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An efficient method of genomic DNA extraction that provides high quality and yield is a crucial pre-requisite and limiting factor in plant genetic analysis. However, pure genomic DNA can be challenging to obtain from some plant species due to their sugar and secondary metabolite contents. Lippia alba is an important aromatic and medicinal plant, chemically characterized by the presence of tannins, flavonoids, anthocyanins, and essential oils, which interfere with the extraction of pure genomic DNA. In this scenario, optimizing the extraction methods and minimizing the effects of these compounds are necessary. This study compares six plant DNA extraction protocols based on the CTAB method. The quality and quantity of DNA samples obtained were determined by physical appearance by electrophoresis in agarose gels and spectrophotometry. The results highlight the difficulty in obtaining pure and clear bands for all tested methods, except for the polyvinylpyrrolidone (PVP)-based protocol created by our team, which was the better option for obtaining high-quality genomic DNA of L. alba. We conclude that adding PVP-40 into DNA extraction buffers can optimize the DNA extraction of L. alba and indicate this protocol for DNA extraction from other aromatic plants., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Lippia dulcis (Aztec sweet herb) contains the potent natural sweetener hernandulcin, a sesquiterpene ketone found in the leaves and flowers. Utilizing the leaves for agricultural application is challenging due to the presence of the bitter-tasting and toxic monoterpene, camphor. To unlock the commercial potential of L. dulcis leaves, the first step of camphor biosynthesis by a bornyl diphosphate synthase needs to be elucidated. Two putative monoterpene synthases (LdTPS3 and LdTPS9) were isolated from L. dulcis leaf cDNA. To elucidate their catalytic functions, E. coli-produced recombinant enzymes with truncations of their chloroplast transit peptides were assayed with geranyl diphosphate (GPP). In vitro enzyme assays showed that LdTPS3 encodes bornyl diphosphate synthase (thus named LdBPPS) while LdTPS9 encodes linalool synthase. Interestingly, the N-terminus of LdBPPS possesses two arginine-rich (RRX 8 W) motifs, and enzyme assays showed that the presence of both RRX 8 W motifs completely inhibits the catalytic activity of LdBPPS. Only after the removal of the putative chloroplast transit peptide and the first RRX 8 W, LdBPPS could react with GPP to produce bornyl diphosphate. LdBPPS is distantly related to the known bornyl diphosphate synthase from sage in a phylogenetic analysis, indicating a converged evolution of camphor biosynthesis in sage and L. dulcis. The discovery of LdBPPS opens up the possibility of engineering L. dulcis to remove the undesirable product, camphor., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The aerial parts of Lippia integrifolia (incayuyo) are widely used in northwestern and central Argentina for their medicinal and aromatic properties. The essential oil composition of thirty-one wild populations of L. integrifolia covering most of its natural range was analyzed by GC and GC-MS. A total of one hundred and fifty two terpenoids were identified in the essential oils. Sesquiterpenoids were the dominant components in all but one of the collections analyzed, the only exception being a sample collected in San Juan province where monoterpenoids amounted to 51%. Five clearly defined chemotypes were observed. One possessed an exquisite and delicate sweet aroma with trans-davanone as dominant component (usually above 80%). Another with an exotic floral odour was rich in oxygenated sesquiterpenoids based on the rare lippifoliane and africanane skeletons. The trans-davanone chemotype is the first report of an essential oil containing that sesquiterpene ketone as the main constituent. The absolute configuration of trans-davanone from L. integrifolia was established as 6S, 7S, 10S, the enantiomer of trans-davanone from 'davana oil' (Artemisia pallens). Wild plants belonging to trans-davanone and lippifolienone chemotypes were propagated and cultivated in the same parcel of land in Santa Maria, Catamarca. The essential oil compositions of the cultivated plants were essentially identical to the original plants in the wild, indicating that the essential oil composition is largely under genetic control. Specimens collected near the Bolivian border that initially were identified as L. boliviana Rusby yielded an essential oil practically identical to the trans-davanone chemotype of L. integrifolia supporting the recent view that L. integrifolia (Gris.) Hieron. and L. boliviana Rusby are synonymous., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

The goal of the present study was to evaluate the acaricidal potential of Lippia alba essential oil, citral chemotypes (LA-10 and LA-44 genotypes) and carvone chemotypes (LA-13 and LA-57 genotypes), as well as purified citral and enantiomers of carvone and limonene. Efficacy against Rhipicephalus microplus was assessed by the larval packet and the engorged female immersion tests. Citral chemotypes had greater larvicidal activity than carvone chemotypes, and this was further supported by larvicidal and adulticidal activity of purified citral with LC50 values of 7.0 and 29.8 mg/mL, respectively. While purified enantiomers of carvone exhibited greater larvicidal activity than those of limonene, enantioselectivity of limonene was observed with R-(+) displaying significantly higher efficacy (LC50 of 31.2mg/mL) than S-(-) (LC50 of 54.5mg/mL). The essential oils and purified compounds were much less toxic toward engorged adult females, with the exception of citral, and this may be due to limited cuticular penetration., (Published by Elsevier B.V.)

Lippia gracilis, popularly known in Brazil as 'alecrim-de-tabuleiro', is used for many purposes, especially antimicrobial and antiseptic activities. The leaves of three L. gracilis genotypes, including LGRA-106, LGRA-109 and LGRA-110 were collected from the Active Germplasm Bank located in the "Campus Rural da UFS" research farm at the São Cristóvão country, Sergipe State, Brazil. The essential oils were obtained from leaves of L. gracilis plants by hydrodistillation. Chemical analysis of the essential oils was performed by gas chromatography-mass spectrometry (GC-MS). The susceptibility of Trichophyton rubrum strains, MYA3108 and TruMDR2, to the two L. gracilis genotypes (LGRA-106 and LGRA-109) essential oils was determined by the serial microdilution method. Leishmanicidal activity of essential oil from LGRA-106 and LGRA-110 was assayed by tetrazolium-dye (MTT) colorimetric method. The oxygenated monoterpene thymol was the main component of the essential oil from genotype LGRA-106, while Carvacrol was more abundant in LGRA-109 and LGRA-110. The concentrations of LGRA-106 and LGRA-109 essential oils that completely eliminate the fungi were determined and these concentrations were similar to those observed for fluconazole, a common antifungal drug. Among the genotype tested, LGRA-106 essential oil exhibited the best fungicidal activity at 46.87μgmL(-1). Regarding to leishmanicidal activity, the IC50, for LGRA-106 and LGRA-110, was 86.32 and 77.26μgmL(-1), respectively. The results showed that L. gracilis essential oil, rich in thymol and thymol itself presented best antidermatophytic activity, while the best leishmanicidal activity was obtained with essential oil from genotype rich in Carvacrol and Carvacrol itself., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The present study aimed to evaluate the activity of Lippia gracilis Schauer essential oil obtained from different L. gracilis genotypes and their major components, carvacrol and thymol against Rhipicephalus (Boophilus) microplus (cattle tick) larvae and engorged females. The larval test was performed parallel to the adult immersion test for engorged females for four L. gracilis genotypes. Similar tests were further performed for their major compounds carvacrol and thymol. Carvacrol (LC50 of 0.22 and 4.46 mg/mL, to larvae and engorged females, respectively) was more efficient than thymol (LC50 of 3.86 and 5.50 mg/mL, to larvae and engorged females, respectively). The lethal concentrations obtained for the isolated essential oil from genotypes LGRA-201 against larvae (1.31 mg/mL) and LGRA-106 against engorged females (4.66 mg/mL) confirmed the acaricidal activity of L. gracilis essential oil and its effectiveness in controlling the southern cattle tick., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Hernandulcin, a C15 sesquiterpene ketone, is a natural sweetener isolated from the leaves of Lippia dulcis. It is a promising sugar substitute due to its safety and low caloric potential. However, the biosynthesis of hernandulcin in L. dulcis remains unknown. The first biochemical step of hernandulcin is the synthesis of (+)-epi-α-bisabolol from farnesyl diphosphate, which is presumed to be catalyzed by a unique sesquiterpene synthase in L. dulcis. In order to decipher hernandulcin biosynthesis, deep transcript sequencings (454 and Illumina) were performed, which facilitated the molecular cloning of five new sesquiterpene synthase cDNAs from L. dulcis. In vivo activity evaluation of these cDNAs in yeast identified them as the sesquiterpene synthases for α-copaene/δ-cadinene, bicyclogermacrene, β-caryophyllene, trans-α-bergamotene, and α-bisabolol. The engineered yeast could synthesize a significant amount (~0.3 mg per mL) of α-bisabolol in shake-flask cultivation. This efficient in vivo production was congruent with the competent kinetic properties of recombinant α-bisabolol synthase (K(m) 4.8 μM and k(cat) 0.04 s(-1)). Detailed chemical analyses of the biosynthesized α-bisabolol confirmed its configuration to be (+)-epi-α-bisabolol, the core skeleton of hernandulcin. These results demonstrated that enzymatic, stereoselective synthesis of (+)-epi-α-bisabolol can be achieved, promising the heterologous production of a natural sweetener, hernandulcin., (Copyright © 2012 Elsevier Inc. All rights reserved.)