Mladen Miloš

Chemical composition and antioxidant effect of glycosidically bound volatile compounds from oregano (Origanum vulgare L. ssp. hirtum).

Abstract The present work examines the content and chemical composition of the glycosidically bound volatiles from oregano as well as their antioxidative properties. The glycosidically bound volatiles amounted to 20 mg kg−1 in dried leaves and flowers of oregano. Fourteen volatile aglycones were identified with thymoquinone as the major component. Other important aglycones were benzyl alcohol, eugenol, 2-phenyl-ethanol, thymol, 3-hexen-1-ol and carvacrol. It was found that all of the aglycones have an antioxidant effect when tested by measuring peroxide values of lard stored at 60°C. These results were compared to the antioxidative activity of oregano essential oil, pure thymol, thymoquinone and also to α-tocoferol which is well known among natural antioxidant compounds.

Chemical Composition and In Vitro Evaluation of Antioxidant Effect of Free Volatile Compounds From Satureja montana L

As a part of an investigation of natural antioxidants from Dalmatian aromatic plants, in this paper we report a study of the antioxidant activity related to the chemical composition of savory free volatile compounds. Twenty-one compounds were identified in the essential oil without fractionation, representing 97.4% of the total oil. The major compound was phenolic monoterpene thymol (45.2%). Other important compounds were monoterpenic hydrocarbons p-cymene (6.4%) and γ-terpinene (5.9%) and oxygen-containing compounds carvacrol methyl ether (5.8%), thymol methyl ether (5.1%), carvacrol (5.3%), geraniol (5.0%) and borneol (3.9%). The evaluation of antioxidant power was performed in vitro by the β-carotene bleaching and thiobarbituric acid (TBA) methods. As determined with both methods, the total savory essential oil as well as different fractions or pure constituents containing hydroxyl group exhibited relatively strong antioxidant effect. The hydrocarbons, when isolated as CH fraction, showed the poorest effectiveness in spite the fact that this fraction contained γ-terpinene, α-terpinene, p-cymene and terpinolene which previously were identified as potential antioxidants.

The effects of essential oils and aqueous tea infusions of oregano (Origanum vulgare L. spp. hirtum), thyme (Thymus vulgaris L.) and wild thyme (Thymus serpyllum L.) on the copper-induced oxidation of human low-density lipoproteins

In this study, the antioxidative capacity effect of essential oils and aqueous tea infusions obtained from oregano, thyme and wild thyme on the oxidation susceptibility of low-density lipoproteins (LDL) has been studied. The results indicate a dose-dependent protective effect of the tested essential oils and aqueous tea infusions on the copper-induced LDL oxidation. The protective effect of essential oils is assigned to the presence of phenolic monoterpenes, thymol and carvacrol, which are identified as the dominant compounds in these essential oils. The strong protective effect of aqueous tea infusions is proposed to be the consequence of large amounts of polyphenols, namely rosmarinic acid and flavonoids (quercetin, eriocitrin, luteolin-7-O-glucoside, apigenin-7-O-glucoside, luteolin, apigenin), with the most pronounced effect in the case of oregano. These findings may have implications for the effect of these compounds on LDL in vivo.

Gas chromatography mass spectral analysis of free and glycosidically bound volatile compounds from Juniperus oxycedrus L. growing wild in Croatia.

The essential oils in fresh needles and green and mature berries of Juniperus oxycedrus L. (Cupressaceae) were analyzed by GC–MS. A total of 36 compounds were identified from needles, representing 94.91% of the total oil. 15 compounds were identified in the green berry oil and 22 in the mature berry oil, which accounted for 94.33 and 90.94% of the total oil composition. The major component was α-pinene. The glycosidically bound volatile compounds amounted to 21 mg kg−1 in needles and 4 mg kg−1 from green berries. Only traces of aglycones were identified in mature berries. Sixteen volatile aglycones were identified in needle sample with 2-hydroxy-5-methylacetophenone as major component. A total of nine aglycones were identified in green berries. The major aglycones were 3-phenyl-2-propen-1-ol and myrtenol. There was no similarity between the glycosidically bound aglycones and the corresponding free compounds found in the essential oil.

Molecular and chemical characterization of the most widespread Ocimum species

DNA fingerprinting (AFLP) and chemical analyses of essential oils were utilized to define the extent of variation existing in the genus Ocimum. Research was carried out on 22 Ocimum accessions representing seven species. Concerning the essential oil composition of all investigated accessions, 115 compounds were identified. UPGMA cluster analysis, based on Euclidian distances of essential oil constituents between all pairs of accessions, showed four well-supported clusters (O. tenuiflorum, O. basilicum/O. africanum, O. basilicum, and O. americanum/O. africanum). Relating to the essential oil composition of all of the investigated accessions, 17 compounds were identified as the main ones, and according to them 13 chemotypes were determined. AFLP relationships were determined by neighbor-joining (NJ) cluster analysis based on Dice’s distance matrix and by maximum parsimony (MP) analysis. O. basilicum, O. americanum/O. africanum, O. tenuiflorum, and O. gratissimum represented four clusters supported with high bootstrap values. A neighbor-net diagram allowed the visualization of apparently conflicting data by revealing relationships between genotypes and chemotypes. Concerning the O. africanum species, two distinct chemotypes, geranial/neral (accession 11) and estragol (accession 10), have been established, while all the studied O. americanum accessions belong to the geranial/neral chemotype. This could be additional evidence that O. americanum is one of the parents of O. africanum. Furthermore, the fact that the O. africanum accession (10) as well as O. basilicum ‘Purpurascens’ and O. basilicum ‘Erevanskii’ accessions belong to the estragol chemotype supports the theory that O. africanum is one of the parents of these two O. basilicum accessions.

The impact of the locality altitudes and stages of development on the volatile constituents of Salvia officinalis L. from Bosnia and Herzegovina

Abstract The essential oils from fresh plant material of Salvia officinalis L. (sage) were subjected to GC/MS analysis in order to determine the impact of the locality altitudes and seasonal variations on their volatile constituents. The sage plant material was collected from two different localities (altitudes 110 and 400 m) in central Herzegovina near Mostar and at four different stages of development: vegetative period (leaves and stalks, January 2003), prior to flowering (leaves and stalks, April 2003), in the course of flowering (flowering tops, leaves and stalks, May 2003) and after flowering (leaves and stalks, August 2003). The oil yields varied from 0.29% to 1.07%. The qualitative composition of the components appeared to be constant. However, there were notable differences in the amounts of several compounds depending on the stages of plant development. The main components were α-thujone (9.3–35.6%), camphor (6.9–29.1%) and viridiflorol (6.0–24.0%). Other important components were α-humulene (3.1–13.6%), manool (3.0–13.3%), 1,8-cineole (8.6–12.7%) and borneol (2.0–5.5%).

Chemical characterization and genetic relationships among Ocimum basilicum L. cultivars.

Twenty‐seven Ocimum basilicum cultivars were subjected to a chemical characterization of essential oil components by gas chromatography/mass spectrometry (GC/MS) and a genetic characterization using the amplified fragment‐length polymorphism (AFLP) technique. Since the same 27 accessions had previously been classified into six morphotypes, these analyses allowed us to make detailed comparisons of chemistry, genetics, and morphology. The chemical composition and morphology of the studied cultivars appeared to have a strong genetic component. The AFLP analysis revealed a distinction between the green and purple morphotypes. The green morphotypes predominantly utilized the terpene biosynthetic pathway, while most purple morphotypes primarily utilized the phenylpropene biosynthetic pathway. The GC/MS analysis led to identification of 87 volatiles. Among the 27 cultivars, five chemotypes were identified. A detailed characterization of the essential oil constituents indicated the existence of both specific combinations of compounds and ‘private’ compounds with the potential to be used in many aspects of human life. The established relationship between a genetic profile, chemical composition, and morphology represents an important step in future breeding programs and in the cultivation of this species.

Dipotassium-trioxohydroxytetrafluorotriborate, K2[B3O3F4OH], is a potent inhibitor of human carbonic anhydrases

Abstract The boron heterocyclic compound dipotassium-trioxohydroxytetrafluorotriborate (K2[B3O3F4OH]) was investigated as inhibitor of the zinc enzyme, carbonic anhydrase (CA, EC 4.2.1.1). Eleven human (h) CA isoforms, hCA I–IV, VA, VI, VII, IX and XII–XIV, were included in the investigations. The anion, similar to tetraborate or phenylboronic acid, inhibited most of them. hCA III was not inhibited by K2[B3O3F4OH], whereas hCA VA, hCA VI, hCA IX and hCA XIII were inhibited in the submillimolar range, with KIs of 0.31–0.63 mM. hCA I and II (cytosolic, widespread isoforms), hCA IV (membrane-bound isoform), hCA XII (tumor-associated, transmembrane) and hCA XIV (transmembrane) were much more effectively inhibited by this anion, with inhibition constants ranging from 25 to 93 µM. hCA VII, a cytosolic enzyme present in the brain and associated to oxidative stress, was very effectively inhibited by K2[B3O3F4OH], with a KI of 8.0 µM. We propose that K2[B3O3F4OH] binds to the metal ion from the enzyme active site, coordinating to the Zn(II) ion monodentately through its B-OH functionality. We hypothesize that some of the beneficial antitumor effects reported for K2[B3O3F4OH] may be due to the inhibition of CAs present in skin tumors.

The effect of air-drying on glycosidically bound volatiles from seasonally collected origano (Origanum vulgare ssp. hirtum) from Croatia

The glycosidically bound volatiles were isolated from fresh and airdried origano by exhaustive percolation with ethyl acetate. After the purification of glycosidic fraction, the enzymatic hydrolysis with beta-glucosidase was performed. The obtained aglycones were analysed by GC-MS. Eighteen compounds were identified. The seasonal variations of main aglycone contents from the fresh plant material were: thymoquinone (3.10-6.18 mg/kg), benzyl alcohol (1.33-3.62 mg/kg), 3,5,5-trimethyl-4-(3-hydroxy-1-buthenyl)-2-cyclohexen-1-on (0.51-3.35 mg/kg), 2-phenyl ethanol (0.42-2.98 mg/kg), eugenol (0.93-2.55 mg/kg), thymol (0.70-1.40 mg/kg) and carvacrol (0.88-1.40 mg/kg). The season of collecting influenced the content and composition of the glycosidically bound volatiles of origano. Air-drying effected mostly the yield of these compounds.

Essential oil and glycosidically bound volatiles of Origanum vulgare L. ssp. hirtum (Link) Ietswaart

The glycosidically bound volatiles were isolated from dried plant material by percolation with ethyl acetate and by extraction with water during hydrodistillation of essential oil. Fifteen volatile aglycones were identified by gas chromatography-mass spectrometry (GC-MS) on two columns with different polarity of the stationary phases. The main aglycones were: thymoquinone, benzyl alcohol, thymol, 2-phenylethanol, carvacrol and 1-octen-3-ol. The content of aglycones was 19 and 21 mg kg−1 with respect to the method of isolation. The chemical composition of aglycones was compared with the chemical composition of essential oil, which consisted mainly of thymol and carvacrol. After the hydrolysis of glycosides, D-(+)-glucose and unknown disaccharide were identified. The hydrolysis of disaccharide, D-(+)-glucose and D-(+)-galactose were detected by TLC and GC-MS as trimethylsilylethers. Copyright