Among forced-flow layer chromatographic techniques, the centrifugally-driven variety was first developed, and named centrifugal layer chromatography, in 1947 by Hopf. In the 1980s Nyiredy further developed the technique and renamed it rotation planar chromatography. This paper summarizes the classifications and applicability of versions of this technique.
The greater celandine (
L.), a well-known source of isoquinoline alkaloids, has a long history of use as a medicinal plant. Although the antimicrobial activity of
alkaloids against pathogenic bacteria has been reported, the mechanism of this action is almost unknown. The BioArena system, which integrates the modern method and biological results of bioautography with TLC and/or OPLC, is especially suitable for investigating biochemical interactions in the adsorbent layer after chromatographic separation. The antimicrobial effect of alkaloids obtained from
root has been demonstrated by use of this system. It was assumed that the antibiotic activity of
alkaloids was a result of formation of formaldehyde. It was also assumed that addition of endogenous HCHO-capture molecules, for example l
-arginine and glutathione, to the culture medium reduces the antibacterial activity of
alkaloids whereas Cu(II) ions enhance the effect. The results obtained support these assumptions and our earlier observations that HCHO and its reaction products are very important in the antibiotic action of these compounds. These small molecules (l
-arginine and glutathione) can capture HCHO molecules mobilized by alkaloids and possibly by pathogen cells, and may be responsible for reduced antibacterial effect. The HCHO-mobilizing power of Cu(II) ions dramatically enhanced the antibiotic effect. The BioArena system is highly suitable for studying special interactions in the adsorbent layer.
Singlet oxygen (
)-catalyzed oxidation of water leads to a variety of oxidants (e.g. ozone, O
) in host-parasite relationships as well. It can be detected indirectly in TLC or OPLC zones by use of the simple BioArena system and O
-eliminating molecules (e.g.
-limonene and indigo carmine) in the culture medium. It follows from these new findings that not only formaldehyde but also O
and related bioreactive compounds may play a crucial role in the mechanism of antibacterial activity of antibiotic-like compounds. The toxic potential of a molecule, however, originates from the ratio of the oxidants produced in the chromatographic spots.
The antibacterial effect of the components of clary sage (Salvia sclarea L.) and spearmint (Mentha spicata L. var. crispa (Bentham) Danert) was investigated by means of high-performance thin-layer chromatography-direct bioautography against the Gram-positive soil bacterium Bacillus subtilis (Bs) and Gram-negative bacteria such as a pepper pathogen Xanthomonas euvesicatoria (Xe), a luminescence gene-tagged Arabidopsis pathogen Pseudomonas syringae pv. maculicola (Psm) and a luminescent marine Aliivibrio fischeri (Af). Sclareol, linalool, and linalyl acetate were identified as active components of clary sage and carvone as the antibacterial substance in spearmint. Sclareol inhibited all tested bacteria, linalool and carvone showed antibacterial effect against all Gram-negative strains tested, while linalyl acetate only against Xe and Af. Some minor components of the clary sage essential oil also gave a zone of inhibition when tested on Gram-negative bacterium strains.
Direct bioautography performed with luminescence gene-tagged bacteria enables almost real-time detection of antimicrobial compounds in plant extracts. This method for the detection of chamomile (
) components with antibacterial effect against Bacillus subtilis soil bacteria was more sensitive than commonly used bioautographic visualization by staining with a tetrazolium salt. Some compounds had a strong inhibiting effect only via the bioluminescence measurement. Extraction of antibacterial components of chamomile flowers was most effective with 50% ethanol; slightly lower efficiency was achieved with acetone and methanol, and hexane was least effective. The results were confirmed by using luminescent
Pseudomonas syringae pv. maculicola
plant pathogen bacteria.
Overpressured layer chromatography (OPLC), ensuring pumpforced constant mobile phase flow and the possibility of overrun, offers the expanded exploitation of fine-particle adsorbent layers for a longer development distance. Using an infusion—transfusion OPLC method with a 26-cm long development, the separation of clove, rosemary, eucalyptus, tea tree, spearmint, thyme, and cinnamon bark essential oil components was achieved with good resolutions. In the combination of OPLC and Aliivibrio fischeri assay, the main essential oil components eugenol, borneol, (−)-R-carvone, thymol, and trans-cinnamaldehyde exhibited antibacterial effect. The OPLC—2,2-diphenyl-1-picrylhydrazyl (DPPH*) test showed two antioxidant components: eugenol and thymol.
In this study, the antibacterial profiling of the ethanolic leaf extract of greater burdock (Arctium lappa L.) is demonstrated, applying thin-layer chromatography (TLC) coupled bioassays against the Gram-positive soil bacterium Bacillus subtilis and the Gram-negative pepper pathogen Pseudomonas syringae pv. maculicola. The main active component was isolated by eluting from the adsorbent bed and subjected to a targeted characterization by high-performance liquid chromatography–diode array detection–electrospray ionisation–mass spectrometry. The identification of the germacranolide sesquiterpene lactone onopordopicrin was based on its retardation factor, bioactivity in TLC-based methods, and retention tim as well as ultraviolet (UV) and mass spectra, compared to those of the reference substance isolated earlier in our laboratory from Onopordum acanthium leaf.
A group of thirteen newly synthesized potential herbicides,
-aryltrichloroacetamides or 2-(chlorophenoxy)acyl derivatives, have been initially investigated by reversed phase (RP) TLC. The lipophilicity of the substances was described by retention factors in water, log
, calculated from experimental RP TLC data, and by log
values calculated by use of software. Biological activity was examined by use of the BioArena system of TLC separation then (micro)biological detection. The potential role of formaldehyde (HCHO) in the toxic antibacterial action of substances against
bacterial cells was investigated. The effect of HCHO capturers (L
-arginine and reduced glutathione) and Cu
ions on the bioactivity and mechanism of toxicity of the substances was examined. It was established that HCHO and its potential reaction products (e.g.
) are crucial in the mechanism of action of these molecules. Correlations between the lipophilicity and bioactivity of the test compounds were also analyzed. It seems that hydrophobicity is not the decisive factor affecting the bioactivity of these substances.