Artemisinin Content Research Articles

Expression of artemisinin biosynthesis genes in eight Artemisia species at three developmental stages

In this study, the researchers have chosen eight species of Artemisia in order to find out their artemisinin content and also the expression levels of eight genes involved in the artemisinin biosynthetic pathway at three different developmental stages, i.e., vegetative, budding and flowering stages. Artemisinin was produced in all the species, albeit in various amounts (0.45–5.3mg/g DW). In four species (Artemisia absinthium, Artemisia diffusa, Artemisia sieberi and Artemisia spicigeria), maximum level of artemisinin production was observed at the flowering stage, while in the other three species (Artemisia annua,Artemisia campestris and Artemisia vulgaris) artemisinin production reached its maximum level at the budding stage. However, Artemisia scoparia is the only species showing highest artemisinin content at the vegetative stage that was correlated with high density of glandular trichomes in leaf tissue. The higher amount of artemisinin, produced in A. annua, was mainly a result of higher expression of the amorpha-4,11-diene synthase (ADS) and artemisinic aldehyde Δ11(13) reductase (DBR2) genes. While, for A. absinthium an increased expression of alcohol dehydrogenase 1 (ALDH1) gene along with decreased expression of dihydroartemisinic aldehyde reductase (RED1) gene resulted in a desirable increase in artemisinin yield. The results of the study indicate that there is a relationship between increased expression of some genes and enhancement of artemisinin content. However, in order to validate the relationship, further enzymatic analysis needs to be performed.

MUTATION INDUCTION FOR IMPROVING OF ARTEMISININ CONTENT IN EACH PART OF ARTEMISIA CINA MUTAN LINES

Mutation induction on Artemisia cina herbal medicine by gamma rays with the doses of 10, 20 and 40 Gy have been conducted at PAIR – BATAN. The purpose of mutation for improving of plant traits which were mutant lines more artemisinin content than control plant. The mutant lines interest were collected based on morphological characters (flowers, leaves, roots and stems). Artemisinin contents so far are only found in the leaves, the improvement are expected not only to increase artemisinin content in leaves but also in the roots, stems and flowers of A.cina mutant lines by mutation induction using gamma rays by the doses of 10, 20 and 40 Gy, every dose was selected 8 mutant lines for analysis artemisinin content.To obtain artemisinin from all selected mutant lines, flowers, leaves, roots and stems were extracted by using n-hexan, and then fractionation by ethyl acetate. Artemisinin content were analyzed by High Pressure Liquid Chromatography (HPLC), and pure artemisinin used as standard. Mutation induction by the dose of 40 Gy have been improving dry weight of roots from 38.85 to 76.19g, dry weight of leaves of mutant lines from 10 Gy irradiation was two times higher than leaves of parent plant, and three times by the dose of 40 Gy compared to dry weight of leaves parent plant, and no different by the dose of 20 Gy. The highest artemisinin content is 73.13mg/g in leaves of A26a3 mutant line and 36.68mg/g in flowers of A17.1 A.cina mutant lines, both of mutant lines from mutation induction by the dose of 10 Gy. Mutation induction by the dose of 20 Gy could improve artemisinin content in stems part of B12.1 mutant line from non detection in parent plant to 0.83mg/g. Mutation induction by the dose of 40 Gy improved artemisinin content to 1.90mg/g in roots and 1.50mg/g in stems of C27b1 and C8.3 mutant lines respectively. Artemisia cina un-irradiated as parent plant only contain artemisinin in flowers, leaves and roots were 0.13; 0.45 and 0.05mg/g respectively. Dose of 40 Gy is the best dose for enhancement of artemisinin content in each part of A.cina mutant lines. Key word : mutation, production of artemisinin, roots, stems, flowers

Antiprotozoal Effect of Artemisia indica Extracts and Essential Oil.

Diverse solvent extracts of Artemisia indica leaves originating from the West Bengal region (India) were assessed for the content of artemisinin and characteristic Artemisia polymethoxyflavonoids, namely eupatin (1), casticin (2), chrysoplenetin (3), cirsilineol (4), chrysophenol-D (5), and artemetin (6). HPLC-DAD and HPLC-MS were used to investigate the extracts macerated by solvents of increasing polarity, i.e., petroleum ether, n-hexane, dichloromethane, acetone, MeOH, or EtOH (either 96, 80, or 60 % v/v), and hot water. Artemisinin was absent in all extracts. The acetone and EtOH extracts comprised the highest levels of polymethoxyflavonoids, whereas no flavonoid could be detected in the infusion. None of the remaining extracts contained chryosphenol-D (5) or artemetin (6), while chrysoplenetin (3) was found in all extracts. The essential oil of the plant was also obtained by hydrodistillation and analysed by gas chromatography and gas chromatography-mass spectrometry simultaneously. Of the 92 compounds detected in the oil, camphor (13.0 %) and caryophyllene oxide (10.87 %) were the major components. All solvent extracts and the volatile oil showed in vitro antimalarial activity, plus a potential malaria prophylactic effect by inhibiting at least two recombinant plasmodial fatty acid biosynthesis (PfFAS-II) enzymes. Except for the infusion, all extracts were also active against other parasitic protozoa and displayed low cytotoxicity against mammalian cells. This is the first detailed study investigating both artemisinin and polymethoxyflavonoid content as well as in vitro malaria prophylactic and detailed antiprotozoal potential of A. indica extracts against a panel of protozoan parasites. This is also the first report of antiparasitic activity of the essential oil of the plant.

Effect of high light intensity on photoinhibition, oxyradicals and artemisinin content in Artemisia annua L.

Artemisia annua L. produces a compound called artemisinin that is a potent anti-malarial compound. However concentration of artemisinin within the plant is typically low (less than 0.8% of dry mass) and currently supply of the drug by the plant does not meet world demand. This investigation was carried out to determine whether high intensity light treatment would increase production of artemisinin in leaves of A. annua. Photoinhibition (14%) was induced in leaves of A. annua when they were subjected to 6 h of high-intensity light [2,000 μmol(photon) m-2 s−1]. Maximum photochemical efficiency of PSII showed a recovery of up to 95% within 24 h of light induced inhibition. During the light treatment, photochemical efficiency of PSII in leaves of the high-intensity light-treated plants was 38% lower than for those from leaves of plants subjected to a low-intensity-light treatment of 100 μmol(photon) m-2 s−1. Nonphotochemical quenching of excess excitation energy was 2.7 times higher for leaves treated with high-intensity light than for those irradiated with low-intensity light. Elevation in oxidative stress in irradiated leaves increased presence of reactive oxygen species (ROS) including singlet oxygen, superoxide anions, and hydrogen peroxide. Importantly, the concentration of artemisinin in leaves was two-fold higher for leaves treated with high-intensity light, as compared to those treated with low-intensity light. These results indicate that A. annua responds to high irradiance through nonphotochemical dissipation of light energy yet is subject to photoinhibitory loss of photosynthetic capacity. It can be concluded that A. annua is capable of rapid recovery from photoinhibition caused by high light intensity. High light intensity also induced oxidative stress characterized by increased concentration of ROS which enhanced the content of artemisinin. Such a light treatment may be useful for the purpose of increasing artemisinin content in A. annua prior to harvest.

Transplanting and harvesting effect on artemisinin biosynthesis and herb yield in Artemisia annua L.

This study evaluates the effect of differential transplantation (5th January, 28th January and 20th February; Ist T, IInd T and IIIrd T, respectively) and harvesting time (after 100 days of transplantation and 126–204 days of age) on artemisinin and herb yield in Artemisia annua in order to optimize field conditions. During the investigation, maximum plant height with branching intensity was observed in the third transplantation. Such a pattern of morphological appearance is classified under oval canopy, and such traits can be useful as visible morphotypic markers. Maximum plants with high artemisinin content were observed in the IInd transplantation. Interestingly, a wide range of artemisinin content (0.1–1 %) was observed in all three populations. Hence, to achieve maximum growth and high artemisinin yield within the shortest possible time, the second transplantation date is recommended for large-scale cultivation.

TRICHOME AND ARTEMISININ REGULATOR 1 Is Required for Trichome Development and Artemisinin Biosynthesis in Artemisia annua

Trichomes, small protrusions on the surface of many plant species, can produce and store various secondary metabolic products. Artemisinin, the most famous and potent medicine for malaria, is synthesized, stored, and secreted by Artemisia annua trichomes. However, the molecular basis regulating the biosynthesis of artemisinin and the development of trichomes in A. annua remains poorly understood. Here, we report that an AP2 transcription factor, TRICHOME AND ARTEMISININ REGULATOR 1 (TAR1), plays crucial roles in regulating the development of trichomes and the biosynthesis of artemisinin in A. annua. TAR1, which encodes a protein specially located in the nucleus, is mainly expressed in young leaves, flower buds, and some trichomes. In TAR1-RNAi lines, the morphology of trichomes and the composition of cuticular wax were altered, and the artemisinin content was dramatically reduced, which could be significantly increased by TAR1 oeverexpression. Expression levels of several key genes that are involved in artemisinin biosynthesis were altered when TAR1 was silenced or overexpressed. By the electrophoretic mobility shift, yeast one-hybrid and transient transformation β-glucuronidase assays, we showed that ADS and CYP71AV1, two key genes in the biosynthesis pathway of artemisinin, are likely the direct targets of TAR1. Taken together, our results indicate that TAR1 is a key component of the molecular network regulating trichome development and artemisinin biosynthesis in A. annua.

Genetic engineering of artemisinin biosynthesis: prospects to improve its production

Approaches involved in production of Artemisinin in different hosts. Artemisinin, a potent antimalarial natural compound, is obtained from aerial parts of Artemisia annua L. plants. The demand (101–119 MT) for artemisinin is exponentially increasing every year because of increased incidence of drug-resistant malaria throughout the world, especially in Africa and Asia. However, the presence of low concentrations (0.01–1.1 %) of the compound in A. annua L. plants poses a major constraint in the commercialization of artemisinin-based combination therapies (ACTs) recommended by WHO for the treatment of multidrug-resistant and cerebral malaria. Further, the improvement in the yield of artemisinin through conventional breeding, in vitro culture, cell suspension culture and total organic synthesis still poses a challenge. However, possibilities are there to enhance the artemisinin biosynthesis either by overexpression of the genes encoding enzymes associated with the rate-limiting steps of the mevalonate and artemisinin biosynthetic pathways or by the suppression of genes encoding enzymes of other competing pathways. Based on the current understanding of the pathway and cloning of the related genes, efforts have been made for the past few years to increase the artemisinin content in A. annua L., Cichorium intybus L. and microbes through metabolic engineering. In the present review, we have discussed the metabolic engineering strategies in both plant and microbial systems for artemisinin accumulation in bioengineered hosts.

The activity of the artemisinic aldehyde Δ11(13) reductase promoter is important for artemisinin yield in different chemotypes of Artemisia annua L.

The artemisinic aldehyde double bond reductase (DBR2) plays an important role in the biosynthesis of the antimalarial artemisinin in Artemisia annua. Artemisinic aldehyde is reduced into dihydroartemisinic aldehyde by DBR2. Artemisinic aldehyde can also be oxidized by amorpha-4,11-diene 12-hydroxylase and/or aldehyde dehydrogenase 1 to artemisinic acid, a precursor of arteannuin B. In order to better understand the effects of DBR2 expression on the flow of artemisinic aldehyde into either artemisinin or arteannuin B, we determined the content of dihydroartemisinic aldehyde, artemisinin, artemisinic acid and arteannuin B content of A. annua varieties sorted into two chemotypes. The high artemisinin producers (HAPs), which includes the '2/39', 'Chongqing' and 'Anamed' varieties, produce more artemisinin than arteannuin B; the low artemisinin producers (LAPs), which include the 'Meise', 'Iran#8', 'Iran#14', 'Iran#24' and 'Iran#47' varieties, produce more arteannuin B than artemisinin. Quantitative PCR showed that the relative expression of DBR2 was significantly higher in the HAP varieties. We cloned and sequenced the promoter of the DBR2 gene from varieties of both the LAP and the HAP groups. There were deletions/insertions in the region just upstream of the ATG start codon in the LAP varities, which might be the reason for the different promoter activities of the HAP and LAP varieties. The relevance of promoter variation, DBR2 expression levels and artemisinin biosynthesis capabilities are discussed and a selection method for HAP varieties with a DNA marker is suggested. Furthermore, putative cis-acting regulatory elements differ between the HAP and LAP varieties.

Effects of cobalt nanoparticles on artemisinin production and gene expression in Artemisia annua

The low artemisinin content in Artemisia has caused this compound to be expensive among medicines. Several attempts have been made to increase its production by altering the expression of different genes. However, no approach has been cost-effective. In this study, the expression levels of SQS and DBR2 genes were quantified by qRT-PCR, and artemisinin content was measured by HPLC in Artemisia annua cell suspension culture under nano cobalt particles elicitation. For this purpose, nano cobalt particles were used in 0.25, 2.5, and 5 mg L-1 concentrations and samples were collected after 8, 24, 48, and 72 h. The highest artemisinin content was observed 24 h after 5 mg L-1 nano cobalt treatment. In this case, artemisinin production was 2.25-fold (113.35 mg g-1 dw) higher than that of the control. Simultaneously, the expression levels of SQS and DBR2 genes decreased. It appears that the decrease in the expression of SQS and DBR2 genes caused the artemisinin content to increase by high concentrations of the nano cobalt particles.

Time-course analysis of the artemisinin biosynthesis, and expression of genes coding for artemisinin pathway enzymes, in response to exogenous salicylic and gibberellic acids

Artemisinin represents medicinally employed compound of Artemisia annua (L.) and is widely used against a num- ber of diseases. This study presents the effect of an exogenous application of salicylic acid (SA) and gibberellic acid (GA3) on the expression of genes coding for artemisinin pathway enzymes as well as on the biosynthesis of artemisinin. We found a temporary increase in artemisinin content in A. annua leaves 12 h after SA application. However, the transcript levels of all studied genes coding for artemisinin pathway enzymes decreased throughout the period studied and artemisinin content dropped down to an even lower level than in control plants at 168 h after treatment. In addition, artemisinin content increased, together with up-regulation of genes coding for en- zymes of artemisinin biosynthesis pathway, owing to GA3 application. However, down-regulation of all investi- gated genes was also observed at 72 h after treatment, probably due to the negative feedback control. As can be seen, both SA and GA3 exogenous applications can increase artemisinin accumulation in different ways. Because of a temporary increase in artemisinin content after SA or GA3 application, it is critical to determine the best harvesting time. Future research should focus on removing the negative feedback mechanism to achieve high- yield Artemisia plants.

Screening of Different <i>Artemisia</i> spp. from Western Ghats of Maharashtra for an Anti-Malarial Compound—Artemisinin

Artemisinin, an endoperoxide sesquiterpene lactone has proven effective in treating drug resistant cases of malaria and cancer. Artemisia annua [sweet wormwood] is the sole source for artemisinin production in many countries. To counter the low content in leaves and costly chemical synthesis process in India, alternative ways to produce artemisinin have been sought. In current study, we collected A. pallens, A. japonica and A. nilagirica from Western Ghats of Maharashtra, India and analyzed artemisinin content. Samples were extracted from leaves and florets in various extraction conditions and analyzed using different chromatographic techniques. Thin layer chromatography (TLC) and high performance thin layer chromatography (HPTLC) analysis showed the presence of compound with endoperoxide linkage in A. nilagirica. High performance liquid chromatography (HPLC) analysis showed the detection of artemisinin in methylene dichloride florets extract of A. japonica, but the concentration was too low [1.3 mg/g dry wt.] for further analyses. Gas chromatography/mass spectrometry (GC/MS) analysis identified structurally important components in the A. nilagirica ethyl acetate extract which explores the biosynthetic pathway of artemisinin from its most important precursor amorpha-4,11-diene. This is the first report of chromatographic screening of these Indian varieties of Artemisia spp. for artemisinin content.

Enhancement of artemisinin content and relative expression of genes of artemisinin biosynthesis in Artemisia annua by exogenous MeJA treatment

Methyl jasmonate (MeJA) is one of the most potent elicitors that can induce over accumulation of many natural products including artemisinin in plants. The 12 known genes (HMGR, DXS, DXR, HDS, HDR, FPS, ADS, CYP71AV1, DBR2, ALDH1, ORA and ERF1) of terpene metabolism in Artemisia annua were dynamically analyzed at the transcriptional levels in the treatment of MeJA from 0 to 48 h. HMGR (MVA pathway) showed higher expression level when the plants were treated with MeJA from 1 to 9 h and had the highest expression level at 3 h MeJA treatment. The expression levels of DXS and DXR (MEP pathway) reached the peak at 9 h. The last two genes of the MEP pathway, such as HDS and HDR, had the highest expression levels at 24 h. The expression of FPS increased significantly in the treatment of MeJA from 1 to 48 h, and the highest expression level appeared at 24 and 48 h after the MeJA treatment. Four genes in artemisinin-specific biosynthetic pathway including ADS, CYP71AV1, DBR2 and ALDH1 had higher expression levels in the treatment of MeJA from 1 to 48 h. The expression levels of two transcription factors such as ORA and ERF1 were also enhanced. The contents of artemisinin in the plants treated with MeJA for 24 and 48 h were respectively 0.971 and 0.973 mg/g DW, about 1.16-fold of the control (0.809 mg/g DW). Taken together, these results suggested that MeJA induced artemisinin biosynthesis by up-regulating the expression of the genes involved in artemisinin biosynthesis and the transcription factor.

Transcriptional changes in Artemisia annua L. induced by UV-B radiation

Artemisinin, the endoperoxide sesquiterpene lactone, is an effective antimalarial drug isolated from Artemisia annua L. In this study, A. annua plants were treated with low dose UV-B radiation to enhanced artemisinin content by nearly 30.5%. We investigated the gene expression responses of A. annua plants subjecte to UV-B radiation during 10-day period using a new custom made Agilent GeneChip with 43,692 probe sets. A total of 358 significantly differentially expressed genes, involved in signal transduction, antioxidant system and regulation of transcription were identified. The results indicated that the UV-B response was related to redox changes and synthesis of phytohormone abscisic acid (ABA). This study also provided a list of potential candidate genes regulating multiple pathways involved in the response. Acknowledgements: This study is supported by NNSF Project (No. 81273487), PAPD and SRF for ROCS (No. K513201011).

Extraction of Artemisinin, an Active Antimalarial Phytopharmaceutical from Dried Leaves of Artemisia annua L., Using Microwaves and a Validated HPTLC-Visible Method for Its Quantitative Determination

A simple, rapid, precise, and accurate high-performance thin-layer chromatographic method coupled with visible densitometric detection of artemisinin is developed and validated. Samples of the dried Artemisia annua leaves were extracted via microwaves using different solvents. This method shows the advantage of shorter extraction time of artemisinin from leaves under the influence of electromagnetic radiations. Results obtained from microwave-assisted extraction (MAE) were compared with hot soxhlet extraction. Chromatographic separation of artemisinin from plant extract was performed over silica gel 60 F254 HPTLC plate using n-hexane : ethyl acetate as mobile phase in the ratio of 75 : 25, v/v. The plate was developed at room temperature 25 ± 2.0°C. Artemisinin separation over thin-layer plate was visualized after postchromatographic derivatization with anisaldehyde-sulphuric acid reagent. HPTLC plate was scanned in a CAMAG’s TLC scanner 3 at 540 nm. Artemisinin responses were found to be linear over a range of 400–2800 ng spot−1 with a correlation coefficient 0.99754. Limits of detection and quantification were 40 and 80 ng spot−1, respectively. The HPTLC method was validated in terms of system suitability, precision, accuracy, sensitivity (LOD and LOQ), and robustness. Additionally, calculation of plate efficiency and flow constant were included as components of validation. Extracts prepared from different parts of the plant (leaves, branches, main stem, and roots) were analyzed for artemisinin content, in which, artemisinin content was found higher in the leaf extract with respect to branches and main stem extracts; however, no artemisinin was detected in root extract. The developed HPTLC-visible method of artemisinin determination will be very useful for pharmaceutical industries, which are involved in monitoring of artemisinin content during different growth stages (in vitro and in vivo) of A. annua for qualitative and quantitative assessment of final produce prior to commercial-scale processing for assessment of cost-benefit ratio.

Impact of Exogenous Elicitors on Artemisinin Production and Trichome Density in <i>Artemisia annua</i> L. under Subtropical Conditions

Artemisinin is a sesquiterpene lactone found in Artemisia annua having many medicinal properties. Therefore the factors affecting trichome initiation are important. In the present study, several strategies have been adapted to increase artemisinin production by treating them with nutrients, hormones and elicitors. Abscisic acid, Gibberellic acid (GA3), a combination of GA3+IAA (Gibberellic acid+indole acetic acid), Chitosan, Methyl jasmonate, Acetyl salicylic acid, Lead acetate, sodium chloride, and Nanozime were sprayed on plant at regular intervals. The glandular trichomes of leaves from Artemisia annua (Asteraceae) were examined by light and scanning electron microscopy and the percentage artemisnin content was analyzed by HPTLC. Maximum artemisinin content % and trichome index (0.128) was found when methy jasmonate was applied and minimum artemisinin content % and trichome index was reported in NaCl & acetyl salicylic acid.

Impact of Exogenous Elicitors on Artemisinin Production and Trichome Density in <i>Artemisia annua</i> L. under Subtropical Conditions

Artemisinin is a sesquiterpene lactone found in Artemisia annua having many medicinal properties. Therefore the factors affecting trichome initiation are important. In the present study, several strategies have been adapted to increase artemisinin production by treating them with nutrients, hormones and elicitors. Abscisic acid, Gibberellic acid (GA3), a combination of GA3+IAA (Gibberellic acid+indole acetic acid), Chitosan, Methyl jasmonate, Acetyl salicylic acid, Lead acetate, sodium chloride, and Nanozime were sprayed on plant at regular intervals. The glandular trichomes of leaves from Artemisia annua (Asteraceae) were examined by light and scanning electron microscopy and the percentage artemisnin content was analyzed by HPTLC. Maximum artemisinin content % and trichome index (0.128) was found when methy jasmonate was applied and minimum artemisinin content % and trichome index was reported in NaCl & acetyl salicylic acid.

Mutagenic effects of heavy-ion beam irradiation on in vitro nodal segments of Artemisia annua L.

Artemisia annua L. is a commercial source of artemisinin. Nevertheless, artemisinin content within the plant is relatively low and varies depending on genotype and environment. To broaden the genetic variability, the mutation effect of 12C-ion beam irradiation on A. annua was examined. Irradiation at 2.5 Gy had a slight lethal effect to nodal segments while a noticeable lethal effect was observed at 5 and 10 Gy. Furthermore, at higher doses (20 and 50 Gy), a severe lethal effect was observed. Mutations at the DNA level of axillary bud-derived shoots were performed by RAPD. The mutation frequency at 10 Gy was about 1.7 and 2.1 times higher than that at 2.5 and 5 Gy, respectively. After growth and artemisinin production observation of 72 irradiated mutants, around 14 and 7 % of them showed higher artemisinin content and artemisinin yield compared to the controls, respectively. The highest artemisinin content in a mutant was 1.43 % DW, which was 3.2-fold higher than the original wild type. Additionally, the highest artemisinin yield in mutants was 3.68 mg/plant, which was around 1.4-fold higher than in the wild type. Moreover, irradiated mutants exhibited antibacterial activity against S. aureus, but the wild types did not. This study presents an effective application of heavy ion beam irradiation to create variations and improve artemisinin production in A. annua.

The physiological response of Artemisia annua L. to salt stress and salicylic acid treatment.

Salinity has a great influence on plant growth and distribution. A few existing reports on Artemisia annua L. response to salinity are concentrated on plant growth and artemisinin content; the physiological response and salt damage mitigation are yet to be understood. In this study, the physiological response of varying salt stresses (50, 100, 200, 300, or 400mM NaCl) on A. annua L. and the effect of exogenous salicylic acid (0.05 or 0.1mM) at 300-mM salt stress were investigated. Plant growth, antioxidant enzyme activity, proline, and mineral element level were determined. In general, increasing salt concentration significantly reduced plant growth. Superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) were stimulated by salt treatment to a higher enzyme activity in treated plants than those in untreated plants. Content of proline had a visible range of increment in the salt-treated plants. Distribution of mineral elements was in inconformity: Na(+) and Ca(2+) were mainly accumulated in the roots; K(+) and Mg(2+) were concentrated in leaves and stems, respectively. Alleviation of growth arrest was observed with exogenous applications of salicylic acid (SA) under salt stress conditions. The activity of SOD and POD was notably enhanced by SA, but the CAT action was suppressed. While exogenous SA had no discernible effect on proline content, it effectively inhibited excessive Na(+) absorption and promoted Mg(2+) absorption. Ca(2+) and K(+) contents showed a slight reduction when supplemented with SA. Overall, the positive effect of SA towards resistance to the salinity of A. annua will provide some practical basis for A. annua cultivation.

Breeding strategy for genetic improvement up to four generations in relation to artemisinin with canopy and other secondary metabolites in Artemisia annua L

This study represents a classical breeding approach for evaluating and systematically selecting high yielding and specific canopy holding plants through the expression of enhanced secondary metabolite biosynthesizing capability for the pharmaceutically important phyto-molecules-Artemisinin. During the 4 years of study (C0-C4 cycle) it was observed that, plant height ranging from about 100cm to more than 300cm, and the plants touching around 300cm height did fall in the three distinct canopy types i.e., pyramidal, spreading and oval in a population. In C0 cycle artemisinin content was in the range of 0.1-0.2% and artemisinic acid (AA) was high (above 0.03%). Gradually due to high selection pressure in favour of artemisinin content, plants containing high artemisinin content showed low AA (less than 0.01%) in C3 cycle. It was also observed that essential oil did not have any significant relationship with artemisinin content. In C0 cycle, oil content was medium (0.2–0.59%) or low (0.0–0.19%) along with artemisinin content. There was a clear-cut shift from three predominant distinct canopies towards oval and spreading in whole selection procedure and then ultimately going in the favour of the oval canopy type for high artemisinin concentration. In a classical breeding fashion, on the principle of recurrent mass selection and progeny testing, the genetic improvement of population through gene pool exploitation using restricted mating among selected progenies of chemotypically superior plant and high yielding genotype (variety ‘CIM Arogya’) was developed.

English

Artemisia annua L. is an important medicinal plant from which artemisinin was extracted to treat malaria effectively. Artemisinin is isolated from the aerial part of the A. annua harvesting from the wild is causing loss of genetic diversity and habitat destruction. The use of controlled environments can overcome cultivation difficulties and could be a means to manipulate phenotypic variation in bioactive compounds and toxins. The aim of this review is to bring together most of the available scientific research papers about the cultivation and breeding conducted on the genus A. annua, which is currently scattered across various publications. Through this review the authors hope to attract the attention of all agronomist and breeders throughout the world to focus on the unexplored potential of A. annua species. Also, the future scope of this plant has been emphasized with a view of the importance of cultivation of A. annua for increasing of artemisinin content. New cultivation approach of A. annua offers the opportunity to optimize yield and achieve a uniform, high quality product. Key words: Artemisia annua, breeding, artemisinin, cultivation, medicinal plant.