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Objective: To analyze the DNA methylations between two chemical types of Pogostemon cablin using MSAP technique. Methods: Methylation sensitive amplification polymorphism (MSAP) technique was used to detect the DNA methylation degree of two chemical type and a total of 197 P. cablin samples. Results: The highest or higher values of the Shannon polymorphism index of the four types of MSAP locus information were produced in the five areas of Yangchun, Deqing, Gaozhou Datong, Lubu, and Guangning. It formed two branches with P. cablin Shipaiensis (Pogostone-type) and the rest (Patchouliol-type). The percentage of variation among the populations was 60.66%, which was far greater than that within the population. A total of 10 differential fragments were screened out, sequenced and analyzed, one of those belongs to the serine/threonine protein kinase family. Conclusion: The MSAP technique can be used to identify the different origins of P. cablin. The formation of different chemical types of P. cablinis closely related to its DNA methylation level, and further research is needed.
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Objective To find out evidences for the identification of Ajuga decumbens Thunb.(Jinchuangxiaocao) and Pogostemon cablin (Blanco.) Benth.(Guanghuoxiang) . Methods Fresh stems and leaves of the two species were collected and used as the samples. We used stereoscopy to observe and compare the macroscopic appearance of the original plants, and used light microscope to observe the stem cross-section, petiole cross-section, leaf epiderm and powder of the two species for microscopic identification. Results The results of macroscopic character identification showed that the petiole of Jinchuangxiaocao had narrow wing, while that of Guanghuoxiang had no narrow wing; the angles between leaf main vein and lateral vein of Jinchuangxiaocao were less than 45°, while the angles of Guanghuoxiang were more than 45°. In respect of microscopic identification, Jinchuangxiaocao had the glandular scales with multicellular head, and Guanghuoxiang had the glandular scales with one-celled head; the intercellular space and crystals were found in the cortex of Guanghuoxiang stems and mesophyll, while Jinchuangxiaocao has no such structures. Jinchuangxiaocao had diacytic and anomocytic stomata, and Guanghuoxiang only had diacytic stomata. Conclusion The appearance and microscopic differences can be used for the identification of Jinchuangxiaocao from Guanghuoxiang.
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Objective: To study the effects of exogenous salicylic acid (SA) on the seed germination and seedling growth of Pogostemon cablin under salt stress and the optimal concentration of SA for alleviating the salt stress. Methods: Under salt stress simulated by 50 mmol/L NaCl solution, P. cablin was used to investigate the effects of exogenous SA on the seed germination and seedling growth with paper media. The germination energy (GE), germination rate (GR), germination index (GI), vigor index (VI), seedling length (SL), root length (RL), seedling fresh weight (SFW), seedling dry weight (SDW), and contents of soluble protein (SP) and chlorophyll in the leaves were determined, and the relative salt-damage rate of these indexes was analyzed. Results: The seed germination and seedling growth of P. cablin were significantly inhibited under 50 mmol/L NaCl stress. Compared with NaCl stress, the seed germination and seedling growth were improved when SA were 0.10 and 0.25 mmol/L, while reduced when SA was 0.50 mmol/L. GE, GR, GI, VI, SL, RL, SFW, SDW, and contents of SP and chlorophyll were significantly improved by 46.02%, 27.45%, 30.16%, 92.63%, 39.80%, 47.94%, 26.48%, 18.85%, 15.94%, and 14.70% with 0.25 mmol/L SA, as well as MDA content was significantly reduced by 18.35%. Conclusion: SA could improve the ability of the seed germination and seedling growth of P. cablin to adapt the environment and alleviate the inhibitory effects of salt stress. Among all the treatments, the optimal concentration of SA is 0.25 mmol/L.
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Objective: In order to elucidate the carotenoid metabolic pathway in Pogostemon cablin, a phytoene desaturase (PDS) of P. cablin (PcPDS1) gene was cloned and analyzed bioinformatically. Methods: The full length PDS gene sequence was retrieved from transcriptomic database of P. cablin, the full length primer was designed for PCR verification, and PcPDS1 gene was analyzed using several bioinformatical softwares. Results: PcPDS1 (NCBI accession, KC854409) contained 1960 bp length of open reading frame (ORF) and encoded 569 amino acids postulated. Additionally, the physical and chemical properties of PcPDS1-coded protein were analyzed by bioinformatical softwares. Phylogenetic analysis showed that PcPDS1 was tightly clustered with PDS genes in Gentiana lutea, Nicotiana tabacum, and Ipomoea batatas, which is consistent to the phylogenetic relationship of species. Conclusion: PcPDS1 is successfully coloned and molecularly characterized.
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OBJECTIVE: To develop a method for the determination of ethanol extracts content of Pogostemon cablin (Blanco) Benth. using near-infrared spectroscopy. METHODS: The ethanol extracts contents of 139 samples were determined by the method of ethanol cold-soak extraction. Near-infrared spectrograms were obtained, then multiplicative signal correction, first derivative method and Savitzky-Golay filter method were used to process the spectrograms, at last the calibration model of ethanol extracts content was established by partial least squares regression analysis method. RESULTS: The correlation coefficients, root-mean-square error of calibration, root-mean-square error of prediction and root-mean-square error of cross-validation of the calibration model of ethanol extracts content were 0.98958, 0.590, 0.623 and 0.97518, respectively. CONCLUSION: The method is stable, accurate, and can be used to predict ethanol extracts content of Pogostemon cablin (Blanco) Benth. rapidly.
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Objective: To isolate the pathogenic Ralstonia solanacearum from the infected Pogostemon cablin plants and to study the bacteria characteristics, so as to lay a foundation for the control of bacterial wilt. Methods: Pathogenic strains of R. solanacearum were isolated from the infected P. cablin plants by being streaked on a tetrazolium chloride (TZC) medium. These strains were used for the classification based on oxidative utilization of three kinds of disaccharide (lactose, maltose, and cellobiose) and three kinds of hexy I alcohol (mannitol, dulcitol, and sorbitol). The pathogenicity was tested by the methods of cutting and soaking roots. Results: Seven strains of pathogenic R. solanacearum were isolated from the infected P. cablin plants which contain four biotypes. Strains of HX5 and HX7 belong to biotype I, HX1 and HX6 belong to biotype II, HX2 and HX4 belong to biotype III, and HX3 belongs to biotype V. The result of pathogenicity test indicated that the most strains had strong pathogenicity. HX4 and HX5 have more serious virulence and can cause higher plant death rate. Conclusion: Strains of R. solanacearum with different biotypes from the P. cablin plants have different pathogenicity.
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[Objective] To compare the difference of fingerprints of Pogostemon cablin (Blanco) Benth. from different places of origin. [Methods] The chemical components of Pogostemon cablin (Blanco) Benth. from 6 places of origin were detected by thin-layer chromatography. [ Results ] The thin-layer chromatogram of volatile components of Pogostemon cablin (Blanco) Benth. from different places of origin was similar. The contents of methanol-extract and petroleum-ether-extract from Pogostemon cablin (Blanco) Benth. planted in Gaoyao, Guangdong differed greatly from those in other places of origin, which can be used to identify the herb of Pogostemon cablin ( Blanco) Benth. from Gaoyao, Guangdong. [Conclusion] Thin-layer chromatographic fingerprints can be used as the parameter to differentiate Pogostemon cablin (Blanco) Benth. from different places of origin.
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Objective Callus induction and plantlets regeneration of Pogostemon cablin(Blanco)Benth.are studied to establish a high-efficiency regeneration system for tissue culture.Methods The leaf segments,nodular stem segments,stem segments and root tips from Pogostemon cablin(Blanco)Benth.were cultured in vitro.The factors which affect the induction,multiplication and differentiation of callus were studied.Results Benzylaminopurine(BA) or thidiazuron(TDZ) combined with ?-naphthylacetic acid(NAA) was helpful to induce callus.The optimal concentrations of BA were 0.2～0.5mg/L.Nodular stem segments and stem segments were the most suitable explants for callus induction.The medium containing BA and NAA promoted the multiplication of callus.The multiplication rate and differentiation rate of 30-day callus were much higher than those of 60-day callus.BA was helpful to induce callus differentiation.The callus from leaf segments exerted a higher differentiation rate.Conclusion A high-efficiency regeneration system of callus induction and plant regeneration from Pogostemon cablin(Blanco)Benth.has been established.
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Object To study comparatively the fingerprint of Pogostemon cablin (Blanco) Benth. in different producing area and investigate the effect of pyrolysis temperature and pyrolysis time on the fingerprint Methods Pyrolysis gas chromatography (PGC) and fuzzy cluster analysis were used Results Both similarity and charecteristics were found in the fingerprint of P. cablin in different producing area. The methodological evalulation showed that this method had a good reproducibility, RSD≤2.75% Conclusion This method is simple, rapid and accurate. It is suitable for the quality control of the Chinese materia medica with the important role of pyrolysis temperature and pyrolysis time
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Objective The volatile oil was further purified to get the patchouli alcohol by molecular distillation taking patchouli oil which contained 23.97% of patchouli alcohol as raw material.Methods The best process parameters,including temperature,feed rate,and scraping rate,of molecular distillation technique for separating patchouli oil were studied by orthogonal test.The content was detected by GC under the conditions of elastic quartz capillary column HP-5,detector temperature 280 ℃,gasification temperature 280 ℃,distributary refer 20∶1.The effects of evaporation temperature on content and extraction rate of the patchouli oil had been investigated and the difference between the molecular and vapor distillations was studied.Results Content and extraction rate of patchouli alcohol under the best evaporation temperature 65 ℃ by molecular distillation reached to 40.71% and 76.55%,respectively.Conclusion The molecular distillation technique that could get the product with higher purity and extracting rate than that by vapor distillation in patchouli oil purification has a very good prospect in industry.
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Objective To study the synthesizing site,transporting mode,and storing position of essential oil in Pogostemon cablin stems and leaves.Methods Ultrathin slicing and histochemical methods were used.Results In parenchyma cells in the stem of P.cablin,dark osmiophilic material was observed in plastids and mitochondria,otherwise,some small gray vesicles were also found in cytoplasm.Some dark osmiophilic droplets were found to adhere to outer membrane of small gray vesicles,and many small gray vesicles with osmiophilic droplets gathered in outer membrane of large vacuole.In leaves,it was different case.Because palisade tissue cells were arranged closely,which had smaller volume and rich chloroplasts,so there was not large vacuole in palisade tissue cells.Plastids,mitochondria,and small vacuole were distributed in high density,but the first two organelles contained more dark osmilphilic substance than latter.Conclusion The plastid is the main synthesis site of essential oil,and the essential oil was mainly transferred by small gray vesicles into large vacuole to store in the stem.In leaves,chloroplast not only has photosynthesis,but also is the main synthesis site of essential oil,and essential oil is permeated into small vacuole to store.
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Objective To study the stem development and essential oil distribution in different deve- lopment stages of Pogostemon cablin and to provide theoretical information and practical guidance for reasonable harvest and use of this medicinal plant. Methods Paraffin method, semi-thin sectioning, and histochemical method were used. Results Essential oil was not found in the promeristem of stem; in the primary meristem of stem and the early stage of stem's primary structure, the essential oil was mainly distributed in glandular hairs; in the primary structure of stem, the essential oil was mainly distributed in cortex parenchyma cells; in the secondary structure of stem, the essential oil was mainly distributed in remaining cortex parenchyma cell. Conclusion The essential oil is mainly distributed in the stem bark, so the thickness of a stem bark can be used as a standard of selective breeding index.
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Objective: To establish the method of fingerprint analysis on volatile oil of Pogostemon cablin (Blanco) Benth. cultivated in Shipai country by GC MS, and to study the main characteristic components. Methods: The main components of the volatile oil of Pogostemon cablin (Blanco) Benth. from different colletion time in Shipai country were determined by GC MS, and compared with the volatile oil of “Gaoyao huoxiang” and “Hainan huoxiang” cultivated in the same GAP plots. Results: RSD of precision and reproducibility and stability is in the range of 5%, and the main components of the volatile oil of “Shipai huoxiang” from different collection time are nearly similar, and the content of Pogostone increases along with cultivation time, and the content of Pogostone and Patchoulic alcohol are significantly differences compared with the “Gaoyao huoxiang” and “Hainan huoxiang”. Conclusion: The method is reliable, accurate and can be used for quality control of volatile oil of Pogostemon cablin (Blanco) Benth.
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Objective: To establish the characteristic fingerprint of volatile oil of Pogostemon cablin (Blanco) Benth.Methods: The main components of 21 samples were determined by GC MS. Results: ? Patchoulene, ? Elemene, cis Caryophyllene, trans Caryophyllene, seychellene, ? Guaiene, ? Patchoulene, ? Guaiene, unidentification component A, patchoulic alcohol, pogostone were determined in all samples, and the relative content of 11 components add up to 80 percent. Compared with the samples produceds in Gaoyao and Hainan, Shipai samples were significantly differences.Conclusion: The retention time and relative content of 11 components bulidup the characteristic fingerprint of volatile oil of Pogostemon cablin (Blanco) Benth, it can be used for quality control and identification to Pogostemon cablin (Blanco) Benth.