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Molecular Cloning and Characterization of Phenylalanine Ammonia-lyase and Cinnamate 4-Hydroxylase in the Phenylpropanoid Biosynthesis Pathway in Garlic (Allium sativum)

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Abstract

The cDNAs encoding phenylalanine ammonia-lyase (PAL) and cinnamate 4-hydroxylase (C4H) were cloned from garlic ( Allium sativum ) using reverse transcription-polymerase chain reaction (RT-PCR) with degenerate primers and 5' and 3' rapid amplification of cDNA ends (RACE) PCR. Amino acid sequence alignments showed that AsPAL and AsC4H have more than 70% amino acid identity with their homologues in other plants. The expression of AsPAL and AsC4H transcripts was highest in the roots but surprisingly low in the bulbils, where phenylpropanoid compounds are most concentrated. These results suggest that some phenylpropanoids are synthesized in the roots and subsequently transported to the bulbils of A. sativum .

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... Kaempferol, a flavonoid that has been isolated from many plants [1][2][3][4], has many important health-promoting effects, such as antioxidant, antimicrobial, and anti-inflammatory effects, and is known to decrease the risk of cancers, including bladder, colon, and squamous cell cancers [5][6][7][8][9]. Flavonoid glycosides are known to be more abundant than their flavonoid monomers in plants and exhibit a broad range of biological activities. ...
... On the basis of previous studies by our group, we know that the expression of different phenylpropanoid biosynthetic genes in different organs is species-specific [1,2,4,29,30]. For example, in Prunella vulgaris [29] and Lycium chinense [4], the highest expression of the first enzyme-coding gene in the phenylpropanoid pathway PAL was observed in flowers. ...
... For example, in Prunella vulgaris [29] and Lycium chinense [4], the highest expression of the first enzyme-coding gene in the phenylpropanoid pathway PAL was observed in flowers. However, in Fagopyrum esculentum [1] and Allium sativum [2], the expression of PAL was highest in roots, and was also highly expressed in the stems of F. esculentum. Similarly, in Scutellaria baicalensis, the highest and lowest expressions of FLS were observed in the roots and leaves, respectively [30]. ...
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Kenaf (Hibiscus cannabinus) is cultivated worldwide for its fiber; however, the medicinal properties of this plant are currently attracting increasing attention. In this study, we investigated the expression levels of genes involved in the biosynthesis of kaempferitrin, a compound with many biological functions, in different kenaf organs. We found that phenylalanine ammonia lyase (HcPAL) was more highly expressed in stems than in other organs. Expression levels of cinnamate 4-hydroxylase (HcC4H) and 4-coumarate-CoA ligase (Hc4CL) were highest in mature leaves, followed by stems and young leaves, and lowest in roots and mature flowers. The expression of chalcone synthase (HcCHS), chalcone isomerase (HcCHI), and flavone 3-hydroxylase (HcF3H) was highest in young flowers, whereas that of flavone synthase (HcFLS) was highest in leaves. An analysis of kaempferitrin accumulation in the different organs of kenaf revealed that the accumulation of this compound was considerably higher (>10-fold) in leaves than in other organs. On the basis of a comparison of kaempferitrin contents with the expression levels of different genes in different organs, we speculate that HcFLS plays an important regulatory role in the kaempferitrin biosynthetic pathway in kenaf.
... The expression patterns of IdPALs, IdC4H, Id4CLs, IdCHS, and IdDFR were analyzed in the flower, bud, leaf, stem, and root using qRT-PCR ( Figure 1). Our group previously investigated the expression patterns of phenylpropanoid biosynthetic genes in buckwheat [21], garlic [22], mulberry [14], and Chinese cabbage [16]. In the current study, we observed various expression patterns among phenylpropanoid biosynthetic gene isoforms in different organs. ...
... Thus, the expression of IdPAL1, IdC4H, Id4CL1, IdCHS, and IdDFR was the highest in the leaf. To date, our group has investigated the expression patterns of phenylpropanoid biosynthetic genes in various plants [21][22][23][24][25][26]. Richet et al. [27] suggested phenylpropanoid gene expression in different organs of poplar could be altered by ozone exposure. ...
... Molecules 2017,22, 901 ...
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Members of the genus Ixeris have long been used in traditional medicines as stomachics, sedatives, and diuretics. Phenylalanine ammonia-lyase (PAL), cinnamate-4-hydroxylase (C4H), 4-coumarate: coenzyme-A (CoA) ligase (4CL), chalcone synthase (CHS), and dihydroflavonol 4-reductase (DFR) are important enzymes in the phenylpropanoid pathway. In this study, we analyzed seven genes from Ixeris dentata var. albiflora that are involved in phenylpropanoid biosynthesis, using an Illumina/Solexa HiSeq 2000 platform. The amino acid sequence alignments for IdPALs, IdC4H, Id4CLs, IdCHS, and IdDFR showed high identity to sequences from other plants. We also investigated transcript levels using quantitative real-time PCR, and analyzed the accumulation of phenylpropanoids in different organs of I. dentata var. albiflora using high-performance liquid chromatography. The transcript levels of IdC4H, Id4CL1, IdCHS, and IdDFR were highest in the leaf. The catechin, chlorogenic acid, ferulic acid, and quercetin contents were also highest in the leaf. We suggest that expression of IdC4H, Id4CL1, IdCHS, and IdDFR is associated with the accumulation of phenylpropanoids. Our results may provide baseline information for elucidating the mechanism of phenylpropanoid biosynthesis in different organs of I. dentata var. albiflora.
... 12,13 Recently, the molecular cloning and characterization of PAL and cinnamate 4-hydroxylase (C4H) in garlic have been described. 14 In addition, the complete transcriptome in bud garlic cloves stored at RT using an Illumina sequencing system has been reported; 127 933 unigenes were found and categorized into 25 groups of different physiological functions, where the authors gave more emphasis to those genes involved in the biosynthesis of aroma compounds. 15 Therefore, the objective of this study was to measure changes in the content of phenolic and anthocyanin compounds in garlic bulbs cv. ...
... The results in the present study are higher than those in previous reports that analyzed whole bulbs or cloves of the plant. 14,21,22 It has also been reported that cataphylls of onion bulbs have higher phenolic and anthocyanin content with respect to other plant organs. 23 In general, these data showed that cold conditioning of "seed" cloves has an important effect on the metabolism of phenolic compounds and anthocyanins, promoting their biosynthesis during plant growth in the field. ...
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Low-temperature conditioning of garlic "seed" cloves accelerated the development of the crop cycle, decreased plant growth, and increased the synthesis of phenolic compounds and anthocyanins in the outer scale leaves of the bulbs at harvest time, leading to 3-fold content increase compared with those conditioned at room temperature. Cold conditioning of "seed" cloves also altered the anthocyanin profile during bulb development and at harvest. Two new anthocyanins are reported for the first time in garlic. The high phenolics and anthocyanin contents in bulbs of plants generated from "seed" cloves conditioned at 5 °C for 5 weeks were preceded by overexpression of some putative genes of the phenolic metabolism [6-fold for phenylalanine ammonia lyase (PAL)] and anthocyanin synthesis [1-fold for UDP-sugar:flavonoid 3-O-glycosyltransferase (UFGT)] compared with those conditioned at room temperature.
... cDNA was obtained from the RNA extract using the PrimeScript RT Reagent Kit (Takara Bio, Ohtsu, Japan). In a preliminary experiment, more than 20 combinations of PAL sequences of such plants as Allium sativum (Tuan et al., 2010;Forward (F): 5´-TMCARG GMTACTCH GGCATMMG-3´, Reverse (R): 5´-GCGCTYTNSACRTGGTTNGT VA-3´), Euphorbia pulcherrima (Tao et al., 2011; 5´-GGAAAATTACTTCTCAAG-3´, R: 5´-TACATAGAAGGG ATTACA-3´), Lactuca sativa var. longifolia (Campos et al., 2004; F: 5´-GGCCACGCGTCGA CTAGTAC-3´) was performed. ...
... The molecular mass was calculated as about 78 kDa. The amino acid sequence contained the highly conserved PAL motif (Tuan et al., 2010) including an active site (Ala-Ser-Gly), and showed high homology to PAL genes of Fabaceae plants (Table 1). These results indicated that the cloned gene was a mung bean PAL. ...
Article
Mung bean sprout turns brown during cold storage. Here we cloned phenylalanine ammonia-lyase (PAL) mRNA and examined its expression during cold storage. A PAL cDNA of mung bean encoding 715 amino acids was cloned, which showed high homology to PAL genes of Fabaceae plants and had a highly conserved PAL motif including the active site (Ala-Ser-Gly). The expression of PAL mRNA increased during cold storage of mung bean sprout. The increase in expression was inhibited by the heat-shock treatment of mung bean sprout before storage. trans-Coumaroyltartronic and trans-caffeoyltartronic acids, the major substrates of polyphenol oxidase in mung bean sprout increased during cold storage. These results show that the PAL expression and biosynthesis of phenolics are essential for enzymatic browning of mung bean sprout during cold storage.
... The forward reaction involves deamination of L-phenylalanine (L-Phe) to trans-cinnamic acid (t-Ca) and ammonia, while, the reversal reaction involves biotransformation of trans-cinnamic acid to L-phenylalanine [4,5], as illustrated in Fig. 1A. PAL is widely distributed in plants, acting as a key enzyme in phenylpropanoid pathway [6,7], controlling the production of secondary metabolites such as flavonoids, coumarins, lignins, stilbenes and phytoalexins [7]. PAL has also been reported found in fungi, yeasts and bacteria [4]. ...
... The forward reaction involves deamination of L-phenylalanine (L-Phe) to trans-cinnamic acid (t-Ca) and ammonia, while, the reversal reaction involves biotransformation of trans-cinnamic acid to L-phenylalanine [4,5], as illustrated in Fig. 1A. PAL is widely distributed in plants, acting as a key enzyme in phenylpropanoid pathway [6,7], controlling the production of secondary metabolites such as flavonoids, coumarins, lignins, stilbenes and phytoalexins [7]. PAL has also been reported found in fungi, yeasts and bacteria [4]. ...
Article
Active lipase-producing bacterium Burkholderia gladioli Bps-1 was rapidly isolated using a modified trypan blue and tetracycline, ampicillin (TB-TA) plate. The electro-phoretically pure enzyme was obtained by purification using ethanol precipitation, ion-exchange chromatography, and gel filtration chromatography. The molecular weight was 34.6 kDa and the specific activity was determined to be 443.9 U/mg. The purified lipase showed the highest activity after hydrolysis with p-NPC16 at a pH of 8.5 and 50°C, and the Km, kcat, and kcat/Km values were 1.05, 292.95 s-1 and 279 s-1mM-1, respectively. The lipase was highly stable at 7.5 ≤ pH ≤ 10.0. K+ and Na+ exerted activation effects on the lipase which had favorable tolerance to short-chain alcohols with its residual enzyme activity being 110% after being maintained in 30% ethanol for 1 h. The results demonstrated that the lipase produced by the strain B. gladioli Bps-1 has high enzyme activity and is an alkaline lipase. The lipase has promising chemical properties for a range of applications in the food-processing and detergent industries, and has particularly high potential for use in the manufacture of biodiesel.
... It is reported that asarinin is derived from phenylpropanoid biosynthesis (ko00940) ). The function of enzyme genes such as PAL, C4H, 4CL, and CCR in phenylpropanoid biosynthesis have been thoroughly studied in many plants (Maher et al. 1994;Liu et al. 2006;Tuan et al. 2010). Nevertheless, the key enzyme genes of phenylpropanoid biosynthesis in A. sieboldii have been rarely reported, only PAL and CAD were cloned and further researched (Lin et al. 2018;Liu et al. 2018). ...
... C4H, phenylalanine ammonialyase (PAL), 4-coumarate: CoA ligase (4CL) and chalcone synthase (CHS) also play a key role in the regulation of lignin biosynthesis [5]. The sequences and expression patterns of C4H have been reported in several plants such as poplar (Populus trichocarpa £ P. deltoides) [6], tea (Camellia sinensis) [7], oilseed rape (Brassica napus) [8], garlic (Allium sativum) [9], Scutellaria baicalensis [10], Ornithogalum saundersiae [11] and Angelica gigas [12]. These studies provided basic information of C4H as an enzyme in synthetic pathways in plants. ...
Article
The present study aimed to describe the cDNA cloning of the cinnamate 4-hydroxylase (C4H) gene from ramie, predict the protein sequence and perform phylogenetic and structural analyses of this gene. C4H catalyses the hydroxylation of trans-cinnamic acid to p-coumaric acid, which plays a crucial role in lignin biosynthesis. In the present study, the gene encoding C4H was cloned by polymerase chain reaction (PCR) technology from cultivated ramie (Boehmeria nivea) and named BnGC4H. In addition, bioinformatics and tissue expression analyses were performed. The results showed that the cloned BnGC4H cDNA contained a 1518-bp open reading frame encoding a 505–amino acid protein. The sequence of BnGC4H is available from the GenBank database with accession number KY937946. The amino acid sequence and structural analysis revealed that BnGC4H shared conserved domains with other C4H forms, including cytochrome P450 domain and trans-cinnamate 4-monooxygenase domain. The phylogenetic analysis indicated that BnGC4H was closely related to Aquilaria sinensis C4H and Ruta graveolens C4H. Real-time quantitative reverse transcription–PCR and RNA in situ hybridization showed that BnGC4H was strongly expressed in the xylem during the maturity stage. The findings provided a theoretical basis for further exploring the function of BnGC4H in the lignin biosynthesis and regulation in ramie.
... All experiments were performed in triplicate. The experimental conditions were similar as previously described (Tuan et al., 2010). Carotenoids were extracted from M. charantia samples (0.1 g) with 3 ml of ethanol containing 0.1% ascorbic acid (w/v). ...
Article
Momordica charantia, a tropical plant, produces a fruit that has a β-carotene concentration five times higher than that of carrot. To elucidate the molecular basis of β-carotene accumulation in M. charantia, the gene expression levels of phytoene synthase (McPSY) and phytoene desaturase (McPDS) were determined. These levels were particularly high in the flowers of M. charantia. During fruit maturation, the expression levels of McPSY and McPDS decreased during the mid-stages but increased in the fully mature fruit. In addition, carotenoids accumulated as the peel changed from green to orange. Thus, McPSY and McPDS expression correlated with carotenoid accumulation during fruit maturation. Principal component analysis (PCA) also was used to evaluate the differences among the profiles of seven carotenoids identified in the fruit at several maturation stages. Riper fruits had higher carotenoid concentrations than less ripe fruits.
... sativum, the highest expression level occurred in the root, but in plant Agastache. rugosa, ArC4H was expressed highest in flowers [28,29]. In L. chinense, we found that the expression levels of LcC4H in red fruits was significantly higher than that that in other organs. ...
Article
Full-text available
Lycium chinense is well known in traditional Chinese herbal medicine for its medicinal value and composition, which have been widely studied for decades. However, further research on Lycium chinense is limited due to the lack of transcriptome and genomic information. The transcriptome of L. chinense was constructed by using an Illumina HiSeq 2000 sequencing platform. All 56,526 unigenes with an average length of 611 nt and an N50 equaling 848 nt were generated from 58,192,350 total raw reads after filtering and assembly. Unigenes were assembled by BLAST similarity searches and annotated with Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) orthology identifiers. Using these transcriptome data, the majority of genes that are associated with phenylpropanoid biosynthesis in L. chinense were identified. In addition, phenylpropanoid biosynthesis-related gene expression and compound content in different organs were analyzed. We found that most phenylpropanoid genes were highly expressed in the red fruits, leaves, and flowers. An important phenylpropanoid, chlorogenic acid, was also found to be extremely abundant in leaves. Using Illumina sequencing technology, we have identified the function of novel homologous genes that regulate metabolic pathways in Lycium chinense.
... In higher plants, phenylpropanoid biosynthetic pathway produces the important metabolites, e.g. flavonoids, isoflavonoids, lignin, anthocyanin, phytoalexins, antimicrobial furanocoumarins, hydroxyl cinnamate esters and phenolic esters, which are all critical players in development, structural protection, defence responses to microbial attacks and tolerance to abiotic stimuli [10,11]. As phenylpropanoid pathway is a gateway for production of many secondary metabolites [12,13], the investigation of characteristics as well as expression patterns of involved genes in production of these metabolites, e.g. ...
Article
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Safflower (Carthamus tinctorius L.) serves as a reference dicot for investigation of defense mechanisms in Asteraceae due to abundant secondary metabolites and high resistance/tolerance to environmental stresses. In plants, phenylpropanoid and flavonoid pathways are considered as two central defense signaling cascades in stress conditions. Here, we describe the isolation of two major genes in these pathways, phenylalanine ammonia-lyase (CtPAL) and chalcone synthase (CtCHS) in safflower along with monitoring their expression profiles in different stress circumstances. The amino acid sequence of isolated region of CtPAL possesses maximum identity up to 96% to its ortholog in Cynara scolymus, while that of CtCHS retains the highest identity to its ortholog in Callistephus chinensis up to 96%. Experiments for gene expression profiling of CtPAL and CtCHS were performed after treatment of seedlings with 0.1 and 1 mM salicylic acid (SA), wounding, and salinity stress. The results of semi-quantitative RT-PCR revealed that both CtPAL and CtCHS genes are further responsive to higher concentration of SA with dissimilar patterns. Regarding wounding stress, CtPAL gets slightly induced upon injury at 3 hours after treatment (hat), while CtCHS gets greatly induced at 3 hat and levels off gradually afterward. Upon salinity stress, CtPAL displays a similar expression pattern by getting slightly induced at 3 hat, but CtCHS exhibits a biphasic expression profile with two prominent peaks at 3 and 24 hat. These results substantiate the involvement of phenylpropanoid and particularly flavonoids pathways in safflower during wounding and especially salinity stress.
... In addition, some of these isoforms are involved in the biosynthesis of 4-coumaroyl CoA-derived flavonoids [9]. Recently, our research group isolated PAL, C4H, and chalcone isomerase (CHI) from Scutellaria baicalensis [10,11]; PAL from Angelica gigas [12]; and C4H from Allium sativum [13]. Agrobacterium rhizogenes-mediated transformation is a rapid and versatile method to engineer plants genetically, as was reported for S. baicalensis [14][15][16]. ...
Article
Scutellaria baicalensis Georgi, a species of the Lamiaceae family, is considered as one of the 50 fundamental herbs used in traditional Chinese medicine. In order to enhance flavone (baicalein, baicalin, and wogonin) content in S. baicalensis roots, we overexpressed a single gene of cinnamate 4-hydroxylase (C4H) and 4-coumaroyl coenzyme A ligase (4CL) using an Agrobacterium rhizogenes-mediated system. SbC4H- and Sb4CL-overexpressed hairy root lines enhanced the transcript levels of SbC4H and Sb4CL compared with those in the control and also increased flavones contents by approximately 3- and 2.5-fold, respectively. We successfully engineered the flavone biosynthesis pathway for the production of beneficial flavones in S baicalensis hairy roots. The importance of upstream gene C4H and 4CL in flavone biosynthesis and the efficiency of metabolic engineering in promoting flavone biosynthesis in S. baicalensis hairy roots have been indicated in this study.
... All reactions were performed using the SYBR Green Real-Time PCR Mix according to the manufacturer's instructions. The experimental conditions were similar as previously described (Tuan et al. 2010;Lü et al. 2015). Each 20 μL qRT-PCR reaction mixture containing 1 μL cDNA, 1 μL forward primer (10 μmol L -1 ), 1 μL reverse primer (10 μmol L -1 ), 10 μL 2× SYBR Green Real-Time PCR Mix, nuclease-free water to final volume of 20 μL, was preheated at 95°C for 5 min, followed by 45 cycles of 95, 60 and 72°C for 30 s, respectively. ...
Article
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The difference between lycopene and phytohormone levels among diploid, triploid and tetraploid plants of two watermelon cultivars during fruit growth and ripening was studied. The expression pattern of five genes (phytoene synthase (PSY1), phytoene desaturase (PDS), ζ-carotene desaturase (ZDS), carotenoid isomerase (CRTISO), and lycopene β-cyclase (LCYB)) was analyzed in details. In red-fleshed cultivar Mimei, lycopene content increased rapidly from 25 to 35 days after pollination (DAP), and then decreased at 40 DAP. Triploid and tetraploid fruit had higher levels of lycopene than diploid. Moreover, triploid tended to contain more lycopene than tetraploid during fruit growth and ripening stages. However, little amount of lycopene (0–2 mg kg⁻¹ fresh weight (FW)) in yellow-fleshed cultivar Huangmei was found during all fruit development stages. In Mimei, transcript level of PSY1 was generally higher than the other four genes, and LCYB gene expression was the lowest among all five genes being tested. PSY1, CRTISO and LCYB genes showed higher transcript levels in polyploid than in diploid fruit. By contrast, in Huangmei, transcript level of LCYB was not the lowest, but only lower than that of PSY1. PSY1, CRTISO and LCYB genes showed higher expression levels in diploid than in polyploid fruit. In Mimei, the negative correlation between gibberellane (GA) content and lycopene accumulation was determined in all three different ploidy fruits, while a positive correlation was observed between abscisic acid (ABA) content and lycopene accumulation only in diploid watermelon. These results indicated that different lycopene contents in different ploidy watermelons is regulated by the differential transcription expression of the lycopene metabolic genes and phytohormones.
... Therefore, the change behaviors of phenolic compounds indicate that day 12 may be the key processing time of "Laba" garlic. On the other hand, dietary intake of avonoids in avonoid-rich foods has presented benets to human health, 44 which implies the potential nutritional value of "Laba" garlic with higher content of phenolic compounds at day 12. Fig. 4 illustrated the metabolic pathways of the 15 phenolic compounds in "Laba" garlic according to the phenylpropanoid biosynthesis pathway, 45 which provides a detailed transformation relationship between these phenolic compounds. For example, one part of cinnamic acid can be transformed into (+)-catechin and apigenin, another part of cinnamic acid is hydroxylated into p-coumaric acid and further converted into a number of phenolic compounds, such as caffeic acid, ferulic acid and quercitrin. ...
Article
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"Laba" garlic is a famous traditional garlic product, particularly popular with the people in Northern China. The processing time plays an important role on the chemical constituents of "Laba" garlic. Here, we investigated the composition of "Laba" garlic during traditional processing using a non-targeted metabolomics approach. Through using gas chromatography mass spectrometry (GC-MS) and liquid chromatography mass spectrometry (HPLC-MS/MS) with multivariate analysis, a total of 20 volatile metabolites, 16 primary metabolites and 15 phenolic compounds were identified as notable changed compounds due to the traditional processing (p < 0.05). The characteristic flavor of "Laba" garlic was formed mainly by the decreased content of organosulfur compounds and the increased content of non-organosulfur compounds. In addition, this study also proposed the metabolic pathway of primary metabolites and phenolic compounds in garlic samples during processing. Most primary metabolites including lactic acid, isocitric acid, l-leucine, l-proline, d-fructose, d-glucose and erythritol increased from day 3, and reached the maximum level at day 12, which were thought to be the foundation for the sweet and sour taste of "Laba" garlic. Although the concentration of the 15 phenolic compounds from day 3 to day 42 was notably higher than the raw garlic (0 day), the antioxidant activities showed a decreasing trend from day 0 to day 28. The correlation analysis result revealed that notable positive associations were presented between organosulfur compounds, organic acids, amino acids and antioxidant activities. Furthermore, day 12 was found to be the most suitable time to obtain the "Laba" garlic considering its colour, flavor, taste and physiological function. These results are helpful in comprehending metabolism changes and physiological function of "Laba" garlic traditional processing.
... described for Arabidopsis thaliana, potato, bean, alfalfa, and poplar (Ohl et al., 1990;Gowri et al., 1991;Jots and Hahlbrock, 1992;Subramaniam et al., 1993). Tissue-specific expression of PALs in plants such as Scutellaria baicalensis (Xu et al., 2012), raspberry (Kumar and Ellis, 2001), common buckwheat , and garlic (Tuan et al., 2010) has shown clearly that predominantly PALs are expressed more in roots and stems than in leaves. In the current study, ChPALs presented similar tissue-specific patterns. ...
Article
Phenylalanine ammonia lyase (PAL) is the first committed step in the formation of phenylpropanoids, and catalyses the deamination of L-phenylalanine (L-Phe) to yield cinnamic acid. While PALs are common in plants, PAL genes involved in alkaloid biosynthesis in Cephalotaxus hainanensis have never been described. To obtain better knowledge of PAL genes and their number and function involved in Cephalotaxus alkaloid biosynthesis four PAL genes were screened and cloned. In vitro enzymatic analysis showed that all four PAL recombinant proteins could convert L-Phe to product trans-cinnamic acid, and showed strict substrate specificity. Moreover, the expression profiles of four ChPALs were analysed using qRT-PCR, which showed that they had higher transcript levels in roots and stems, and that different ChPALs displayed different response sensitivities and change patterns in response to stimuli. Several metabolic compounds were measured in stimulated leaves using UPLC-MS, and indicating the concentration of Cephalotaxus alkaloids and cinnamic acid in leaves subjected to different conditions. These concentrations increased significantly after treatment with 100 mM NaCl, 100 mM mannitol, 100 μM SA and 10 μM ABA. The expression levels of four PAL genes showed indications of upregulation after treatment. These results supply an important foundation for further research on candidate genes involved in the biosynthesis of Cephalotaxus alkaloids.
... Furthermore, the expression studies revealed the tissue-specific expressions where the genes expression was lower in the leaf samples of P. equestris and D. catenatum. Studies on the tissue-specific expression of PALs in plants such as raspberry, common buckwheat, garlic and skullcap have shown that PAL genes are expressed more in other tissues as compared to leaf tissue (Kumar & Ellis, 2001;Li et al., 2010;Tuan et al., 2010;Xu et al., 2012). ...
Article
Phenylalanine ammonia-lyase (PAL) is a key gateway enzyme that connects the phenylpropanoid pathway to primary metabolism. The phenylpropanoid pathway plays a vital role in the growth and environmental adaptation of many plants leading to the production of valuable bioactive compounds with industrial and medical applications. In the present study, nine putative PAL genes from three orchids were identified; five in Apostasia shenzhenica and two each in Dendrobium catenatum and Phalaenopsis equestris. Eighteen motifs and four major conserved functional domains were identified as reported in PAL proteins of other species. All the nine PALs were stable based on their computed physicochemical properties and localized in the cytoplasm. The three-dimensional structures of PALs revealed a homo-tetrameric structure consisting of four identical subunits. A total of 21 cis-regulatory elements with known functions were identified from the promoter regions of all PALs which are responsible for various plant responses to light, stress and growth regulators like auxins, gibberellins and abscisic acid. Phylogenetic analysis showed that the studied PAL proteins clustered in two major clades (clade I and II), placing dicot and monocot PALs in two separate monophyletic clades. In silico gene expression of the identified PALs in different vegetative and reproductive tissues revealed the differential expressions based on tissue type and disclosed that the expression of PAL genes was upregulated in all the tissues examined with an exception of PePAL leaf samples where no expression was detected, however, the same being highly expressed in reproductive tissues (PePAL1-labellum; PePAL2-sepal). In case of AsPALs, the expression was found to be highest in reproductive tissues (AsPAL4-maximum in inflorescence). On the other hand, the expression of DcPALs was found to be highest in vegetative tissues (DcPAL2-maximum in root). Based on the medicinal importance of orchids and the significant role of PAL genes in synthesis of bioactive compounds, the functional characterization of PAL genes can be further exploited in genetic improvement of medicinal orchids. Communicated by Ramaswamy H. Sarma
... PAL is the first rate-limiting enzyme in the pathway, catalyzing the formation of trans-cinnamic acid via the L-deamination of phenylalanine; C4H hydroxylates trans-cinnamic acid at the C4 position to produce p-coumaric acid; next, 4CL catalyzes the production of p-coumaroyl-CoA. Subsequently, the p-coumaroyl-CoA is transformed into phenylpropanoid compounds with diverse functions in plants (Tuan et al. 2010). Recently, exogenous application of plant hormones to elicit increased yield of secondary metabolites has been practiced as a short-term strategy for plantations to improve their yield (Horbowicz et al. 2011;Kurepin et al. 2017). ...
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Prior to Ginkgo leaf harvest, we implemented four exogenous applications of salicylic acid (SA: 50, 80, and 100 mg·L ⁻¹ ) or methyl jasmonate (MJ: 20, 40, and 80 mg·L ⁻¹ ) as leaf spray to investigate their effect on flavonoids content and related enzyme activity. Compared with the control, a significant increase of 12% and 26% in flavonoid content was observed 7 days after the first application of 100 mg·L ⁻¹ SA and 20 mg·L ⁻¹ MJ, respectively; both were determined to be the best treatments. Operationally, administering either elicitor (SA or MJ) 1 week prior to leaf harvest is deemed to be the most practical and economic option. Additionally, enzymes activity involved in flavonoid metabolism (phenylalanine ammonia-lyase, cinnamate 4-hydroxylase, and 4-coumarate:coenzyme A ligase) and antioxidants (peroxidase, superoxide dismutase, and catalase) significantly increased in most SA and MJ treatments. The results strongly support Ginkgo leaf response to SA- or MJ-induced stress through increased nonenzymatic antioxidant compounds (flavonoids) and activity of antioxidant enzymes.
... C4H and 4CL had high expression levels in the root and stem but lower levels in the leaf (Fig. 7). In Allium sativum [23] and Polygonum minus [24], the highest expression of C4H occurred in the root, while the expression of 4CL in different organs differed among species [25,26]. ...
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Background Polygonum cuspidatum Sieb. et Zucc. is a well-known medicinal plant whose pharmacological effects derive mainly from its stilbenes, anthraquinones, and flavonoids. These compounds accumulate differentially in the root, stem, and leaf; however, the molecular basis of such tissue-specific accumulation remains poorly understood. Because tissue-specific accumulation of compounds is usually associated with tissue-specific expression of the related biosynthetic enzyme genes and regulators, we aimed to clarify and compare the transcripts expressed in different tissues of P. cuspidatum in this study. Results High-throughput RNA sequencing was performed using three different tissues (the leaf, stem, and root) of P. cuspidatum . In total, 80,981 unigenes were obtained, of which 40,729 were annotated, and 21,235 differentially expressed genes were identified. Fifty-four candidate synthetase genes and 12 transcription factors associated with stilbene, flavonoid, and anthraquinone biosynthetic pathways were identified, and their expression levels in the three different tissues were analyzed. Phylogenetic analysis of polyketide synthase gene families revealed two novel CHS genes in P. cuspidatum . Most phenylpropanoid pathway genes were predominantly expressed in the root and stem, while methylerythritol 4-phosphate and isochorismate pathways for anthraquinone biosynthesis were dominant in the leaf. The expression patterns of synthase genes were almost in accordance with metabolite profiling in different tissues of P. cuspidatum as measured by high-performance liquid chromatography or ultraviolet spectrophotometry. All predicted transcription factors associated with regulation of the phenylpropanoid pathway were expressed at lower levels in the stem than in the leaf and root, but no consistent trend in their expression was observed between the leaf and the root. Conclusions The molecular knowledge of key genes involved in the biosynthesis of P. cuspidatum stilbenes, flavonoids, and anthraquinones is poor. This study offers some novel insights into the biosynthetic regulation of bioactive compounds in different P. cuspidatum tissues and provides valuable resources for the potential metabolic engineering of this important medicinal plant.
... Compounds were monitored at 310 nm. Analysis of reference standards facilitated the preparation of a standard curve which was used to quantify concentrations of coumarin in plant extracts, as reported previously Tuan et al., 2010). The amount of each compound in the different samples was obtained from the concentration and the known volume (900 μL) and the mass percentages were calculated based on the dry weight of each sample (5.5 mg). ...
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Garlic is characterised by medicinal properties due to the content of over 2000 biologically active substances. Numerous commercially processed garlic forms, which differ in the content of bioactive compounds, especially sulphuric, are available on the market. The knowledge of the types of bioactive substances present in garlic and its products, their changes during treatment and pro-health influence is of crucial importance to the diet supplement producers, doctors, pharmacists and consumers. Therefore, this work has aimed to characterise the most important bioactive substances of garlic, its preparations and describe in detail the role of garlic in dietoprophylaxis and dietotherapy.
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Flavonoids are synthesized in response to developmental and environmental signals and perform many functions in plants. Arabidopsis (Arabidopsis thaliana) roots grown in complete darkness do not accumulate flavonoids since the expression of genes encoding enzymes of flavonoid biosynthesis is light dependent. Yet, flavonoids accumulate in root tips of plants with light-grown shoots and light-shielded roots, consistent with shoot-to-root flavonoid movement. Using fluorescence microscopy, a selective flavonoid stain, and localized aglycone application to transparent testa mutants, we showed that flavonoids accumulated in tissues distal to the application site, indicating uptake and movement systems. This was confirmed by time-course fluorescence experiments and high-performance liquid chromatography. Flavonoid applications to root tips resulted in basipetal movement in epidermal layers, with subsequent fluorescence detected 1 cm from application sites after 1 h. Flavonoid application to midroot or cotyledons showed movement of flavonoids toward the root tip mainly in vascular tissue. Naringenin, dihydrokaempferol, and dihydroquercetin were taken up at the root tip, midroot, or cotyledons and traveled long distances via cell-to-cell movement to distal tissues, followed by conversion to quercetin and kaempferol. In contrast, kaempferol and quercetin were only taken up at the root tip. Using ATP-binding cassette (ABC) transporter and H(+)-ATPase inhibitors suggested that a multidrug resistance-associated protein ABCC transporter facilitated flavonoid movement away from the application site.
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Phenylalanine ammonia-lyase (PAL; EC 4.3.1.5) genomic sequences were isolated from bean (Phaseolus vulgaris L.) genomic libraries using elicitor-induced bean PAL cDNA sequences as a probe. Southern blot hybridization of genomic DNA fragments revealed three divergent classes of PAL genes in the bean genome. Polymorphic forms were observed within each class. The nucleotide sequences of two PAL genes, gPAL2 (class II) and gPAL3 (class III), were determined. gPAL2 contains an open reading frame encoding a polypeptide of 712 amino acids, interrupted by a 1720 bp intron in the codon for amino acid 130. gPAL3 encodes a polypeptide of 710 amino acids showing 72% similarity with that encoded by gPAL2, and contains a 447 bp intron at the same location. At the nucleotide level, gPAL2 and gPAL3 show 59% sequence similarity in exon I, 74% similarity in exon II, and extensive sequence divergence in the intron, 5' and 3' flanking regions. S1 nuclease protection identified transcription start sites of gPAL2 and gPAL3 respectively 99 bp and 35 bp upstream from the initiation codon ATG, and showed that gPAL2 but not gPAL3 was activated by elicitor, whereas both were activated by wounding of hypocotyls. The 5' flanking region of both genes contain TATA and CAAT boxes, and sequences resembling the SV40 enhancer core. gPAL2 contains a 40 bp palindromic sequence and a 22 bp motif that are also found at similar positions relative to the TATA box in 5' flanking regions of other elicitor-induced bean genes.
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The cDNAs (Espals) encoding phenylalanine ammonia-lyase (PAL) were cloned from Ephedra sinica by reverse transcription polymerase chain reaction (RT-PCR) using degenerate primers and by 5′ and 3′-rapid amplification of cDNA ends (RACE). 2166 bp of the open reading frame (ORF) encoded 722 amino acids; sequence analyses of Espal clones suggested that at least four isoforms of EsPAL (EsPAL1, 2, 3, 4) existed, with nine amino acids substitution in their sequences. Phylogenetic analysis of EsPAL and PALs from other plant species revealed that EsPAL and Pinus PAL formed a gymnosperm-type PAL subfamily. The recombinant EsPAL1 to 4 functionally catalyzed a PAL reaction and their Km, Vmax, Kcat and Kcat/Km values did not show significant differences. Semi-quantitative RT-PCR analysis indicated that the expression of Espal genes in the roots was higher than in the plant's aerial parts. In addition, the activity of PAL in the roots was also higher than in the aerial parts. These results suggest that Espal genes are expressed in the whole plant but are dominant in the roots rather than in the aerial parts.
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Bananas are highly susceptible to chilling injury (CI) and phenylalanine ammonia-lyase (PAL), as a key enzyme involved in plant phenylpropanoid metabolism, has been associated with low temperature stress in plant tissues. However, little is known about the role of PAL (including PAL activity, gene and protein expression) in postharvest chilling tolerance of banana fruit. Two partial cDNAs sequences (MaPAL1 and MaPAL2) with about 760bp were cloned from banana pulp by RT-PCR. Western and northern hybridizations were used to investigate expression of PAL protein and PAL genes in fruit stored for 10 days at 7°C (chilling temperature) and then transferred to 22°C (room temperature). The effects of propylene (a functional ethylene analog) on their expression in relation to CI were also examined. Northern and western blot analyses revealed that mRNA transcripts of MaPAL1 and MaPAL2 and PAL protein levels in banana fruit during storage increased, reaching a peak at about day 8, and finally decreased at chilling temperature. Prior to low temperature storage, pretreatment with propylene could alleviate CI and enhance PAL activity, protein amount and mRNA transcripts of MaPAL1 and MaPAL2. Moreover, changes in PAL activity, protein amount and accumulation of MaPAL1 and MaPAL2 exhibited almost the same patterns. The results suggest that the induction of PAL in banana fruit during low temperature storage is regulated at transcriptional and translational levels, and is related to reduction in CI symptoms.
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Flavonol levels in the edible portions of Allium vegetables (leeks, shallots, green onions, garlic and onions) range from <0.03 to > 1 g/kg of vegetable. Shallots contained uniformly high concentration of total flavonols, >800 mg/kg in each of five independent samples. However, onions varied widely in the amounts of flavonoids they contained. White onions contained no detectable flavonols, but 20 cultivars of yellow and red onions contained between 60 mg/kg and >1000 mg/kg. Individual samples of leeks, garlic, and scallions contained undetectable levels of flavonols.
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Five cDNA clones encoding phenylalanine ammonia-lyase from a barley root cDNA library were isolated. Southern analysis of barley genomic DNA also revealed at least five different pal genes in barley genome. Three nearly full-length cDNA clones (hpal2, hpal3, and hpal7) were further characterized and found to be highly similar to wheat and rice PAL sequences. The amount of pal transcripts was highest in the roots and lowest in the leaves. Transcript levels of pal were rapidly and strongly, but transiently, increased after treatment with mercuric chloride, whereas the expression was later after fungal inoculation.
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The antioxidant activities of polar fractions of mature garlic bulbs and immature plants in four different model systems are presented. Antioxidant activity was evaluated as free radical-scavenging capacity (RSC), together with the effect on lipid peroxidation (LP). RSC was assessed by measuring the scavenging activity of garlic extracts on 2,2-diphenyl-1-picrylhydrazyl (DPPH) and hydrogen peroxide. Effects on LP were evaluated by following the activities of examined garlic extracts in Fe2+/ascorbate and Fe2+/H2O2 systems of induction. Investigated extracts reduced the DPPH radical formation (IC50 ranging from 1.03 to 6.01 mg/ml) and neutralised H2O2 (IC50 ranging from 0.55 to 2.01 mg/ml) in a dose-dependent manner. Strong inhibition of LP in both systems of induction was observed for all tested garlic extracts. Various levels of phenolics (0.05–0.98 mg gallic acid equivalents/g of dry extract) and flavonoid aglycones (4.16–6.99 μg quercetin equivalents/g of dry extract) in the investigated extracts of garlic could explain the obtained differences in these results only partially.
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Lignin is a group of complex phenolic polymers that provide important strengthening and waterproofing properties to plant cell walls. Considerable progress has been made in the study of the regulation of phenylpropanoid metabolism in relation to lignification. The prospect of altering plant lignins to improve intrinsic properties, such as biomass digestibility or pathogen resistance, by manipulating phenylpropanoid metabolism represents a rational approach toward plant improvement. The feasibility of artificially manipulating lignin and lignin-dependent processes is largely reliant on our understanding of the regulatory mechanisms controlling lignin biosynthesis. This chapter focuses on the enzymic steps of phenylpropanoid metabolism, namely the general phenylpropanoid and lignin branch pathways, describing attempts to manipulate these steps and alter lignin deposition and lignin-dependent processes. The prospect of attaining control over plant lignification via the manipulation of phenylpropanoid metabolism is addressed, and the potential problems and pitfalls relating to the multifunctionality of lignin, the complexity of the biosynthetic pathways, and the lack of specificity in the enzymes are examined. The problem of judging the significance of the controls that operate at the level of phenylpropanoid metabolism with those that operate at other levels, such as monolignol transport and the polymerization steps, is highlighted.
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Quercetin, a flavonoid, is found in many plants, including edible fruits and vegetables. We examined the effects on cell growth of human malignant cells derived from the gastrointestinal tract and on cell cycle progression. Quercetin markedly inhibited the growth of human gastric cancer cells and the IC50 value was 32–55 μM. DNA synthesis was suppressed to 14% of the control level by the treatment with 70 μM quercetin for 2 days. Furthermore, quercetin blocked cell progression from the G1 to the S phase.
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Substituted γ-chromones were found to weakly inhibit HIV-1 proteinase, an important enzyme in the replication and processing of the AIDS virus. Chromones bearing hydroxyl substituents and a phenolic group at the 2-position (flavones) were the most active compounds and structure-activity relationships for a limited series of flavone inhibitors are presented. Dixon plots are reported and a possible mechanism for flavone-induced inhibition is proposed. The results are also compared with those for some structurally related non-peptidic inhibitors of HIV-1 proteinase. Since some flavonoid compounds have already been shown to have antiviral activity against AIDS, the present observations of anti- HIV-1 proteinase activity may be particularly significant.
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Currently reliance on natural products is gaining popularity to combat various physiological threats including oxidative stress, cardiovascular complexities, cancer insurgence, and immune dysfunction. The use of traditional remedies may encounter more frequently due to an array of scientific evidence in their favor. Garlic (Allium sativum) holds a unique position in history and was recognized for its therapeutic potential. Recent advancements in the field of immunonutrition, physiology, and pharmacology further explored its importance as a functional food against various pathologies. Extensive research work has been carried out on the health promoting properties of garlic, often referred to its sulfur containing metabolites i.e. allicin and its derivatives. Garlic in its preparations are effective against health risks and even used as dietary supplements such as age garlic extract (AGE) and garlic oil etc. Its components/formulations can scavenge free radicals and protect membranes from damage and maintains cell integrity. It also provides cardiovascular protection mediated by lowering of cholesterol, blood pressure, anti-platelet activities, and thromboxane formation thus providing protection against atherosclerosis and associated disorders. Besides this, it possesses antimutagenic and antiproliferative properties that are interesting in chemopreventive interventions. Several mechanisms have been reviewed in this context like activation of detoxification phase-I and II enzymes, reactive oxygen species (ROS) generation, and reducing DNA damage etc. Garlic could be useful in preventing the suppression of immune response associated with increased risk of malignancy as it stimulates the proliferation of lymphocytes, macrophage phagocytosis, stimulates the release of interleukin-2, tumor necrosis factor-alpha and interferon-gamma, and enhances natural killer cells. In this paper much emphasis has been placed on garlic's ability to ameliorate oxidative stress, core role in cardiovascular cure, chemopreventive strategies, and indeed its prospective as immune booster.
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The cDNAs (Espals) encoding phenylalanine ammonia-lyase (PAL) were cloned from Ephedra sinica by reverse transcription polymerase chain reaction (RT-PCR) using degenerate primers and by 5' and 3'-rapid amplification of cDNA ends (RACE). 2166 bp of the open reading frame (ORF) encoded 722 amino acids; sequence analyses of Espal clones suggested that at least four isoforms of EsPAL (EsPAL1, 2, 3, 4) existed, with nine amino acids substitution in their sequences. Phylogenetic analysis of EsPAL and PALs from other plant species revealed that EsPAL and Pinus PAL formed a gymnosperm-type PAL subfamily. The recombinant EsPAL1 to 4 functionally catalyzed a PAL reaction and their K(m), V(max), K(cat) and K(cat)/K(m) values did not show significant differences. Semi-quantitative RT-PCR analysis indicated that the expression of Espal genes in the roots was higher than in the plant's aerial parts. In addition, the activity of PAL in the roots was also higher than in the aerial parts. These results suggest that Espal genes are expressed in the whole plant but are dominant in the roots rather than in the aerial parts.
Article
Phenylalanine ammonia-lyase and cinnamate 4-hydroxylase are important enzymes in allocating significant amounts of carbon from phenylalanine into the biosynthesis of several important secondary metabolites. Tea is an important crop of commerce known for its beverage and medicinally important flavonoid compounds, mainly catechins. As metabolic flux for the operation of the flavonoid pathway is maintained through the activities of PAL and C4H, thus, catechins biosynthesis in tea is critically dependent on the products of these enzymes. We examined the expression of PAL and C4H. Sequence encoding CsPAL was isolated from tea by polymerase chain reaction using sequence information available at the NCBI GenBank. Sequence encoding C4H was isolated from tea by using differential display of mRNA and rapid amplification of cDNA ends technology. CsC4H (AY641731) comprised of 1,352 bp full-length cDNA with open reading frame of 1,173 bp encoding 390 amino acids. Catechin contents decreased in response to drought stress (DS), abscisic acid (ABA), and gibberellic acid (GA(3)) treatments but increased in response to wounding. The expression of CsPAL and CsC4H showed the same behavior under the above treatments and was also in accordance with the catechin contents. A positive correlation between catechin contents and gene expression suggested a critical role of the enzymes in catechins biosynthesis and a crosstalk between phenylpropanoid and flavonoid pathways.
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To investigate the possible role of serine as a precursor of dehydroalanine at the active site of phenylalanine ammonia lyase, two serines, conserved in all known PAL and histidase sequences, were changed to alanine by site-directed mutagenesis. The resulting mutant genes were subcloned into the expression vector pT7.7 and the gene products were assayed for PAL activity. Mutant PALMutS209A showed the same catalytic property as wild-type PAL, whereas mutant PALMutS202A was devoid of catalytic activity, indicating that serine-202 is the most likely precursor of the active site dehydroalanine.
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In the past year progress has been made in the manipulation of phenylpropanoid metabolism but several studies highlight gaps in our understanding of the biochemistry of these pathways. New components involved in transcriptional regulation of phenylpropanoid genes have been identified, including transcription factors and novel proteins that function upstream of DNA-binding proteins.
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The objective of this review is to examine briefly the medical uses of garlic throughout the ages and the role that it was considered to play in prevention and treatment of disease. Interest in the potential benefits of garlic has origins in antiquity and is one of the earliest documented examples of plants employed for treatment of disease and maintenance of health. Garlic was in use at the beginning of recorded history and was found in Egyptian pyramids and ancient Greek temples. There are Biblical references to garlic. Ancient medical texts from Egypt, Greece, Rome, China and India each prescribed medical applications for garlic. In many cultures, garlic was administered to provide strength and increase work capacity for laborers. Hippocrates, the revered physician, prescribed garlic for a variety of conditions. Garlic was given to the original Olympic athletes in Greece, as perhaps one of the earliest "performance enhancing" agents. It is of interest that cultures that developed without contact with one another came to similar conclusions about the efficacy of garlic. Modern science is tending to confirm many of the beliefs of ancient cultures regarding garlic, defining mechanisms of action and exploring garlic's potential for disease prevention and treatment.
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An important traditional Chinese medicine herb, Astragalus membranaceus var. Mongholicus, whose dried root is known as Radix astragali ("Huangqi" in Chinese), has high flavonoid content as an essential active constituent. Phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) catalyzes the first and also a rate-limiting step in phenylpropanoid pathway, which supplies precursors for a variety of secondary metabolites including flavonoids. A PAL gene, designated AmPAL1 (GenBank accession no. AY986506), was isolated from A. membranaceus var. Mongholicus with a full-length cDNA of 2562 nucleotides and an open reading frame of 2154 bp. Northern blot analysis revealed that AmPAL1 expressed universally in different organs, and its expression was markedly induced by UV irradiation, mechanical wounding, and white light irradiation on etiolated seedlings, with some distinctive responsive properties. Content of a typical flavonoid, quercetin, in A. membranaceus var. Mongholicus of different ages correlated with PAL enzymatic activity. Transgenic tobacco plants harboring AmPAL1 under the control of the CaMV35S promoter showed significantly increased PAL activity and correlatively increased quercetin content than those in non-transformed plants. These results indicate that PAL is maybe a key point for flux into flavonoid biosynthesis in the genetic control of secondary metabolism in A. membranaceus var. Mongholicus.
Article
Garlic has been used medicinally since antiquity. In virtually every early civilization known, such as ancient India, Egypt, Rome, China, and Japan, garlic was part of the therapeutic regimen for a variety of maladies. Therefore, the ancient medicinal tradition of garlic use would qualify it as a folk medicine or as an alternative or complementary medicine. But is garlic an alternative to established methods of disease prevention or treatment? Scientists from around the world have identified a number of bioactive substances in garlic that are water soluble (e.g., S-allyl methylcysteine), and fat soluble (e.g., diallyldisulfide). Mechanisms of action are being elucidated by modern technology. The validity of ancient medicine is now being evaluated critically in cell-free systems, animal models, and human populations. Preventive and therapeutic trials of garlic are still in early stages. There are many promising lines of research suggesting the potential effects of garlic. The current state of knowledge does not recognize garlic as a true alternative, but it will likely find a place for garlic as a complement to established methods of disease prevention and treatment. Our goal should be to examine garlic together with other agents to evaluate its possible efficacy and toxicity under conditions of actual use in humans.
Article
The objective of this review is to update and assess the clinical evidence based on rigorous trials of the effectiveness of garlic (A. sativum). Systematic searches were carried out in Medline, Embase, Amed, the Cochrane Database of Systematic Reviews, Natural Standard, and the Natural Medicines Comprehensive Database (search date December 2006). Our own files, the bibliographies of relevant papers and the contents pages of all issues of the review journal FACT were searched for further studies. No language restrictions were imposed. To be included, trials were required to state that they were randomized and double blind. Systematic reviews and meta-analyses of garlic were included if based on the results of randomized, double-blind trials. The literature searches identified six relevant systematic reviews and meta-analysis and double-blind randomized trials (RCT) that were published subsequently. These relate to cancer, common cold, hypercholesterolemia, hypertension, peripheral arterial disease and pre-eclampsia. The evidence based on rigorous clinical trials of garlic is not convincing. For hypercholesterolemia, the reported effects are small and may therefore not be of clinical relevance. For reducing blood pressure, few studies are available and the reported effects are too small to be clinically meaningful. For all other conditions not enough data are available for clinical recommendations.
Article
Phenylalanine ammonia-lyase (PAL) is one of the branch point enzymes between primary and secondary metabolism. It plays an important role during plant development and defense. A PAL gene designated as SmPAL1 was cloned from Salvia miltiorrhiza using genome walking technology. The full-length SmPAL1 was 2,827 bp in size and consisted of an intron and two extrons encoding a 711-amino-acid polypeptide. Sequence alignment revealed that SmPAL1 shared more than 80% identity with the PAL sequences reported in Arabidopsis thaliana and other plants. The 5' flanking sequence of SmPAL1 was also cloned, and a group of putative cis-acting elements such as TATA box, CAAT box, G box and TC-rich repeats were identified. Transcription pattern analysis indicated that SmPAL1 expressed in all tissues examined, but more highly in leaf. Besides, expression of SmPAL1 was found to be induced by various treatments including ABA, wounding, and dehydration. To further confirm its function, SmPAL1 was expressed in Escherichia coli strain M15 with pQE-30 vector. The recombinant protein exhibited high PAL activity and could catalyze the conversion of L: -Phe to trans-cinnamic acid. This study will enable us to further understand the role SmPAL1 plays in the synthesis of active pharmaceutical compounds in S. miltiorrhiza at molecular level.
Flavones are inhibitors of HIV-1 proteinase Flavonoids are differentially taken up and transported long distances in Arabidopsis Implications of long-distance flavonoid movement in Arabidopsis thaliana
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The metabolism of aromatic compounds in higher Weisshaar, B.; Jenkins, G. I. Phenylpropanoid biosynthesis and its regulation
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Mimica-Dukic, N.; Samojlik, I.; Goran, A.; Igic, R. Phenolics as antioxidants in garlic (Allium sativum L., Alliaceae) Food Chem
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Quercetin and its glycosides in Allium vegetables
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Biological properties of garlic and garlic-derived organosulfur compounds
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Iciek, M.; Kwiecien, I.; Wodek, L. Biological properties of garlic and garlic-derived organosulfur compounds. Environ. Mol. Mutagen. 2009, 50 (3), 247-265.
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