Impact of D2O on the peptidization of l‑methionine

This is our follow-up study carried out in an order to collect experimental evidence regarding the impact of heavy water ( D2O) on the spontaneous oscillatory peptidization of the sulfur-containing proteinogenic α-amino acids and this time our target is l-methionine (l-Met). Our earlier study has been focused on l-cysteine (l-Cys) and it seemed interesting for us to compare the effect induced by D2O on these two structurally related compounds. In a broader sense though, our goal is to investigate if spontaneous oscillatory peptidization of various different proteinogenic α-amino acids might play any role in living organisms. As analytical techniques, we used high-performance liquid chromatography with the evaporative light-scattering detection (HPLC-ELSD), mass spectrometry (MS), scanning electron microscopy (SEM), and turbidimetry. The obtained results demonstrate that heavy water seriously hampers the oscillatory peptidization of l-Met, apparently due to the presence of the deuterium cation in the reaction medium and in the first instance, due to isotopic substitution of two functionalities participating in peptidization, i.e., –COOH and –NH2. This effect is fully analogous to that earlier reported for l-Cys. A cautious conclusion can be drawn that far more experimental evidence is still needed to be able to judge about an importance (or otherwise) of the oscillatory peptidization of proteinogenic α-amino acids for various different life processes

Impact of D2O on peptidization of L‑proline

This is our follow-up study carried out in an order to collect experimental evidence regarding the impact of heavy water (D2O) on the spontaneous oscillatory peptidization of l-proline (l-Pro). Our earlier studies have been focused on the two sulfurcontaining proteinogenic α-amino acids, i.e., l-cysteine (l-Cys) and l-methionine (l-Met), and it seemed interesting to assess the effect induced by D2O on one more proteinogenic α-amino acid, i.e., l-Pro. It needs to be added that unlike l-Met, but similar to l-Cys, the oscillatory peptidization of l-Pro dissolved in the organic-aqueous solvent characterizes with the circadian rhythm. As analytical techniques, we used high-performance liquid chromatography with the evaporative light-scattering detection (HPLC-ELSD), mass spectrometry (MS), scanning electron microscopy (SEM), and turbidimetry. The obtained results can in certain sense be viewed as analogous to those earlier reported for l-Cys and l-Met by demonstrating that heavy water considerably hampers the oscillatory peptidization of l-Pro. However, an unexpected observation was also made than unlike the cases with l-Cys and l-Met, the observed hampering effect of D2O on the oscillatory peptidization of l-Pro is not monotonously dependent on the concentration of D2O in the system, but it is the strongest pronounced for 10% (v/v) D2O in the employed binary methanol–water solvent (with the investigated proportions of D2O in this solvent changing from 0 to 30%). Although we have no rational explanation for this striking effect, we believe that it should not pass unnoticed and therefore it is emphasized in this study. Maybe this firm quantitative result will prove an inspiration for future researchers interested in getting a deeper insight into the role of D2O in life processes, and more specifically in the kinetic and the mechanistic aspects thereof

Impact of D2O on peptidization of L-Cysteine

An attempt was made to answer the question if spontaneous oscillatory conversion and peptidization of proteinogenic a-amino acids might be essential for living organisms. To this effect, we investigated an impact of heavy water (D2O) on the peptidization of L-Cys. As analytical techniques, we used high-performance liquid chromatography, mass spectrometry, scanning electron microscopy, and turbidimetry. The results obtained demonstrate that heavy water seriously hampers the oscillatory peptidization of L-Cys, apparently due to the presence of the deuterium cation in the reaction medium. A cautious conclusion can be drawn that thorough reflection is needed on possible importance of the oscillatory peptidization of proteinogenic a-amino acids for various different life processes

Impact of D2O on the peptidization of L‑alanine

This is our fifth consecutive study carried out in an order to collect experimental evidence on the impact of heavy water ( D2O) on the spontaneous peptidization of proteinogenic α-amino acids and this time its subject matter is L-alanine (L-Ala). Our four earlier studies have been focused on the two sulfur-containing α-amino acids (i.e., L-cysteine (L-Cys) and L-methionine (L-Met)), and on two structurally related α-amino acids (i.e., L-proline (L-Pro) and L-hydroxyproline (L-Hyp)). It seemed interesting to assess the effect exerted by D2O on L-Ala, the simplest chiral (endogenous and proteinogenic) α-amino acid with as low molar weight, as 89.09 g mol−1 only. As analytical techniques, we used high-performance liquid chromatography with the diode array detection (HPLC–DAD), mass spectrometry (MS), and scanning electron microscopy (SEM). The obtained results make it clear that the impact of heavy water on the dynamics of the spontaneous peptidization of L-Ala is even stronger than with the four other α-amino acids discussed earlier (although in all five cases, heavy water significantly hampers spontaneous oscillatory peptidization). Unlike in the four previous cases, though, the solubility of L-Ala in pure D2O is quite low and it takes twice as much time to dissolve it in D2O than in MeOH + X, 70:30 (v/v). Consequently, the peptidization of L-Ala in heavy water is even more obstructed than it was the case with the other investigated α-amino acids and it results in considerable yields of the L-Ala crystals (most probably at least partially deuterated) at the expense of the L-Ala-derived peptides. Perhaps it might be interesting to add that out of five α-amino acids investigated so far, which can be divided into two groups of endogenous and exogenous species, two endogenous species (L-Cys and L-Pro) undergo spontaneous oscillatory peptidization in an aqueousorganic solvent (i.e., in the absence of D2O) following the circadian rhythm, whereas two exogenous ones (i.e., L-Met and L-Hyp) do not. The third endogenous species (L-Ala) first undergoes two initials oscillations which are damped (not periodic) and the oscillatory changes are on a scale of ca. 10 h (as estimated with use of the Fourier transform approach) and after that, the system reaches a steady state

Thin-layer chromatographic quantification of magnolol and honokiol in dietary supplements and selected biological properties of these preparations

Two isomeric biphenyl neolignans, magnolol and honokiol, are considered as constituents responsible for the healing effect of magnolia bark, a traditional Oriental medicine. To survey the increasing number of dietary supplements that contain magnolia bark or its extract, an affordable quantitative thin-layer chromatography (TLC) –densitometry method was developed. The methanol extracts were analyzed on the silica gel plates after manual sample application using n -hexane –ethyl acetate –ethanol (16:3:1, v/v/v) as a mobile phase. For quantitation, the chromatograms were scanned in the absorbance mode at the wavelength λ= 290 nm. The limits of detection and quantitation were 90 and 280 ng/zone for magnolol and 70 and 200 ng/zone for honokiol, respectively. None of the two targeted neolignans were detected in two of the six analyzed supplements. In the other four samples, the measured amounts were between 0.95-114.69 mg g −1 for magnolol and 4.88-84.86 mg g −1 for honokiol. Moreover, separations of these two neolignans on the TLC and high-performance TLC (HPTLC) layers were compared and HPTLC was combined with antioxidant (DPPH) and antibacterial ( Bacillus subtilis and Aliivibrio fischeri ) assays and mass spectrometry (MS), using the elution-based interface. Both magnolol and honokiol exhibited effects in all bioactivity assays. The HPTLC-MS tests confirmed purity of neolignan zones in the extracts of dietary supplements and supported tentative identification of the alkaloid piperine and the isoflavone daidzein as additional bioactive components of the investigated dietary supplements. Using the same mobile phase in the orthogonal directions 2D-HPTLC-MS experiments proved degradation, i.e., instability of magnolol and honokiol on the silica gel adsorbent

The Hampering Effect of Heavy Water (D2O) on Oscillatory Peptidization of Selected Proteinogenic α-Amino Acids

We present an overview of our studies on the hampering effect of heavy water (D2O) on spontaneous oscillatory peptidization of selected proteinogenic a-amino acids. The investigated set of compounds included three endogenous and two exogenous species. The experiments were carried out with use of high-performance liquid chromatography (HPLC), mass spectrometry (MS) and scanning electron microscopy (SEM). These techniques were chosen to demonstrate spontaneous oscillatory peptidization of a-amino acids in an absence of D2O (HPLC) and the hampering effect of D2O on peptidization (HPLC, MS and SEM). The HPLC analyses were carried out at 21 ± 0.5◦C with each a-amino acid freshly dissolved in the binary liquid mixture of organic solvent + H2O, 70:30 (v/v) or in pure D2O for several dozen hours or several hours, respectively. The analyses with use of MS and SEM were carried out, respectively, after 7 days and 1 month of sample storage period in the darkness at 21±0.5◦C and for these experiments, each a-amino acid was dissolved in the liquid mixture of organic solvent + X, 70:30 (v/v), where X: H2O + D2O in volume proportions from 30:0 to 0:30. The results obtained with use of HPLC, MS and SEM point out to the strong hampering effect of D2O on the oscillations and peptidization yields, yet the dynamics of these processes significantly depends on chemical structure of a given a-amino acid

Impact of D2O on peptidization of L-histidine

This is our sixth consecutive study carried out in an order to collect an experimental evidence on the impact of heavy water (D2O) on spontaneous peptidization of the proteinogenic α-amino acids and this time it is L-histidine (L-His). Scientists have not yet achieved a full consensus regarding the source of this very important amino acid in human and mammalian tissues, and on this particular question rather contradictory answers in form of experimental results are produced, equally supporting its exogenous and endogenous origin. Although this issue still remains unsolved, for practical demands of life sciences the two UN agencies, FAO and WHO, have both tentatively accepted that L-His is an exogenous α-amino acid. As analytical techniques, in our studies we employed high-performance liquid chromatography with the diode array detection (HPLC–DAD), mass spectrometry (MS), and scanning electron microscopy (SEM). Spontaneous peptidization of L-His dissolved in methanol + H2O,7:3 (v/v) was carried out at 22 ± 0.5 °C in the darkness for a relatively long period of 314 h, and its progress was chromatographically checked by targeting concentration of the L-His monomer in the 12-min intervals. This investigation revealed alternating yet non-periodic concentration changes, indicating changeable formation and hydrolytic decay of the L-His-derived oligopeptides in the function of time, and a fast net concentration fall of the L-His monomer (witnessing to quite vigorous peptidization). Moreover, the MS results confirmed formation of the relatively high oligopeptides, falling within the range of two or more dozen L-His monomer units. Impact of D2O on peptidization of L-His was traced with use of MS and SEM for the L-His samples dissolved in aqueous methanol solvents containing 5,10, 20, and 30% D2O, and also in pure D2O. Similar to the results earlier presented for five other proteinogenic α-amino acids, heavy water exerts a powerful inhibitory effect on spontaneous peptidization of L-His, equally perceptible when assessed with aid of mass spectrometry (with the mass spectra in the first instance playing the role of quasi-quantitative fingerprints), and based on purely qualitative micrographs derived with use of SEM

The 42nd Symposium Chromatographic Methods of Investigating Organic Compounds : Book of abstracts

The 42nd Symposium Chromatographic Methods of Investigating Organic Compounds : Book of abstracts. June 4-7, 2019, Szczyrk, Polan

Zastosowanie chromatografii cienkowarstwowej do analizy wybranych pigmentów roślinnych

The doctoral dissertation is divided into two parts: theoretical and experimental. In the theoretical part, primary and secondary plant metabolites are discussed and individual groups of phytochemical compounds are characterized. The following chapters are devoted to the compounds from the group of anthocyanins, betacyanins and neolignans. The experimental part includes descriptions of the research and analyses carried out using thin-layer chromatography (TLC and HPTLC methods), scanning densitometry, UV-Vis spectrophotometry and mass spectrometry (TLC-MS). The aim of the study was to develop new conditions for the chromatographic separation of compounds, as well as to apply the method for their qualitative and quantitative determination. The first stage is focused on the analysis of selected anthocyanins: cyanin, keracyanin, delphinidin and pelargonidin. In addition, a method of triple chromatographic development for the separation of anthocyanins was carried out. The tested compounds were determined in food samples. The next stage of research focuses on betacyanins, including betanin. Horizontal separation of betacyanin pigments was observed during the chromatographic analysis, which is a "fingerprint" of those compounds. The last part of the experimental concerns compounds from the group of neolignans: magnolol and honokiol, which were determined in samples of dietary supplements, and their antioxidant and antibacterial properties were examined