Estimation for Raw Material Plants of a Henna Product Using LC-High Resolution MS and Multivariate Analysis.

Henna is a plant-based dye obtained from the powdered leaf of the pigmented plant Lawsonia inermis, and has often been used for grey hair dyeing, treatment, and body painting. As a henna product, the leaves of Indigofera tinctoria and Cassia auriculata can be blended to produce different colour variations. Although allergy from henna products attributed to p-phenylenediamine, which is added to enhance the dye, is reported occasionally, raw material plants of henna products could also contribute to the allergy. In this study, we reported that raw material plants of commercial henna products distributed in Japan can be estimated by LC-high resolution MS (LC-HRMS) and multivariate analysis. Principal Component Analysis (PCA) score plot clearly separated 17 samples into three groups [I; henna, II; blended henna primarily comprising Indigofera tinctoria, III; Cassia auriculata]. This grouping was consistent with the ingredient lists of products except that one sample listed as henna was classified as Group III, indicating that its ingredient label may differ from the actual formulation. The ingredients characteristic to Groups I, II, and III by PCA were lawsone (1), indirubin (2), and rutin (3), respectively, which were reported to be contained in each plant as ingredients. Therefore, henna products can be considered to have been manufactured from these plants. This study is the first to estimate raw material plants used in commercial plant-based dye by LC-HRMS and multivariate analysis.

Activation and Identification of a Griseusin Cluster in Streptomyces sp. CA-256286 by Employing Transcriptional Regulators and Multi-Omics Methods.

Streptomyces are well-known producers of a range of different secondary metabolites, including antibiotics and other bioactive compounds. Recently, it has been demonstrated that "silent" biosynthetic gene clusters (BGCs) can be activated by heterologously expressing transcriptional regulators from other BGCs. Here, we have activated a silent BGC in Streptomyces sp. CA-256286 by overexpression of a set of SARP family transcriptional regulators. The structure of the produced compound was elucidated by NMR and found to be an N-acetyl cysteine adduct of the pyranonaphtoquinone polyketide 3'-O-α-d-forosaminyl-(+)-griseusin A. Employing a combination of multi-omics and metabolic engineering techniques, we identified the responsible BGC. These methods include genome mining, proteomics and transcriptomics analyses, in combination with CRISPR induced gene inactivations and expression of the BGC in a heterologous host strain. This work demonstrates an easy-to-implement workflow of how silent BGCs can be activated, followed by the identification and characterization of the produced compound, the responsible BGC, and hints of its biosynthetic pathway.

In Vitro: Cytotoxicity, Apoptosis and Ameliorative Potential of Lawsonia inermis Extract in Human Lung, Colon and Liver Cancer Cell Line.

Cancer has emerged as a major public health issue in developed as well as in developing countries. Plant-derived molecules are widely being used in the treatment of cancer due to their minimum side effects. Lawsonia inermis (Henna) is one of the medicinal plants containing many therapeutic properties. In the present study, bioactive components of L. inermis extract were analyzed by LCMS/MS method and validated. Lawsone (3.5%) is primarily responsible for cytotoxic and anti-cancerous activities. These properties were studied on human lung carcinoma (A549), colorectal cancer (DLD1) and Hepatocellular carcinoma (HepG2) cancer cell lines. The activities were assessed by MTT assay, evaluation of apoptosis by measuring the production of Reactive Oxygen Species (ROS) and mitochondrial membrane potential of the cancer cell lines. Moreover, apoptosis in the respective cancer cell lines was also determined by chromatin condensation and DNA fragmentation using Hoechst 33528 and propidium iodide (PI) staining. The preliminary in vitro result of MTT showed that the henna extract induces cytotoxic properties against A549, DLD1, HepG2 with IC50values 490, 480 and 610 µg/ml respectively (more than 40% growth inhibition). In addition, the extract induced a concentration-dependent rise in ROS production which was 84, 102, and 110% in HepG2, DLD1 AND A549 respectively at 300 µg/ml, whereas at 400 µg/ml concentration it was 86, 102, and 106% in respective cell lines while decreasing mitochondrial membrane potential was more than 20% in the investigated cell lines. The extract also provoked changes associated with apoptosis and the data indicate that the ROS production leads to a diminution in mitochondrial membrane potential and this correlated with the extract cytotoxicity.

Metabolomics Analysis Reveals Tissue-Specific Metabolite Compositions in Leaf Blade and Traps of Carnivorous Nepenthes Plants.

Nepenthes is a genus of carnivorous plants that evolved a pitfall trap, the pitcher, to catch and digest insect prey to obtain additional nutrients. Each pitcher is part of the whole leaf, together with a leaf blade. These two completely different parts of the same organ were studied separately in a non-targeted metabolomics approach in Nepenthes x ventrata, a robust natural hybrid. The first aim was the analysis and profiling of small (50-1000 m/z) polar and non-polar molecules to find a characteristic metabolite pattern for the particular tissues. Second, the impact of insect feeding on the metabolome of the pitcher and leaf blade was studied. Using UPLC-ESI-qTOF and cheminformatics, about 2000 features (MS/MS events) were detected in the two tissues. They showed a huge chemical diversity, harboring classes of chemical substances that significantly discriminate these tissues. Among the common constituents of N. x ventrata are phenolics, flavonoids and naphthoquinones, namely plumbagin, a characteristic compound for carnivorous Nepenthales, and many yet-unknown compounds. Upon insect feeding, only in pitchers in the polar compounds fraction, small but significant differences could be detected. By further integrating information with cheminformatics approaches, we provide and discuss evidence that the metabolite composition of the tissues can point to their function.

Evaluation of in vitro testing strategies for hazard assessment of the skin sensitization potential of "real-life" mixtures: The case of henna-based hair-colouring products containing p-phenylenediamine.

BACKGROUND: Allergic contact dermatitis caused by henna-based hair-colouring products has been associated with adulteration of henna with p-phenylenediamine (PPD). OBJECTIVES: To develop a testing approach based on in vitro techniques that address key events within the skin sensitization adverse outcome pathway in order to evaluate the allergenic potential of hair-colouring products. METHODS: The following in vitro assays were used to test the sensitizing capacity of hair dye ingredients: the micro-direct peptide reactivity assay (mDPRA); the HaCaT keratinocyte-associated interleukin (IL)-18 assay; the U937 cell line activation test (U-SENS)/IL-8 levels; the blood monocyte-derived dendritic cell test; and genomic allergen rapid detection (GARD skin). Those techniques with better human concordance were selected to evaluate the allergenic potential of 10 hair-colouring products. RESULTS: In contrast to the information on the label, chromatographic analyses identified PPD in all products. The main henna biomarker, lawsone, was not detected in one of the 10 products. Among the techniques evaluated by testing hair dye ingredients, the mDPRA, the IL-18 assay, GARD skin and the U-SENS correlated better with human classification (concordances of 91.7%-100%) and were superior to the animal testing (concordance of 78.5%). Thus, these assays were used to evaluate hair-colouring products, which were classified as skin sensitizers by the use of different two-of-three approaches. CONCLUSIONS: Our findings highlight the toxicological consequences of, and risks associated with, the undisclosed use of PPD in henna-based "natural" "real-life" products.

Lapachol biotransformation by filamentous fungi yields bioactive quinone derivatives and lapachol-stimulated secondary metabolites.

Lapachol is a natural naphthoquinone with a range of biological effects, including anticancer activity. Microbial transformations of lapachol can lead to the formation of new biologically active compounds. In addition, fungi can produce secondary metabolites that are also important for drug discovery. The goal of this study was to evaluate the ability of filamentous fungi to biotransform lapachol into biologically active compounds and identify secondary metabolites produced in the presence of lapachol. Seven out of nine strains of filamentous fungi tested exhibited the ability to biotransform or biodegrade lapachol. The bioactive derivatives norlapachol and isolapachol were identified among biotransformation products. Moreover, lapachol stimulated the production of pyrrolo-[1,2-a] pyrazine-1,4-dione, hexahydro-3-(2-methylpropyl) and phenol-2,4-bis-(1,1-dimethylethyl), secondary metabolites already known to have antimicrobial and antioxidant activities. These results open the perspective of using these strains of filamentous fungi for lapachol biotransformation and efficient production of several biologically active compounds.

Naphthoquinone Metabolites Produced by Monacrosporium ambrosium, the Ectosymbiotic Fungus of Tea Shot-Hole Borer, Euwallacea fornicatus, in Stems of Tea, Camellia sinensis.

The tea shot-hole borer beetle (TSHB, Euwallacea fornicatus) causes serious damage in plantations of tea, Camellia sinensis var. assamica, in Sri Lanka and South India. TSHB is found in symbiotic association with the ambrosia fungus, Monacrosporium ambrosium (syn. Fusarium ambrosium), in galleries located within stems of tea bushes. M. ambrosium is known to be the sole food source of TSHB. Six naphthoquinones produced during spore germination in a laboratory culture broth of M. ambrosium were isolated and identified as dihydroanhydrojavanicin, anhydrojavanicin, javanicin, 5,8-dihydroxy-2-methyl-3-(2-oxopropyl)naphthalene-1,4-dione, anhydrofusarubin and solaniol. Chloroform extracts of tea stems with red-colored galleries occupied by TSHB contained UV active compounds similar to the above naphthoquinones. Laboratory assays demonstrated that the combined ethyl acetate extracts of the fungal culture broth and mycelium inhibited the growth of endophytic fungi Pestalotiopsis camelliae and Phoma multirostrata, which were also isolated from tea stems. Thus, pigmented naphthoquinones secreted by M. ambrosium during spore germination may prevent other fungi from invading TSHB galleries in tea stems. The antifungal nature of the naphthoquinone extract suggests that it protects the habitat of TSHB. We propose that the TSHB fungal ectosymbiont M. ambrosium provides not only the food and sterol skeleton necessary for the development of the beetle during its larval stages, but also serves as a producer of fungal inhibitors that help to preserve the purity of the fungal garden of TSHB.

Application of ionic-liquid-magnetized stirring bar liquid-phase microextraction coupled with HPLC for the determination of naphthoquinones in Zicao.

In this study, a green, rapid, and simple method, ionic-liquid-magnetized stirring bar liquid-phase microextraction was developed for the determination of naphthoquinones, including shikonin and ß,ß'-dimethylacrylshikonin, in Zicao. This method permits active magnetic stirring, extraction, and pre-enrichment in a single device simultaneously, so the extract is conveniently collected. The analytes were extracted from the sample to ionic liquid-magnetized stirring bar, then the analyte-adsorbed magnetized stirring bar can be readily isolated from the sample solution by a magnet. The key experimental parameters were investigated and optimized, including the type and volume of ionic liquid, extraction time, salt concentration, stirring speed, and pH. The recoveries were in the range of 89.47-102.38%, and good reproducibilities were obtained with relative standard deviation below 5.36%. Compared with the conventional extraction methods, the proposed method is quicker and more effective.

Monitoring of Lawsone, p-phenylenediamine and heavy metals in commercial temporary black henna tattoos sold in Turkey.

BACKGROUND: Henna has a very low allergic potential, and severe allergenic contact dermatitis is mainly caused by p-phenylenediamine (PPD), which is added to temporary black 'henna tattoos', and potentially also by some heavy metals. OBJECTIVE: To determine the presence of, and quantify, Lawsone, PPD and heavy metal contaminants (cobalt, nickel, lead, and chromium) in commercial temporary black henna tattoo mixtures (n = 25) sold in Turkey. METHODS: Lawsone and PPD concentrations were analysed with high-performance liquid chromatography, and heavy metal quantification was performed with inductively coupled plasma mass spectrometry. RESULTS: PPD was found in all 25 black henna tattoo samples purchased from tattoo shops; levels varied between 3.37% and 51.6%. Lawsone was detected (0.002-88.2%) in 21 of the 25 temporary black henna tattoo samples analysed. Heavy metal contaminant levels were 0.44-3.11 ppm for Co, 1.13-2.20 ppm for Ni, 1.59-17.7 ppm for Pb, and 35.0-76.9 ppm for Cr. CONCLUSIONS: Our results suggest that commercial temporary black henna mixtures containing PPD levels up to 51.6% pose a risk of contact sensitization and severe allergic contact dermatitis among users. It is important to identify both the additives and metallic contaminants of black henna tattoo products; the significance of metal contaminants has still to be assessed.

Diversity of Polyhydroxynaphthoquinone Pigments in North Pacific Sea Urchins.

Using high-performance liquid chromatography with diode-array detection and mass spectrometry (HPLC-DAD/MS) we investigated the composition of polyhydroxynaphthoquinone (PHNQ) pigments from sea urchins Strongylocentrotus pallidus, St. polyacanthus, St. droebachiensis, Brisaster latifrons and Echinarachnius parma, collected in the Sea of Okhotsk and the Bering Sea. Identification of PHNQ pigments from sea urchins St. polyacanthus, B. latifrons, and E. parma was performed for the first time. Among the usual PHNQ pigments, mono- and dimethoxy derivatives of spinochrome E, not previously found in other sea urchins, were discovered in St. polyacanthus and St. droebachiensis. In St. droebachiensis, two monomethoxy derivatives of echinochrome A were detected, isolated previously from only tropical sea urchins. It was found that the composition and total content of pigments of St. droebachiensis depends on the collection area of the sea urchins and its depth and varies from 88 to 331 µg/g of dry shells. Sea urchins St. pallidus, B. latifrons and E. parma had average values for PHNQ pigment content, approximately 30 µg/g, and St. polyacanthus had a low PHNQ content, 13 µg/g.

[Dynamic variation of components in exocarp of Juglans mandshurica with browning based on UPLC-Q-TOF/MS].

To analyze the dynamic changes in components in exocarp of Juglans mandshurica at different browning periods. Twenty-six batches of exocarp of J. mandshurica samples from thirteen browning periods were assessed by UPLC-Q-TOF-MS/MS. The formula of different compounds were determined by accurate mass and isotopic abundance ratio from target screening function of Peakview 2.0/masterview1.0 software. Then their structures were determined by analysis of MS/MS fragment or comparison with standard substances and references. The contents of chemical components were changed significantly in different browning periods and twenty five compounds were identified or inferred. Of the 13 naphthoquinone compounds, the contents of 6 compounds with similar parent nucleus as juglone and 3 naphthoquinone glycosides compounds were decreased significantly, and 4 naphthoquinone derivatives such as regiolone were produced; the contents of four flavones and two phenolic acids compounds were decreased significantly; and the contents of 6 diarylheptanoids compounds were increased significantly. UPLC-Q-TOF/MS method can be used to identify and analyze the chemical constituents from exocarp of J. mandshurica rapidly and accurately, and analyze the rules of dynamic changes, to reveal the browning of Chinese medicinal materials and its effects on compositions of fruits and vegetables.

[Simultaneous determination of eight hydroxyl naphthoquinones in different parts of Arnebiae euchroma by HPLC].

This present study is to develop an HPLC method for simultaneous determination of eight hydroxyl naphthoquinones, shikonin, ß-hydroxy-isovalerylshikonin, acetylshikonin, ß-acetoxy-isovalerylshikonin, deoxyshikonin, isobutyrylshikonin, ß,ß'-dimethylacrylshikonin and isovalerylshikonin. The eight constituents were measured on a Waters Xbridge C18 column (4.6 mm×250 mm，5 µm) with isocratic elution of acetonitrile-0.05% formic acid solution (70∶30) at a flow rate of 1.0 mL•min⁻¹. The detection wavelength was 275 nm and the column temperature was 30 ℃. The results of content determination indicated that significant differences of the eight compounds exist in every part of Arnebia euchroma，in which the highest part was the root bark, followed with the root, then the stem residues. The content of the xylem of root and aerial part was lower than the above parts. The results provided scientific basis for the medicinal parts of A. euchroma.

Determination of lead, cadmium and nickel in hennas and other hair dyes sold in Turkey.

The concentrations of lead, nickel and cadmium in various hennas and synthetic hair dyes were determined by high-resolution continuum source graphite furnace atomic absorption spectrometry (HR-CS GFAAS). For this purpose, 1 g of sample was digested using 4 mL of hydrogen peroxide (30%) and 8 mL of nitric acid (65%). The digests were diluted to 15 mL and the analytes were determined by HR-CS GFAAS. All determinations of Pb and Cd were performed using NH4H2PO4 as a modifier. The analytes in hair certified reference materials (CRMs) were found within the uncertainty limits of the certified values. In addition, the analyte concentrations added to hair dye were recovered between 95 and 110%. The limits of detection of the method were 48.90, 3.90 and 12.15 ng g(-1) for Pb, Cd and Ni, respectively and the characteristic concentrations were 8.70, 1.42 and 6.30 ng g(-1), respectively. Finally, the concentrations of the three analytes in various synthetic hair dyes with different brands, shades and formulae as well as in two henna varieties were determined using aqueous standards for calibration. The concentrations of Pb, Cd and Ni in hair dyes were in the ranges of LOD-0.56 µg g(-1), LOD-0.011 ng g(-1) and 0.030-0.37 µg g(-1), respectively, whereas those in the two hennas were 0.60-0.93 µg g(-1), 0.033-0.065 ng g(-1) and 0.49-1.06 µg g(-1), respectively.

Concentrations of buparvaquone in milk and tissue of dairy cows.

AIM: To determine the concentration of the anti-theilerial drug buparvaquone in the milk and tissue of dairy cattle following treatment with two different formulations, and to assess the effect of clinical theileriosis on the concentration of buparvaquone in milk. METHODS: Healthy lactating dairy cows (n=25) were injected once (Day 0) I/M with 2.5 mg/kg of one of two formulations of buparvaquone (Butalex; n=12 or Bupaject; n=13). Milk samples were collected from all cows daily until Day 35. Five cows were slaughtered on each of Days 56, 119, 147, 203 and 328, and samples of liver, muscle and injection site tissue collected. Milk samples were also collected from cows (n=14) clinically affected with theileriosis for up to 21 days after treatment with buparvaquone. Milk and tissue samples were analysed by liquid chromatography-mass spectrometry; limits of detection (LOD) were 0.00018 mg/kg for muscle and 0.00023 mg/L for milk. Concentrations of buparvaquone in milk and tissues were log10-transformed for analysis using multivariate models. RESULTS: In healthy cows, concentrations of buparvaquone in milk declined with time post-treatment (p<0.001), but were above the LOD in 11 of 25 cows at Day 35. Concentration in milk was higher one day after treatment in cows treated with Butalex than in cows treated with Bupaject, but not different thereafter (p=0.007). Concentrations of buparvaquone in muscle were below the LOD for four of five animals at Day 119 and for all animals by Day 147, but were above the LOD at the injection site of one cow, and in the liver of three cows at Day 328. Tissue concentrations did not differ with formulation nor was there a formulation by time interaction (p>0.3). Concentrations of buparvaquone in the milk of clinically affected animals were not different from those of healthy animals at 1 and 21 days post-treatment (p=0.72). Between 21 and 25 days post-treatment concentrations were below the LOD in 9/14 milk samples from clinically affected cows. CONCLUSIONS: Detectable concentrations of buparvaquone were found in the milk of some cows for at least 35 days and in the liver and injection site of some cows until at least 328 days after injection. There were no biologically meaningful differences in milk or tissue concentrations between the formulations, or in the milk concentrations for cows that were clinically affected compared with those that were healthy at the time of treatment.

[Determination of five naphthaquinones in Arnebia euchroma by quantitative analysis multi-components with single-marker].

This study is to determine five naphthaquinones (acetylshikonin, ß-acetoxyisovalerylalkannin, isobutylshikonin, ß,ß'-dimethylacrylalkannin,α-methyl-n-butylshikonin） by quantitative analysis of multi-components with a single marker (QAMS). ß,ß'-Dimethylacrylalkannin was selected as the internal reference substance, and the relative correlation factors (RCFs) of acetylshikonin, ß-acetoxyisovalerylalkannin, isobutylshikonin and α-methyl-n-butylshikonin were calculated. Then the ruggedness of relative correction factors was tested on different instruments and columns. Meanwhile, 16 batches of Arnebia euchroma were analyzed by external standard method (ESM) and QAMS, respectively. The peaks were identifited by LC-MS. The ruggedness of relative correction factors was good. And the analytical results calculated by ESM and QAMS showed no difference. The quantitative method established was feasible and suitable for the quality evaluation of A. euchroma.

[Dynamic variation of major effective components in fresh rejuvenated fruit of Juglans mandshurica based on UPLC-Q-TOF-MS].

The changes in effective components of Juglans mandshurica at different harvest periods were analyzed by UPLC-Q-TOF-MS/MS. Eighteen batch samples of J. mandshurica from six harvest periods were assessed by multivariate statistical analysis with Markerview software. The formula of different compounds were determined by accurate mass and isotopic abundance ratio from target screening function of Peakview 2.0/Masterview1.0 software. Then their structure were determined by analysis of MS/MS fragment or comparison with standard substances and references. Naphthoquinone are the major markers in samples of Juglans mandshurica from different harvest periods. Thirty-eight of naphthalenequinones were identified or inferred in J. mandshurica and contents decline gradually. UPLC-Q-TOF-MS method which develops a new strategy can identify and analyze chemical constituents from J. mandshurica rapidly and accurately, main chemical constituents can be used for quality evaluation and efficacy material research. The dynamic changes in the metabolite accumulation of J. mandshurica the basic data for harvesting medicinal plants at different times.

The chemical biology of naphthoquinones and its environmental implications.

Quinones are a group of highly reactive organic chemical species that interact with biological systems to promote inflammatory, anti-inflammatory, and anticancer actions and to induce toxicities. This review describes the chemistry, biochemistry, and cellular effects of 1,2- and 1,4-naphthoquinones and their derivatives. The naphthoquinones are of particular interest because of their prevalence as natural products and as environmental chemicals, present in the atmosphere as products of fuel and tobacco combustion. 1,2- and 1,4-naphthoquinones are also toxic metabolites of naphthalene, the major polynuclear aromatic hydrocarbon present in ambient air. Quinones exert their actions through two reactions: as prooxidants, reducing oxygen to reactive oxygen species; and as electrophiles, forming covalent bonds with tissue nucleophiles. The targets for these reactions include regulatory proteins such as protein tyrosine phosphatases; Kelch-like ECH-associated protein 1, the regulatory protein for NF-E2-related factor 2; and the glycolysis enzyme glyceraldehyde-3-phosphate dehydrogenase. Through their actions on regulatory proteins, quinones affect various cell signaling pathways that promote and protect against inflammatory responses and cell damage. These actions vary with the specific quinone and its concentration. Effects of exposure to naphthoquinones as environmental chemicals can vary with the physical state, i.e., whether the quinone is particle bound or is in the vapor state. The exacerbation of pulmonary diseases by air pollutants can, in part, be attributed to quinone action.

Chromatographic behavior of small organic compounds in low-temperature high-performance liquid chromatography using liquid carbon dioxide as the mobile phase.

Low-temperature high-performance liquid chromatography, in which a loop injector, column, and detection cell were refrigerated at -35ºC, using liquid carbon dioxide as the mobile phase was developed. Small organic compounds (polyaromatic hydrocarbons, alkylbenzenes, and quinones) were separated by low-temperature high-performance liquid chromatography at temperatures from -35 to -5ºC. The combination of liquid carbon dioxide mobile phase with an octadecyl-silica (C18 ) column provided reversed phase mode separation, and a bare silica-gel column resulted in normal phase mode separation. In both the cases, nonlinear behavior at approximately -15ºC was found in the relationship between the temperature and the retention factors of the analytes (van't Hoff plots). In contrast to general trends in high-performance liquid chromatography, the decrease in temperature enhanced the separation efficiency of both the columns.

A Novel Four-step Approach for Systematic Identification of Naphthoquinones in Juglans cathayensis Dode using Various Scan Functions of Liquid Chromatography-Tandem Mass Spectrometry along with Data Mining Strategies.

INTRODUCTION: Systematic analyses of naphthoquinones in Juglans cathayensis have not yet been reported. It is very challenging to identify naphthoquinones with various structural diversities, especially those at trace levels. OBJECTIVE: To develop an efficient analytical approach for systematic discovery and identification of naphthoquinones in Juglans cathayensis. METHODOLOGY: A novel four-step approach was evaluated by utilizing various scan functions of liquid chromatography-triple quadrupole-linear ion trap mass spectrometry (LC-QTRAP-MS/MS) and liquid chromatography-quadrupole time-of-flight tandem mass spectrometry (LC-QTOF-MS/MS) along with data mining strategies. First, MS/MS fragmentation behaviors of naphthoquinones were investigated. Second, multiple ion monitoring triggered enhanced product ion scan (MIM-EPI) with specified ions was conducted to identify targeted naphthoquinones. Third, other scan functions of QTRAP-MS/MS and data mining strategies were explored to identify untargeted naphthoquinones. Fourth, structural rationalization and confirmation of naphthoquinones were performed using QTOF-MS/MS via its accurate mass measurement and MS/MS fragmentation functions. RESULTS: Optimal scan methods and data mining strategies using QTRAP-MS/MS were obtained for identification of targeted and untargeted naphthoquinones. Consequently, 48 naphthoquinones including 24 novel ones were identified or tentatively identified from Juglans cathayensis. CONCLUSION: A novel four-step approach for efficient discovery and identification of naphthoquinones was developed by exploring various scan functions of current LC-MS/MS technologies and data mining strategies, providing an example for systematic characterization of certain classes of phytochemicals, especially trace analytes in complex samples.

[Multi-component quantitative analysis combined with chromatographic fingerprint for quality assessment of Onosma hookeri].

A method for simultaneous determination of the shikonin, acetyl shikonin and ß, ß'-dimethylpropene shikonin in Onosma hookeri and the chromatographic fingerprint was estabished by HPLC-DAD on an Agilent Zorbax SB-column with a gradient elution of acetonitrile and water at 0.8 mL x min(-1), 30 degrees C. The quality assessment was conducted by comparing the content difference of three naphthoquinone constituents, in combination with chromatographic fingerprint analysis and systems cluster analysis among 7 batches of radix O. hookeri. The content of the three naphthoquinone constituents showed wide variations in 7 bathces. The similarity value of the fingerprints of sample 5, 6 and 7 was above 0.99, sample 2 and 3 above 0.97, sample 3 and 4 above 0.90, and other samples larger than 0.8, which was in concert with the content of three naphthoquinone constituents. The 7 samples were roughly divided into 4 categories. The results above indicated that the using of this medicine is complex and rather spotty. The established HPLC fingerprints and the quantitative analysis method can be used efficiently for quality assessment of O. hookeri.