본 연구는 낙지다리(Penthorum chinense Pursh) 추출물을 기능성 화장품소재로 개발하기 위해 (−)‑epicatechin gallate를 지표성분으로 선정하고, 품질관리를 위해 High Performance Liquid Chromatography (HPLC)를 이용하여 분석법을 개발하였다. 분석에 사용된 칼럼은 Unison US-C18 (4.6 × 250 mm, 5 μm, Imtakt, USA)을 사용하여 0.05% (v/v) trifluoroacetic acid (TFA)와 메탄올을 이동상 조건으로 컬럼 온도는 30 ℃ 에서 유속은 1.0 mL/min 로 검출파장은 280 nm에서 검출하였다. International Conference on Harmonization (ICH) 가이드라인(version 4, 2005)을 근거로 하여 특이성, 직선성, 정밀성, 정확성, 검출한계 및 정량한계를 분석하여 분석방법을 검증하였다. 검출한계 및 정량한계는 각각 0.11 mg/mL 및 0.33 mg/mL로 나타났으며, 검량곡선은 상관계수값이 0.9999로 양호한 직선성을 보였고, 정밀성 분석결과 도 0.6% 이하로 확인하였다. 또한, 회수율은 99.51 ~ 101.92% 범위로 정확성이 있음을 알 수 있다. 따라서, 본 분석법은 낙지다리 추출물의 지표성분의 분석법은 적합한 시험법임이 검증되었다.

Background: Camellia sinensis L.(CS) is a perennial evergreen species of plant whose leaves are used to produce tea. In this plant species, the parts used are the leaves, sub-branch parts are thrown out. Methods and Results: Ethanol extract of sub-branch parts was used for isolation of major compounds by column chromatography. Structures were identified as caffeine (1), (-)-epicatechin (2) and (-)-epicatechin gallate (3) by interpretation of spectroscopic analysis, including 1H- and 13C-NMR. High-performance liquid chromatography (HPLC) method was used to compare the quantitative level of marker compounds in various extraction solvents of sub-branch parts of CS. The content of caffeine, (-)-epicatechin, and (-)- epicatechin gallate in 30% ethanol extract showed higher value with 3.28 ± 0.57 ㎎/g, 5.53 ± 0.88 ㎎/g, and 1.29 ± 0.24 ㎎/g, respectively. Conclusions: These results indicated that not only leaves parts but also sub-branch, could be a good source for the functional material and pharmaceutical industry.

Background : Alpinia Officinarum Rhizome (高良薑) is recorded as a roots of Alpinia officinarum Hance (Zingiberaceae) in Korea Pharmacopoeia and there is no established confirmation test and method for quantitative determination using marker compound. Using the six marker compounds, pinocembrin, (5S)-5-hydroxy-7- (4-hydroxy-3-methoxyphenyl) -1-phenyl-3-heptanone, pinocembrin, galangin, kaemferide, galangin 3-methyl ether, A simultaneous analysis method was developed to utilize the quality control. Methods and Results : In this study, YMC-pack column (ODS C18, 250 × 4.6 ㎜, 5 ㎛) from YMC was used and the temperature was set at 30℃. A soultion of 0.1% formic acid was using water (mobile phase A) and acetonitrile (mobile B), respectively, and the mobile phase B was set to 15 - 35% for 0 - 15 minutes and 35 - 40% for 15 - 60 minutes. And 270 ㎚ was set as the detection wavelength. The order of the components is pinocembrin (1, Rt: 23.2 min), (5S)-5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)-1-phenyl-3-heptanone (2, Rt: 37.8 min), pinocembrin (3, Rt: 44.9 min), galangin (4, Rt: 46.7 min), kaemferide (5, Rt: 48.6 min), galangin 3-methyl ether (6, Rt: 52.8 min) and Based on the retention time, ultraviolet spectrum and MS value, it was confirmed by comparison with standard compounds. Conclusion : In this study, we developed the simultaneous analysis method by six marker components of the Alpina officinarum Hance extract to develop a quality test method to scientifically identify Alpinia officinarum Hance.

Background : Coffee is one of the favorite brewed drink in the world where is distributed in Latin America, Southeast Asia, Southern Asia and Africa. Coffee has an effective antioxidant ability and reported about that. In this study, it was analyzed by using high performance liquid chromatography (HPLC) to establish the method about content of caffeine, chlorogenic acid, caffeic acid and p-coumaric acid in coffee. Methods and Results : Coffee was extracted with 70% EtOH in room temperature and evaporated at 45℃. All standard and sample extract were melted and diluted with 15% MeOH. Mobile phase was prepared using water with 0.01% phosphoric acid and MeOH. All standard and sample were analyzed with gradient elution (0 min : 15% MeOH, 35 min : 30% MeOH). The chromatograms were monitored at 272 and 320 ㎚. HPLC reported linear equation that based on the calibration curve for each standard compound (caffeine : Y = 1.04e + 004X – 3.21e + 003, R2 = 0.999890. chlorogenic acid : Y = 2.86e + 004X – 8.24e + 003, R2 = 0.999891. caffeic acid : Y = 2.07e + 004X – 1.21e + 004, R2 = 0.999894. p-coumaric acid : Y = 3.24e + 004X – 1.10e + 004, R2 = 0.999897). Standard compounds were determined with qualitative and quantitative analysis. The retention time of each peak of standard compounds were separated by chromatogram. Conclusion : In this study, we determined that the analysis method of compounds in coffee. In addition, we have confirmed that separation about the retention time of each peak of caffeine and chlorogenic acid in different solvent condition depending on acid buffer. This method can be use to determine standard compound in coffee.

Background : Morus alba L. (M. alba L.), belonging to the family Moraceae, is widely distributed in East Asia. Fruits of M. alba L. have been used in traditional herbal medicine due to their antioxidant, anticancer, and antidiabetic properties. Phenolics play a main role for the growth, development, and pigment accumulation of plants. In this study, metabolic profiling of white (M. alba L. ‘Turkey’) and red (M. alba L. ‘Cheongil’) fruits during maturation. Methods and Results : Phenolic compounds are secondary metabolites found in most of the higher plants. In the current study, the levels of phenolic compounds decreased during the maturation of Turkey and Cheongil fruits. Particularly, the Turkey fruits showed a dramatic decrease in the accumulation of phenolics. Principal component analysis (PCA) is one of powerful tools to identify overall patterns in the multivariat experimental data. The PCA score plots results revealed a clear classification between Cheongil and Turkey. Additioanlly, each group spread left to right in the X-axis by maturity. Two principal components of the score plot explained 71.1% of the total variance. Principal component 1 was associated with the separation of each group by maturity and isolation of Turkey 1. Conclusion : In this study, we investigated primary metabolites and secondary metabolites (phenolics) in the white fruits (M. alba L. ‘Turkey’) and red fruits (M. alba L. ‘Cheongil’) in order to provide information on change in metabolite patterns during maturation.

Background : Agastache rugosa (A. rugosa), belonging to the Lamiaceae family, is a medicinal plant mainly distributed in Korea and contains various phenolic compounds revealing anti-fungal and anti-HIV properties. This study is aim to investigate change in phenylpropanoid content of flowers at different developmental stages using high performance liquid chromatography (HPLC) and quantitative real time polymerase chain reaction (qRT-PCR). Methods and Results : The variation in the transcriptional level of phenylpropanoid biosynthetic genes and phenylpropanoid contents in the flowers of A. rugosa at different developmental stages was analyzed. The transcript levels of phenylpropanoid biosynthesis genes, including ArPAL (phenylalanine ammonia-lyase), ArC4H (cinnamate 4-hydroxylase), and ArCHS (Chalcone synthase), were high in flowers at 1st stage compared with flowers at 2nd and 3rd stages. On the other hand, the expression levels of flavonoid biosynthesis genes, including ArTAT (tyrosine amino transferase), ArHPPR (hydroxyl phenylpyruvate reductase), and ArRAS (rosmarinic acid synthase), were higher in flowers at 3rd stage than those of flowers at 1st and 2nd. These results were consistent with HPLC analysis revealing that most phenolic compounds were higher in flowers at 1st and 2nd stage but the level of rosmarinic acid was the highest in 3rd stage. Conclusion : Our findings provide the information on change in phenylpropanoid biosynthesis in A. rugosa flowers at different developmental stages.

Background : Galantamine is mainly obtained from the bulbs and flowers of Galanthus caucasicus, Galanthus woronowii, and other related genera such as Narcissus tazetta, Narcissus pseudonarcissus, Leucojum aestivum, and Lycoris radiata. Galantamine is used to treat Alzheimer’s disease (AD) and as an AD painkiller. Narcissus tazetta (N. tazetta), belonging to the Amaryllidaceae family, is a ornamental plant containing galantamine. In this study, metabolic profiling of N. tazetta different organs was performed. Methods and Results : The amount of galantamine in bulb of N. tazetta is the highest levels. About 0.61 ± 0.09 ㎎/g in bulb, 0.15 ± 0.17 ㎎/g in root, and 0.10 ± 0.0 ㎎/g in leaf. Contents of galantamine in root and leaf are not statistically significant. The total phenolic contents in leaf are the highest level. Rutin and kaempferol are identified all part of N. tazetta. On the other hands, 4-hydroxybenzoic acid is existed in leaf and caffeic acid is only existed in root. None of the bulbs except rutin and kaempferol are identified. Because plant secondary metabolism is closely related to plant primary metabolism, we used GC-TOF-MS on the levels of hydrophilic low-molecular-weight molecules in the N. tazetta. A total of 41 metabolites, including sugars, amino acids, organic compounds, and phenolic acids, were identified and measured, and the resulting quantitative data were subjected to principal components analysis (PCA). The results of PCA of metabolic profiles clearly showed the lack of marked variance among different organs of L. radiata. Two principal components of the score plot explained 86.79% of the total variance (component 1; 55.40%, component 2; 31.39%). Component 1 resolved the separation of leaves from the other plant parts. Conclusion : Narcissus tazetta belongs to amaryllidaceae family. These family has various alkaloids, in particular, galantamine is beneficial to Alzheimer patients. All parts of N. tazetta produce galantamine, in particular, the highest level is existed in the bulb. In contrast, phenolic compounds are identified

Background : Members of Amaryllidaceae family produce several alkaloids with unique structures and a variety of medicinal properties. Galantamine, in particular, is one of the alkaloids approved by the Food and Drug Administration (FDA), and the European Registration Bureau for treatment of Alzheimer’s disease. Lycoris radiata (L. radiata), belonging to the Amaryllidaceae family, is a bulbous plant containing galantamine, which exhibits selective and dominant acetylcholinesterase inhibition. In this study, metabolic profiling of L. radiata different organs was performed. Methods and Results : Galantamine in root, bulb, and leaf of L.radiata analyzed by high performance liquid chromatography (HPLC). The amount of galantamine in leaf is about 1.07 ± 0.17 ㎎/g and it is the higher than bulb (0.88 ± 0.01 ㎎/g) and root (0.75 ± 0.01 ㎎/g). These results are statistically significant. Six phenolics are identified in L. radiata through high performance liquid chromatography. Total amounts of phenolics are the highest in bulb. Because plant secondary metabolism is closely related to plant primary metabolism, we used GC-TOF-MS on the levels of hydrophilic low-molecular-weight molecules in the L. radiata. A total of 41 metabolites, including sugars, amino acids, organic compounds, and phenolic acids, were identified and measured, and the resulting quantitative data were subjected to principal components analysis (PCA). The results of PCA of metabolic profiles clearly showed the lack of marked variance among different organs of L. radiata. Two principal components of the score plot explained 89.4% of the total variance (component 1, 51.86%; component 2, 37.54%). Component 1 resolved the separation of leaves from the other plant parts. Conclusion : Amaryllidaceae family synthesize galantamine belonging to alkaloids. Particularly, in bulb of Lycoris radiata, galantamine contents are the highest level. Thus, bulb is very beneficial for Alzheimer’s disease because the galantamine is well known as treatment of dementia of Alzheimer type.

Background : Lycoris radiata belongs to the Amaryllidaceae family and is a bulbous plant native to South Korea, China, and Japan. Galantamine, a representative alkaloid of Amaryllidaceae plants, including L. radiata, exhibits selective and dominant acetylcholinesterase inhibition. In this study, transcriptome analysis of L. radiata was performed. Methods and Results : Genes for galantamine biosynthesis were used to design primers for qRT-PCR. Quantitative RT-PCR was performed with LrActin as a reference gene for normalization. The RT-PCR results reveal the expression of LrPAL A and LrC4H at an early stage in the pathway. Interestingly, the expression of these genes was significantly higher in roots. However, the expression levels of LrNNR and LrN4OMT, which are closely involved in galantamine biosynthesis, were significantly higher in bulbs than leaves and roots. The expression levels of LrPAL B, LrTYDC, LrCYP98A3 and LrCYP76T were not significantly different among the different parts of the plants tested. HPLC analysis confirmed the presence of galantamine in all the organs, including the root (0.53 ± 0.07 ㎎/g dry weight), bulb (0.27 ± 0.04 ㎎/g dry weight), and leaf (0.75 ± 0.09 ㎎/g dry weight). The galantamine level in the bulb was 1.42 and 2.78 times higher than that in the root and leaf, respectively. The results of qRT-PCR for the eight galantamine genes revealed relatively high levels of genes expressed early, including LrPAL A, LrPAL B, LrC4H, and LrTYDC in the roots. However, in the bulbs, the levels of LrNNR and LrN4OMT were higher, which are crucial for galantamine biosynthesis. It also explains why bulbs contain high amounts of galantamine, which is likely due to the increased expression of LrNNR and LrN4OMT and the high levels of LrCYP96T, although the genes expressed early were expressed at high levels in the root. Conclusion : Transcript data of plants grown in a growth chamber revealed high expression levels of LrNNR and LrN4OMT genes that are closely involved in galantamine biosynthesis, and, as expected, we observed higher amounts of galantamine in the bulbs than in the root and leaves.

A QuEChERS method was developed for the analysis of diazinon, chlorfenapyr, and lufenuron in Napa cabbage. These pesticides represent three different chemical classes and are commonly used in cabbage production in Korea. The objective of the proposed method is a fast, inexpensive, and easy extraction of pesticides, followed by rapid analysis. The proposed method involves a microscale extraction using acetonitrile and dispersive solid phase extraction (SPE), allowing for time and materials savings. The pesticides were separated and quantified using reversed-phase HPLC-UV at 220 nm. The calibration curves showed good linearity (R2>0.97), and the limits of detection and quantification were ≤0.05 and 1 mg/kg, respectively. Intraday and interday recoveries were in the range 97-116% and 101-112% with RSD% ≤9% for concentrations between 0.5-5 mg/kg. Abnormal recoveries and a substantial matrix effect were initially observed for lufenuron, signaling that optimization of lufenuron recovery requires a slight modification of the method. The proposed method was tested on cabbage samples sold at local markets, which showed no detectable residues of the target pesticides. The proposed method could thus be used for monitoring these pesticides in cabbage and similar vegetables.

Background : Eleutheroside E (Syringaresinol-di-O-glucoside), one of the internal standard in Eleutherococcus senticosus (Rupr. & Maxim.) Maxim., showed effects on the anti-inflammation of arthritis and the decline in blood sugar. In this study, it was analyzed by using high performance liquid chromatography (HPLC) to find out the optimum experimental condition which indicated the highest content of eleutheroside E. Methods and Results: In total of 15 different experimental conditions were used to extract samples. Briefly, there were three different conditions in the temperature (room temperature, 70℃ and 100℃) and five solvent conditions (100% water, 30% EtOH, 50% EtOH, 70% EtOH and 100% EtOH) were used. The extraction condition of all samples were extracted in every 4 hours and repeated three times with a reflux cooling system. The HPLC was reported as eleutheroside E standard equivalents using the following linear equation based on the calibration curve : equation : Y = 7.72e + 0.04X – 7.83e + 004, R2 = 0.999918. Among 15 conditions, eleutheroside E was obtained with the highest amount (10.36 ± 3.81 ㎎/g of extract) at 100% EtOH extracted and room temperature condition. In this study, the eleutheroside E content was increased with increasing of EtOH concentration. And it can be detected by heating at 100% water extraction condition. Conclusion : These results demonstrated that the experimental condition at room temperature in 100% EtOH could be used in further studies to obtain the highest content of eleutheroside E in Eleutherococcus senticosus (Rupr. & Maxim.) Maxim.

Aralia elata Seemann (AE) has long been used as a folk medicine for the treatment of various diseases including diabetes mellitus, anti-arthritic, and anti-gastric ulcer agent in Korea, Japan, and China. This study was performed to establish a simple and reliable HPLC/UV analytical method for determination of most active anti-hypertensive compound, a 3-O-α-L-rhamnopyranosyl(1→2)-α-L-arabinopyranosyl hederagenin 28-O-β-D-xylopyranosyl(1→6)-β-D-glucopyranosylester (HE) for the standardization of the shoot extract of AE as a health functional food ingredient. The quantitative analytical method of HE was optimized by HPLC analysis using reverse-phase C18 column at 40°C with H2O and acetonitrile (70:30, v/v) as an isocratic mobile phase at a flow rate of 1.0 mL/min and detection wavelength of UV 205 nm. This HPLC/UV analytical method showed good specificity and high linearity in the tested range of 0.03125-2.0mg/ml with excellent coefficient of determination (R2) of 0.9999. The limit of detection and limit of quantification were 12.0 μg/mL and 36.5 μg/mL, respectively. Relative standard deviation (RSD) values of data from intra- and inter-day precision were less than 0.2% and 0.1%, respectively. These results indicate that the established HPLC/UV analytical method is very simple, specific, precise, accurate, and reproducible and thus can be useful for the quantitative analysis of HE as a functional anti-hypertensive compound in AE extract.

Background : Cirsium plants have been used for perennial edible plants and grow wild in the mountainous regions of Korea, including Cirsium setidens and C. pendulum. In particular, C. setidens is commonly called 'gondre', and it has been used medicinally for diseases such as hematuria, hepatitis and hypertension. Hairy roots cultivation can be used as a method for increasing production through mass culture of medicinal plants, and elicitors such as methyl jasmonate (MeJa) can be treated to increase the content of certain useful ingredients. In this study, the hairy root was derived from the leaf tissues of C. setidens and C. pendulum, and the HPLC pattern was compared by MeJa treatment. Methods and Results : Agrobacterium rhizogenes R1000 was used to induce hairy roots in 1/2× MS medium. In addition, the hairy roots was treated with MeJa for different time (0, 1, 2, 5, 10, 24, 48, 72 h) and with various concentrations (0, 10, 50, 100, 200, 300 μM). The HPLC pattern changes were analyzed by on-line HPLC-ABTS. Four new peaks were observed in both Cirsium setidens and C. pendulum hairy roots, all of which showed antioxidant activity. In the case of C. pendulum, chlorogenic acid content was about 4 times higher than that of leaves. These peaks, including chlorogenic acid, were all affected by MeJa treatment. Conclusion : Four peaks were detected in the hairy roots of C. setidens and C. pendulum not in the leaves, and they were confirmed to be affected by the treatment of MeJa. It is necessary to clarify the structure through the subsequent compound separation.

Background : Agrimonia pilosa (A. pilosa) Ledebour has been registered in The Korean Herbal Pharmacopeia (KHP). In the recent study, A. Coreana showed antioxidant and anti-inflammatory effect. However, Studies of components in Agrimonia coreana (A. Coreana) Nakai was not much. So, we compared A. pilosa and A. coreana by High performance liquid chromatography (HPLC). we perfomed Thin layer chlromatography (TLC) including the anatomical characteristics by using microscope. Methods and Results : The anatomical characteristics of A. pilosa were similar to them of A. coreana. But, fascicular fivers of A. Coreana was broader than it of A. pilosa. TLC were performed to identify the Rutin and Apigenin-7-glucuronide compound. In the extract of Agrimoniae Herba, they were identified on the spot of Rf 0.2, 0.4 in Ethyl acetate - Formic acid – Water (8 : 1 : 1). The Rutin and Apigenin-7-glucuronide were analysed by HPLC/UV with Thermo Column (5 μm, 4.6 × 250 mm, C18), the column temperature at 40 ℃ and a diode-array detector (DAD) seted at 255 nm and 338 nm. The mobile phase was composed of water and acetonitrile containing 0.1% formic acid with the flow rate 1 mL/min. All compounds showed good linearity (R2>0.999) within test ranges. Agrimoniae herba was extrated by four kinds of extraction methods: MeOH, 50% MeOH, EtOH and water. The highest extraction rate occurred, when it was treated with 50% Methanol for refluxing extraction (60min). Content of Rutin was found to be 0.07±0.00 mg/g in A. pilosa and 0.02±0.00 mg/g in A. coreana. Content of Apigenin-7-glucuronide was found to be 0.12±0.00 mg/g in A. pilosa and 0.11±0.00 mg/g in A. coreana. Conclusion : The anatomical characteristics of A. pilosa were similar to them of A. coreana. Contents of Rutin and Apigenin-7-glucuronide in A. pilosa was higher than them in A. coreana slightly. But there were observed the similar patterns of Agrimonia pilosa Ledebour and Agrimonia coreana Nakai on the finger print anelysis.

대한민국 식품의약품안전처(식약처)는 포름알데하이드 분석법으로 2,4-dinitrophenylhydrazine (DNPH) 유도체화-고성능액체크로마토그래프법(HPLC)을 고시하고 있다. 본 연구는 고시법의 복잡한 시료 전처리 과정을 개선하여 화장품 분석에 편리하게 사용할 수 있는 유도체화법을 개발하고자 수행되었다. 전처리 법을 간단하게 하기 위하여 pH, 시간 및 온도 등 반응조건을 최적화하였다. 이 전처리법은 초산염 완충액(pH 5.0)을 사용한 검액의 pH 조정, 디클로로메탄을 사용한 액-액 분획 그리고 감압농축기를 사용한 증발건조와 같이 식약처 고시법의 복잡한 과정이 필요 없다. 유도체화 과정을 통하여 생성된 formaldehyde dinitrophenylhydrazone (formaldehyde-DNP)는 식약처의 시험방법을 약간 변형한 역상 HPLC법으로 분리하고 정량하였다. 2 ∼ 40 ppm 농도 범위의 표준액들을 가지고 수행한 검량선 작성 결과, 본 시험법은 상관계수 값 이 0.9999로 좋은 직선성을 보여주었다. 본 실험의 최소검출한계(LOD)와 최소정량한계(LOQ)는 각각 0.2 ppm과 0.5 ppm이었다. 또한 회수율 실험결과는 실험방법이 매우 정확하고 재현성이 높음을 보여주었다. 따라 서 본 연구에서 제안된 시험법은 화장품 중 포름알데하이드를 신속하게 분석하는데 적용될 수 있을 것이다.

A new extraction method-heated ultrasonic extraction was qualitatively and quantitatively analyzed for the extraction of major ginsenosides from ginseng extract; this new high-performance liquid chromatography (HPLC) method was compared with the official extraction method of Korean industrial standards and standard for health functional food. Methods and Results : Ginsenoside compounds were analyzed for 35 minutes by the new HPLC analysis method using a Halo® RP-Amide column. The new HPLC analysis method was validated by the measurement of intra-day and inter-day precision, accuracy, limit of detection (LOD), and limit of quantification (LOQ) of each ginsenoside. The correlation coefficients (r2) for the calibration curves of the ginsenoside compounds were over 0.9997 in terms of linearity. The heated ultrasonic extraction method using ultrasonication for 30 minutes at 50℃ yielded higher amount of ginsenosides than the extraction method of the Korean industrial standards owing to the enhancement of extraction efficiency. Conclusions : Compared to the other extraction methods, the heated ultrasonic extraction method yielded a higher amount of ginsenoside Rb1 than Rg1 index compounds for the quality evaluation of ginseng roots.

Five caffeoylquinic acids of Aster altaicus var. uchiyamae Kitamura (Compositae) leaves were identified using standard compounds by HPLC and determined as follows: 3,4-di-O-caffeoylquinic acid (4.92 ± 0.06 ㎎/g dried weight), 3,5-di-O-caffeoylquinic acid (3.95 ± 0.13 ㎎/g), 4,5-di-O-caffeoylquinic acid (1.39 ± 0.10 ㎎/g), 5-O-caffeoylquinic acid (chlorogenic acid, 8.05 ± 0.21 ㎎/g), 3-O-caffeoylquinic acid (4.97 ± 0.18 ㎎/g). The total content of five caffeoylquinic acids were calculated as 26.73 ± 0.26 ㎎/g dried weight while the percentage of the five compounds in the MeOH extract was calculated as 25.22 ± 0.25%. The IC50 value of the MeOH extract scavenging peroxynitrite (ONOO - ) was shown as 5.16 ± 0.15 ㎍/㎖.