides have been recently labeled with DNS-C1 [145]. The procedure involves the hydrolysis of the carbamates with 0.1 M sodium carbonate to form a phenol and methylamine [166]. The two hydrolysis products are labeled with DNS-C1 and subsequently detected and determined quantitatively by TLC on silica gel layers by scanning spectrofluorimetry in situ. The reaction conditions were examined, and optimum conditions for hydrolysis and labeling were established [167]. The overall reaction scheme is shown in Fig. 4.62. The phenol derivatives of a number of N-methylcarbamates have been separated by one- and two-dimensional TLC [168], and the fluorescence behaviour and stability of the derivatives have been examined [169]. Most of the DNS derivatives fluoresce at similar wavelengths (excitation, ca. 365 nm emission, ca. 520 nm). The fluorescence spectrum of a typical DNS derivative is shown in Fig. 4.63. The method has been applied successfully to the analysis of low concentrations of carbamates in water and in soil samples with little or no clean-up being required [170,171]. Amounts as low as 1 ng of insecticide can be detected instrumentally. Visual limits of detection are ca. 5-10 ng per spot. 

Raisanen R, Bjork H, Hynninen PH (2000) Two-Dimensional TLC Separation and Mass Spectrometric Identification of Anthraquinones Isolated from the Fungus Dermocybe sanguinea. Z Naturforsch 55c 195... 

---