Biosynthesis of essential oils in aromatic plants: A review

In aromatic plants species, biosynthesis of essential oils occurs through two complex natural biochemical pathways involving different enzymatic reactions. Isopentenyl diphosphate (IPP) and its isomer dimethylallyl diphosphate (DMAPP) are the universal precursors of essential oil biosynthesis and are produced by the cytosolic enzymatic MVA (mevalonic acid) pathway or by plastidic and enzymatic 1-deoxy-d-xylolose-5-phosphate (DXP) pathway, also called the 2-C-methylerythritol-4-phosphate (MEP) pathway. In the particular plant cell part, prenyl diphosphate synthases condense isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) further to form prenyl diphosphates, which are used as substrates for geranyl diphosphate (GPP; C₁₀) or for fernesyl diphosphate (FPP; C₁₅). Essential oils are final terpenoid products and are formed by a huge group of enzymes known as terpene synthases (TPS). Essential oils are important secondary metabolites of plants and have been used not only in different industries but also in ethnobotanical medicines for centuries. Hence, considerable research has been undertaken to understand the essential oil biosynthetic pathways. This review will be a valuable source of information in the field of natural products, as we give detailed insights about biosynthesis of essential oils in plants and thus indicate also new unexplored horizons for further research.     

Supercritical CO2 Extraction of β-Carotene from Sweet Potatoes

Using supercritical CO₂ to extract β-carotene there was approximately a five-fold or a three-fold increase in amount of carotenoids extracted from freeze-dried tissue relative to the amount extracted from oven-dried or fresh tissue, respectively. The most efficient conditions were at 48°C and 41.4 MPa. Of the total carotenoid content ～20% was inaccessible to supercritical CO₂. The HPLC carotenoid profile of sweet potatoes showed that the unextracted tissue contained 90%β-carotene, primarily as all-trans (ca. 99%). Supercritical CO₂ extracts contained up to 94%β-carotene. The isomer composition of β-carotene of supercritical extracts showed ～ 14% 13-cis and 11% 9-cis.     

In vitro synthesis of high molecular weight rubber by Hevea small rubber particles

Hevea brasiliensis is one of few higher plants producing the commercial natural rubber used in many significant applications. The biosynthesis of high molecular weight rubber molecules by the higher plants has not been clarified yet. Here, the in vitro rubber biosynthesis was performed by using enzymatically active small rubber particles (SRP) from Hevea. The mechanism of the in vitro rubber synthesis was investigated by the molecular weight distribution (MWD). The highly purified SRP prepared by gel filtration and centrifugation in the presence of Triton^® X-100 showed the low isopentenyl diphosphate (IPP) incorporation for the chain extension mechanism of pre-existing rubber. The MWD of in vitro rubber elongated from the pre-existing rubber chains in SRP was analyzed for the first time in the case of H. brasiliensis by incubating without the addition of any initiator. The rubber transferase activity of 70% incorporation of the added IPP (w/w) was obtained when farnesyl diphosphate was present as the allylic diphosphate initiator. The in vitro synthesized rubber showed a typical bimodal MWD of high and low molecular weight fractions in GPC analysis, which was similar to that of the in vivo rubber with peaks at around 10⁶ and 10⁵ Da or lower. The reaction time independence and dependence of molecular weight of high and low molecular weight fractions, respectively, indicated that the high molecular weight rubber was synthesized from the chain extension of pre-existing rubber molecules whereas the lower one was from the chain elongation of rubber molecules newly synthesized from the added allylic substrates.     

Effect of water activity and temperature on growth of Alternaria alternata on a synthetic tomato medium

Alternaria alternata is a toxigenic fungus, predominantly responsible for Blackmould of ripe tomato fruits, a disease frequently causing substantial losses of tomatoes, especially those used for canning. The objective of this study was to determine the effect of water activity (a_w, 0.904, 0.922, 0.954, 0.982) and temperature (6, 15, 21 and 35 °C) on germination and radial growth rate on a synthetic tomato medium of a cocktail inoculum of five strains of A. alternata isolated from tomato fruits affected by Blackmould. The shortest germination time (1.5 days) was observed at 0.982 a_w, both at 21 °C and 35 °C. The germination time increased with a reduction on a_w. The fastest growth rate was registered at 0.982 a_w and 21 °C (8.31 mm/day). Growth rates were higher when a_w increased. No growth or germination was observed at the lowest a_w level evaluated (0.904) after 100 days of incubation at 6 °C and 15 °C. A temperature of 6 °C caused a significant reduction in growth rates, even at the optimum a_w level. The knowledge on the ecophysiology of the fungus in this substrate is necessary to elaborate future strategies to prevent its development and evaluate the consumer health risk.     

Study of the changes of trans-resveratrol caused by ultraviolet light and determination of trans- and cis-resveratrol in Spanish white wines

Trans-resveratrol is a polyphenolic compound found in several foods items like wines, legumes, berries, peanuts and pistachios and lately it has attracted great attention due to its beneficial health effects such as anti-cancer, neuroprotective, anti-aging and anti-inflammatory. Therefore, it is important to identify all isomers and derivatives of this compound in order to avoid any errors in their quantification. This paper focus on the study of the transformation of trans-resveratrol into the cis form after subjecting the trans form to UV light. Several conditions were tested and results show the transformation should be made in perfectly controlled conditions (UV light wavelength; distance between the UV lamp and the solution and exposure surface) otherwise the formation of a third compound is observed whose concentration increases as does the light exposure time. This new compound is derived from cis-resveratrol and it appears in a higher concentration when the initial concentration of the trans-resveratrol solution is low. Trans- and cis-resveratrol were determined in white wines from the northwestern of Spain.     

HPLC/MS and HPLC/NMR as hyphenated techniques for accelerated characterization of the main constituents in Chamomile (<Emphasis Type="Italic">Chamomilla recutita</Emphasis> [L.] Rauschert)

An ethanolic extract of Chamomilla recutita [L.] Rauschert was analyzed by the hyphenated techniques HPLC/MS and HPLC/NMR, speeding up a reliable and comprehensive characterization of the main constituents in an extract from chamomile flowers. In addition to the cis- and trans-isomers of 2-(glucosyloxy)-4-methoxy-cinnamic acid and also herniarin and apigenin, apigenin-7-O-β-glucoside itself and further derivatives of the latter could unambiguously be identified as main constituents.     

ORGANOLEPTIC TRIALS WITH HOP BITTER SUBSTANCES

Pure hops-derived substances were added to unhopped beer. Illumination only produced a sun-struck flavour with cis-isohumulone, trans-isohumulone and racemic trans-isohumulone. It seems that 3-methyl-butenyl-mercaptan is not or not alone responsible for sun-struck flavour. Tests trials show that trans-isohumulone, cis-isohumulone, racemic trans-isohumulone, trans-₁-isohumulone and cis-₁-isohumulone do not show significant differences in bitter taste and bittering power. Fresh solutions of these bitter acids are not as bitter as solutions which have been kept for a few weeks.     

Biotransformation of (R)-(+)- and (S)-(−)-limonene to α-terpineol by Penicillium digitatum— investigation of the culture conditions

The biotransformation of (R)-(+)- and (S)-(−)-limonene by Penicillium digitatum was investigated. One strain of P. digitatum was able to convert (R)-(+)-limonene to pure (R)-(+)-α-terpineol in 8 h with a yield of up to 93%. It was found that (R)-(+)-limonene was converted much better into α-terpineol than (S)-(−)-limonene, and that no significant chemical conversion of the substrate occurred in control flasks at pH 3.5. The culture conditions involved such as the type and concentration of co-solvent applied and the sequential addition of substrate were investigated, taking into account some findings on the physical behaviour of the system. The highest bioconversion yields were obtained when the substrate was applied as a diluted solution in EtOH.     

Impact of packaging and storage conditions on colour and β-carotene retention of pasteurised mango purée

The effects of light and oxygen on β-carotene, vitamin A value, and colour stability of mango purée during storage were evaluated. Non-aseptic production at a pilot-plant scale, involving continuous high-temperature-short-time (HTST) heating, caused significant 13-cis-β-carotene formation, resulting in a pro-vitamin A retention of 82.3–89.6%, while peroxidase (POD) activity was reduced to residual levels of 0.4–2.6% in the pasteurised purée. Since oxygen removal from the package headspace by flushing with superheated steam or N₂ marginally affected β-carotene stability during storage, hot-filling was most crucial for exhausting the air. Irrespective of the packaging material, pro-vitamin A loss during storage was chiefly caused by oxidative degradation of β-carotene. Additional photo- isomerisation under exposure to light became manifest in the relative increase in 9-cis-β-carotene with associated decline in the 13-cis isomer portion. Despite photoisomerisation, pro-vitamin A retention was up to 90.9% in bottled purée after 168 days of light-exposed storage, with deaeration of the purée resulting from hot-filling. Marked colour loss mainly caused by accumulation of brown oxidation products was monitored by a rising degree of discolouration (Dd). Although additional headspace deaeration of cans and bottles improved colour retention, dissolved residual oxygen and other reactive compounds provoked browning and β-carotene degradation. As regards pro-vitamin A and colour, headspace minimisation was much more effective than its deaeration. Due to smaller headspace volumes, bottling proved superior to canning under the prevailing conditions, irrespective of light-exposed storage.     

Comparing predicting models for heat inactivation of Listeria monocytogenes and Pseudomonas aeruginosa at different pH

Under the same experimental conditions it has been demonstrated that whereas survival curves of Listeria monocytogenes in the range of temperatures from 54 to 62 °C followed a first-order kinetic, those of Pseudomonas aeruginosa in the range of temperatures from 50 to 56 °C were not linear showing a shoulder followed by a linear region. The first order kinetic model did not describe survival curves of P. aeruginosa. A model based on the Weibull distribution (Log₁₀(N_t/N₀)=(1/−2.303)*(t/b)ⁿ)) accurately described the inactivation kinetics of both microorganisms at the three pHs of 4, 5.5, 7.4 investigated. For both microorganisms, the b value depended on the treatment temperature and the pH of the treatment medium. Whereas for L. monocytogenes the n value was independent of the treatment conditions, for P. aeruginosa the n value depended on the pH of the treatment medium.The model based on the Weibull distribution was capable of accurately predicting the treatment time to inactivate five Log₁₀ cycles of both microorganisms at the three pHs investigated.     

Biochemical and thermal characterization of crude exo-polygalacturonase produced by Aspergillus sojae

Crude exo-polygalacturonase enzyme (produced by Aspergillus sojae), significant for industrial processes, was characterized with respect to its biochemical and thermal properties. The optimum pH and temperature for maximum crude exo-polygalacturonase activity were pH 5 and 55 °C, respectively. It retained 60–70% of its activity over a broad pH range and 80% of its initial activity at 65 °C for 1 h. The thermal stability study indicated an inactivation energy of E_d = 152 kJ mol⁻¹. The half lives at 75 and 85 °C were estimated as 3.6 and 1.02 h, respectively. Thermodynamic parameters, ΔH^*, ΔS^* and ΔG^*, were determined as a function of temperature. The kinetic constants K_m and V_max, using polygalacturonic acid as substrate, were determined as 0.424 g l⁻¹ and 80 μmol min⁻¹, respectively. SDS-PAGE profiling revealed three major bands with molecular weights of 36, 53 and 68 kDa. This enzyme can be considered as a potential candidate in various applications of waste treatment, in food, paper and textile industries.     

Partial characterization of peroxidase from the leaves of thymbra plant (Thymbra spicata L. var. spicata)

Peroxidase (EC 1.11.1.7; donor: hydrogen-peroxide oxidoreductase) catalyses the oxidation of various electron donor substrates (e.g. phenols, aromatic amines). In this study, the peroxidase was extracted from Thymbra spicata L. var. spicata and, then partially purified with (NH₄)₂SO₄ precipitation and dialysis. The substrate specificity of peroxidase was investigated using 2,2′-azino-bis(3-ethylbenz-thiazoline-6-sulfonic acid) (ABTS), o-dianisidine, o-phenylenediamine, catechol and guaiacol as substrates. Furthermore, the effects of buffer concentration, pH, temperature and thermal inactivation on enzyme activity were also studied. The results obtained have shown that (i) the best substrate is o-dianisidine, followed by ABTS, catechol, guaiacol and o-phenylenediamine, respectively; (ii) the best buffer concentration is 40 mM for o-dianisidine and catechol, 10 mM for ABTS and guaiacol, and 100 mM for o-phenylenediamine; (iii) optimum pH is 2.5 for ABTS and o-phenylenediamine, 6.0 for o-dianisidine, and 7.0 for catechol and guaiacol; (iv) optimum temperature is 20 °C for catechol, 40 °C for ABTS, guaiacol and o-dianisidine, and 50 °C for o-phenylenediamine; and (v) the enzyme activity in the thermal inactivation experiments was enhanced with increase in temperature with o-dianisidine as a substrate while its activity decreased with o-phenylenediamine.     

Effect of oxygen concentration on nitrogen removal by Nitrosomonas europaea and Paracoccus denitrificans immobilized within tubular polymeric gel

Tubular gel reactors containing Nitrosomonas europaea and Paracoccus denitrificans, which remove nitrogen from solutions through a process of nitrification and denitrification, require oxygen for ammonia oxidation, the first and rate-limiting step in the process. To accelerate ammonia oxidation, high concentrations of oxygen were applied to the reactors instead of air. Although a 50% O₂:N₂ gas mixture and pure oxygen were both toxic to free N. europaea cells, they actually accelerated ammonia oxidation by N. europaea immobilized within the tubular gel. Indeed, the rate of ammonia oxidation by a tube exposed to pure oxygen was twice that of one exposed to 20% O₂. When the distribution of N. europaea cells within the tubes was investigated using a fluorescently-labeled antibody, colonies were found on the external surface of the tube exposed to 20% O₂, but were located at a depth of 120–300 μm from the external surface in the case of the tube exposed to pure oxygen. The region between the external surface of the gel and the colonies apparently acted as a barrier, reducing the diffusion of oxygen and thus protecting the cells from oxygen cytotoxicity.     

Environmental factors modify carbon nutritional patterns and niche overlap between Aspergillus flavus and Fusarium verticillioides strains from maize

This study examined the utilization patterns of key carbon sources (CS, 24: including key sugars, amino acids and fatty acids) in maize by strains of Aspergillus flavus and Fusarium verticillioides under different water activity (a_w, 0.87–0.98 a_w) and temperature (20–35 °C) values and compared the niche overlap indices (NOI) that estimate the in vitro CS utilization profiles [Wilson, M., Lindow, S.E., 1994. Coexistence among epiphytic bacterial populations mediated through nutritional resource partitioning. Applied and Environmental Microbiology 60, 4468–4477.]. The ability to grow in these key CS in minimal media was studied for 120 h in 12 h steps. The NOI was calculated for inter-species (F. verticillioides–A. flavus) and for intra-species (A. flavus–A. flavus) using CS utilization patterns over the range of interacting environmental conditions. 30 °C, over the whole a_w range examined, was found to be optimal for utilization of the maximum number of CS by A. flavus. In contrast, for F. verticillioides this was more so at 20 °C; 25 °C allowed a suboptimal usage of CS for both species. NOIs confirmed the nutritional dominance of A. flavus at 30 °C, especially at lower a_w levels and that of F. verticillioides at 20 °C, mainly at 0.95 a_w. In other conditions of a_w, based on CS utilization patterns, the data indicated that A. flavus and F. verticillioides occupied different ecological niches. The variability in nutritional sources utilization between A. flavus strains was not related to their ability to produce aflatoxins (AFs). This type of data helps to explain the nutritional dominance of fungal species and strains under different environmental conditions. This could be useful in trying to find appropriate natural biocontrol microorganisms to compete with these mycotoxigenic species.     

Phytate degradation by human gut isolated Bifidobacterium pseudocatenulatum ATCC27919 and its probiotic potential

The growing awareness of the relationship between diet and health has led to an increasing demand for food products that support health above and beyond providing basic nutrition. Probiotics are live organisms present in foods, which yield health benefits related to their interactions with the gastrointestinal tract. Phytases are a subgroup of phosphatases that catalyse the desphosphorylation of phytate, which reduces its negative impact on mineral bioavailability, and generates lower inositol phosphates. The aims of this investigation were to (i) study the ability of the probiotic candidate Bifidobacterium pseudocatenulatum to degrade phytate in synthetic medium, to (ii) identify the lower inositol phosphates generated, to (iii) study its survival under conditions mimicking gastrointestinal passage and finally to (iv) assess adhesion of the bacteria to Caco-2 cells. The first steps of InsP₆ degradation by B. pseudocatenulatum phytate-degrading enzyme/s were preferentially initiated at the DL-6-position and 5-position of the myo-inositol ring. It suggests that the main InsP₆ degradation pathway by B. pseudocatenulatum by sequential removal of phosphate groups was D/L-Ins(1,2,3,4,5)P₅ or D/L-Ins(1,2,3,4,6)P₅; D/L-Ins(1,2,3,4)P₄; to finally Ins(1,2,3)P₃ and D/L-Ins(1,2,4)P₃/D/L-Ins(1,3,4)P₃. This human strain also showed a notable tolerance to bile as well as a selective adhesion capacity (adhesion to control surfaces was zero), to human intestinal Caco-2 cells comparable to the commercial probiotic B. lactis. The phytate-degrading activity constitutes a novel metabolic trait which could contribute to the improvement of mineral absorption in the intestine as a nutritional probiotic feature with potential trophic effect in human gut.     

Analysis of the role of the Listeria monocytogenes F0F1-ATPase operon in the acid tolerance response

As little is known about the genes involved in the induction of an acid tolerance response in Listeria monocytogenes, the role of the F₀F₁-ATPase was analyzed as a consequence of its role in the acid tolerance of a number of other bacteria and its conserved nature. It was found that acid adapted cells treated with N,N′-dicyclohexylcarbodiimide (DCCD) exhibited greatly enhanced sensitivity to low pH stress. Degenerate primers were designed to amplify and sequence a portion of the atpD gene. Subsequently, a PCR product from atpA to atpD was identified. While we were unable to create a deletion in the atpA gene, the plasmid pORI19 was inserted in a region between atpA and atpG to reduce, rather than eliminate, expression of the downstream genes. As expected this mutant displayed enhanced resistance to neomycin and exhibited slower growth than the wild type strain. This mutant could still induce an acid tolerance response and remained susceptible to DCCD treatment, but its relative acid sensitivity was difficult to assess as a consequence of its slow growth.     

基于比较转录组学的多花黄精黄酮类化合物合成基因表达分析

为了解药用植物多花黄精Polygonatum cyrtonema Hua.中黄酮类化合物的生物合成途径,为黄精属植物黄酮类化合物的代谢与调控的分子机制研究提供数据,利用二代高通量测序技术,对分别培养了 3个月和9个月的多花黄精组培苗根茎进行比较转录组学研究.结果显示,经序列组装与功能注释,多花黄精根茎的转录组数据经拼接...     

HUMULINIC ACIDS. VIII. STEREOISOMERS OF HUMULINIC ACIDS

Humulinic acid contains chiral centra at C₄ and C₅ and can therefore theoretically occur in four optically active stereoisomers. The diastereoisomeric forms are at the same time trans and cis isomers of each other (formerly humulinic acids A and B).⁵ This paper concerns the isolation and characterization of the trans and cis co- and ad-humulinic acids and the separation of racemic trans humulinic acid in its enantiomorphs.     

维氏气单胞菌来源几丁质酶的克隆表达及应用

丰年虾是一种小型甲壳类动物,孵化后的幼虫是水产动物的优质饵料,残留的卵壳则作为废弃物处理.然而丰年虾卵壳含有丰富的蛋白质和几丁质,具有极大的应用潜力.为了从丰年虾卵壳中制备几丁寡糖,选择了水解产物分布宽泛和含有几丁质结合域的维氏气单胞菌来源的几丁质酶ChiB565,将其在毕赤酵母Pichia pastoris中进行重组...