(Japanese horseradish, Eutrema japonicum) is the only cultivated species in the genus Eutrema with functional components that provide a strong pungent flavor. To evaluate genetic resources for breeding, we surveyed variations in the two most abundant isothiocyanate (ITC) components in , allyl isothiocyanate (AITC) and 6-methylsulfinyl (hexyl) isothiocyanate (6-MSITC, hexaraphane). We also examined the phylogenetic relationships among 36 accessions of wild and cultivated in Japan using chloroplast DNA analysis. Our results showed that (i) the 6-MSITC content in currently cultivated accessions was significantly higher than in escaped cultivars, whereas the AITC content was not significantly different. (ii) Additionally, the 6-MSITC content in cultivated was significantly lower in the spring than during other seasons. This result suggested that the 6-MSITC content responds to environmental conditions. (iii) The phylogenetic position and the 6-MSITC content of accessions from Rebun, Hokkaido Prefecture had different profiles compared with those from southern Honshu, Japan, indicating heterogeneity of the Rebun populations from other Japanese accessions. (iv) The total content of AITC and 6-MSITC in cultivated was significantly higher than that of wild . In conclusion, old cultivars or landraces of , “zairai”, are the most suitable candidates for immediate use as genetic resources.

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We investigated the effect of rhizome extract on atopic dermatitis (AD) model mice. The extract was fed to the HR-1 hairless mice, which develop AD-like symptoms with a special diet (HR-AD diet). The extract was expected to reduce the symptoms induced. rhizome-containing HR-AD diet (5% and 10%) reduced the scratching behavior, and the 10% rhizome HR-AD diet significantly reduced scratching behavior on days 28, 35 and 42. Plasma components (histamine, eotaxin, IgE and thymus and activation-regulated chemokine (TARC)) were decreased in the 10% rhizome HR-AD diet. In histopathological examinations (toluidine blue (T.B.), major basic protein (MBP), CD4, IL-4, IL-5, eotaxin, TARC and IgE), the rhizome-containing HR-AD diet (5% and 10%) significantly reduced the number of positive stained cells. These results suggested that the rhizome extract improved the AD-like symptoms of HR-1 hairless mice.

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is a plant of Japanese origin. It belongs to the family Brassicaceae and produces various isothiocyanates (ITCs). To clarify the type I allergies inhibited by ITCs, we investigated the inhibitory effect on chemical mediator release from dinitrophenylated bovine serum albumin (DNP-BSA)-stimulated RBL-2H3 rat basophilic leukemia cells. Allyl ITC (AITC), sec-butyl ITC (s-BuITC), and 3-butenyl ITC (3-BuITC), which have 3 or 4 carbon chains, inhibited histamine release but did not inhibit the release of leukotriene B4 (LTB4) or cysteinyl LTs (CysLTs). 4-Pentenyl ITC (4-PeITC) and 5-hexenyl ITC (5-HeITC), which have 5 or 6 carbon chains and an unsaturated bond at the end, inhibited LTB4 release but did not inhibit the release of histamine or CysLTs. 6-Methylthiohexyl ITC (6-MTITC), 6-methylsulfinylhexyl ITC (6-MSITC), and 6-methylsulfonylhexyl ITC (6-MSFITC), which have a sulfur atom inserted at the end of a 6-carbon chain, inhibited the release of histamine, LTB4, and CysLTs and the elevation in intracellular Ca²⁺. These results suggest that ITCs inhibited type I allergies by inhibiting chemical mediator release and that the inhibitory effects on each chemical mediator were due to differences in the side chain structure of the ITCs.

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The antioxidative activities of (Eutrema MAXIM.) root, stem, and leaf extracts were evaluated by several methods. The stem extract was found to be a strong antioxidant when tested by the TBA method using deoxyribose as the substrate, whereas the HPTLC assay using linolenic acid as the substrate indicated that the activity of the leaf extract was greater than those of the stem and root extracts. All three extracts showed strong-OH radical scavenging activity when tested by the ESR method. Superoxide scavenging activity was extremely high in the leaf extract, whereas the stem and root extracts did not show any appreciable effect. These results suggest the occurrence of different antioxygenic principles in the leaf and stem of . As most of the leaf and stem has not been utilized so far, these findings will contribute to the total utilization of plants.

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Only one sentence of an article in Asahi Shinbun made me meet many persons concerned, study many documents and write this article. As mentioned in it, late Mr. Akira Takahashi was eager to cultivate high-grade in New Zealand after retirement. Unfortunately just before retirement he died of pancreatic cancer and left his excellent English Proposal to New Zealand government. Last autumn I happened to be given this proposal by his wife. And I, in his place, decided to get definite and reliable information on the following questions: 1. Who began to cultivate Wasabia japonica on New Zealand? 2. When and where did (do) they cultivate it? 3. Whether did (do) they sell it abroad or not? 4. How is its present situation in New Zealand? After getting many pieces of valuable information, this February I visited the three plantations of in New Zealand and met several persons concerned. In this article I mention 0. Personal motivation for investigation 1. Short history of cultivation in New Zealand 2. Present situation of cultivation in New Zealand 3. Present situation of imported in Japan 4. Present situation of consumption in New Zealand 5. Outlook for cultivation in New Zealand

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Clubroot disease was found on (Eutrema Maxim.) growing in drained paddy fields in Yamaguchi Prefecture. The clubroot incidence was high in plants transplanted from one of two nursery beds set in drained paddy fields, where the occurrence of clubroot on Cardamine flexuosa With., a cruciferous weed, was more frequently observed. In the inoculation test, the clubroot fungus from the weed showed high pathogenicity to as well as that from Chinese cabbage. It is suggested that the fungus from the weed affects in fields.

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The main pungent component of (Eutrema japonica) is known to be isothiocyanate and its derivatives, volatile substances.  Allyl isothiocyanate (AITC) accounts for more than half of isothiocyanate derivatives.  However, there is a little information on the effects of AITC on the immune system by analyzing the number of white blood cells (WBCs) over the course of days of AITC administration.  In the present study, we studied the effects of AITC (dose=20 mg/kg body weight/day for 10 days, subcutaneous: s.c.) on the number of WBCs (total WBCs, lymphocytes, monocyte, neutrophil, basophil and eosinophil) and plasma corticosterone concentrations in adult male rats.  Administration of AITC decreased significantly the number of total WBCs on days 1-4 post s.c. injection by 25-27%.  Administration of AITC also decreased the number of lymphocytes on days 1-10 by 21-36% and monocyte on days 1-8 by 28-78%.  However, administration of AITC increased the number of neutrophil on days 8-10 by 61-112%.  AITC did not change the number of eosinophil and basophil.  Plasma corticosterone concentrations during the experimental period were 4.7-8.4 times significantly higher in the AITC group than in the control group, indicating that AITC induced stress-responses.  The relative weights of thymus and adrenals per body weight were significantly lower and clearly higher in the AITC group than in the control group, respectively.  These results suggest that AITC-mediated stress-responses are at least in part attributable to changes in the number of circulating WBCs.

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玄米にニンニクの小片, オロシ, および揮発性成分抽出物を処理すると, 貯蔵害虫のコクゾウムシとコクヌストモドキに対して忌避効果を示したが, 殺虫活性はなかった. ニンニクおよびその抽出物は供試した農業害虫のコナガ幼虫とモモアカアブラムシに対しては忌避効果も殺虫活性も示さなかった. 一方比較試験に供試したトウガラシとワサビについては, いずれも貯蔵害虫に対して忌避効果がないか, あるいは著しく弱い効果しか示さなかったが, ワサビの揮発性成分は貯蔵害虫両種に対して殺虫活性を示した. ニンニクの揮発性成分を捕集しGC-MSで分析したところ, 4つの主要なピークが分離され, アリシンの急速な分解によって生成したスルフィド化合物と, その脱水素反応によって生成した環状化合物と推定された. 忌避効果がアリシン自体, その分解物, あるいはそれらの混合物に由来するかについては不明である.

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The effect of leafstalk (Wasabia japonica MATTUM.) extract on bone components in the femoral-diaphyseal and -metaphyseal tissues of aged female rats in vitro and in vivo was investigated. Femoral-diaphyseal and -metaphyseal tissues were cultured for 48 hr in Dulbecco's modified Eagle's medium (serum free) containing either vehicle or leafstalk extract (10, 25 or 50 μg/ml of medium). The presence of leafstalk extract (50 μg/ml) caused a significant increase in calcium content, alkaline phosphatase activity and deoxyribonucleic acid (DNA) content in the diaphyseal and metaphyseal tissues in vitro. However, the effect of leafstalk extract (50 μg/ml) in increasing bone components was completely abolished in the presence of cycloheximide (10^-6 M), an inhibitor of protein synthesis. Moreover, rats were orally administered leafstalk extract (10 or 20 mg/100 g body weight) once daily for 7 days. The calcium content, alkaline phosphatase activity and DNA content in the femoral-diaphyseal and -metaphyseal tissues of aged rats was significantly increased by the administration of leafstalk extract (10 or 20 mg/100 g) for 7 days in vivo. Meanwhile, body weight, serum calcium and inorganic phosphorus concentrations of female aged rats were not significantly altered by the administration of leafstalk extract (10 or 20 mg/100 g) for 7 days. The present study demonstrates that leafstalk extract has an anabolic effect on bone components in vitro and in vivo. The intake of leafstalk extract may have a preventive effect on bone loss with increasing age.

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E. ('高井', 2n=14) とA. rusticana ('赤芽',2n=32) の正逆交配による雑種胚の発達と崩壊を組織学的に観察した. 交配後50時間での花粉管の伸長を蛍光顕微鏡で観察したところ, いずれの組み合わせとも, 花粉管は胚珠に達していた. E. ×A.rusticanaの場合, 交配後10日目から崩壊する雑種胚が徐々に増加し, 交配後30日での生存率は6.0%となった. これに対して, A. rusticana×E. では,雑種胚の発達はE. ×A. rusticanaに比べて遅く,また, 崩壊も著しく急速に進み, 交配後30日には生き残った雑種胚を含む胚珠はほとんど観察されなかった.
胚珠培養にあたっては, 交配後, 10, 20および30日目の胚珠を培地に置床した. 培地はMS培地およびWhite培地を用いた. いずれの培地とも3%ショ糖と0.6g寒天を加え, pH5.8に調整した. 交配後20日目および30日目に置床したE. ×A. rusticana由来の胚珠から, 合計14個体の植物体が得られた.しかし, A. rusticana×E. では植物体は得られなかった. また, 本実験で使用した材料の場合, MS培地がWhite培地より適しているように思われた.得られた植物体は根端細胞の染色体観察 (2n=23)とアイソザイム分析から, E. とA. rusticanaの雑種であることが確認された.

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The active component in leafstalk (Wasabia japonica MATSUM.), which has a stimulatory effect on bone calcification in mouse calvaria tissue culture in vitro, was purified. leafstalk extract was obtained from a homogenate with 20% ethanol. The active component, which was found in ethanol extraction, was purified by gel filtration chromatography with HiLoad 26/80 Superdex 30 pg column and reversed-phase chromatography (RPC) on a resource 15 RPC 3 ml column. The result of ESI mass spectra of the purified active component showed that its material had a molecular weight of 158. The absorption spectra of the material gave the maximum absorption at a wavelength of 221 nm. The material in leafstalk extract differed from genistein and 17β-estradiol, which can stimulate bone calcification in vitro, and had a comparatively lower molecular weight. This study demonstrates that the material with a low molecular weight of 158 is an active component in leafstalk which stimulates bone calcification.

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Wasabi

(Wasabia japonica Matsum.) is a Japanese traditional spice. Many kinds of processed foods that include

wasabi

as an ingredient have been developed and produced. Here, we report on the detection of the myrosinase genes of

wasabi

from products containing

wasabi

. Because

wasabi

had multiple copies of myrosinase genes that closely resembled each other, primers were designed so that the gene family could be amplified. Polymerase chain reaction with the primers indicated that fragments of the myrosinase genes were amplified from DNA samples of all

wasabi

cultivars tested, but no bands were found when DNA of other species, such as horseradish, radish, spinach, and pea, were tested. In addition, the system could detect myrosinase genes from food products that included

wasabi

. These results suggest that this technique is applicable for the specific detection of

wasabi

in a variety of products.

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Transgenic plants over-expressing defensin gene were successfully produced in Phalaenopsis orchid by Agrobacterium-mediated transformation method. Embryogenic cell suspension culture of Phalaenopsis Wataboushi ‘#6.13’ was infected with A. tumefaciens strain EHA101 carrying a plasmid containing defensin gene and selectable marker nptII, hpt genes. Plantlets were regenerated through somatic embryogenesis from the calli selected on hygromycin-containing medium. Transformation of plantlets with defensin gene was confirmed by PCR analysis. Southern blot analysis confirmed successful integration of 1–4 copies of the gene. Production of the 5 kDa defensin protein with varying levels was confirmed in the leaf extracts of different transgenic clones using Western blot analysis. Most of the transgenic plants showed strong resistance to Erwinia carotovora, which causes soft rot disease in the control plant. These results suggest the usefulness of this gene for conferring the resistance to various diseases of Phalaenopsis and possibly other orchids.

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Liothrips wasabiae Haga and Okajima（Thysanoptera: Phlaeothripidae）was previously found only on field-cultivated

（Eutrema japonicum）in the Chugoku region（west Honshu, Japan）. However, it was found on water-cultivated in Shizuoka Prefecture（central Honshu, Japan）in 2016, where is cultivated in terraced fields with high humidity saturated by mountain streams. Unlike field-cultivated , sprayed insecticide on water-cultivated is susceptible to dilution due to the high humidity and mist. Therefore, we conducted an experiment to determine the lethal effects of three neonicotinoid insecticides on adult and larval L. wasabiae at concentrations below the normal dilution, since both the adult and larval insects usually occur concurrently. The three neonicotinoid insecticides registered to the insect are dinotefuran, nitenpyram, and imidacloprid. The results revealed that dinotefuran and nitenpyram demonstrated greater insecticidal activities against both the adult and larval L. wasabiae than imidacloprid; however, all three insecticides, when used in lower concentrations than normal, proved less effective. This indicates that dinotefuran and nitenpyram may be used when sprayed insecticide is likely to be diluted, such as in water fields, even when both adult and larval insects occur simultaneously, taking into consideration the reduced insecticidal effect.

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Five potato (Solanum tuberosum L.) cultivars were transformed with Agrobacterium tumefaciens strain EHA101 harboring defensin gene (isolated from Wasabia japonica L.) in a binary plasmid vector, pEKH1. The infected tuber explants co-cultivated for 3 days resulted in higher transformation efficiency (7–50% higher) for all cultivars than 2 days co-cultivation. PCR analysis showed an amplified fragment of defensin gene and the selectable marker, nptII gene in the genomic DNA of all clones rooted on MS medium supplemented with 100 mgl⁻¹ kanamycin, suggesting their transgenic nature. Southern blot analysis confirmed that transgenic plants integrated 1–6 copies of defensin gene into their genome. Expression of defensin protein was confirmed in the leaf extracts of independent transgenic clones by Western blot analysis. Antifungal assay of detached leaves from non-transformed control and transgenic plants indicated that transgenic plants were partially resistant to the fungal pathogen, Botrytis cinerea (gray mold).

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(Wasabia japonica) is a traditional condiment used in Japanese dishes. The rhizome shows potent amylolytic activity. The amylase activities of 12 Brassicaceae vegetables, including and three non-Brassicaceae root vegetables, were determined and compared. Among the 15 vegetables, showed the highest amylase activity (203 U g^-1 fresh weight). The second and third highest activities were found in horseradish (14 U g^-1 fresh weight) and sweet potato (8.6 U g^-1 fresh weight), respectively. The 12 other vegetables exhibited activities less than 3 U g^-1 fresh weight. Immunoblot analyses using an anti-radish (Raphanus sativus) β-amylase antibody indicated that contained an abundance of the antigen. Tissue printing indicated that the β-amylase antigen is generally distributed throughout the rhizome. These results suggest that contains a large amount of β-amylase, and therefore shows strong amylolytic activity.

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Myrosinase (β-thioglucoside glucohydrolase, EC 3.2.3.1) is involved in pungency generation in (Wasabia japonica Matsum.). We isolated full-length cDNA (WjMY1) encoding myrosinase from the petiole of . The deduced polypeptide has 545 amino acids with a molecular weight of 62107. WjMY1 mRNA is accumulated in whole organs of the plant ; the order of expression is petiole≥rhizome>leaf. The order was found to be similar to that of the specific activity of myrosinase. This is the first report de- scribing the isolation of a myrosinase gene from .

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The effect of leafstalk (Wasabia japonica MATSUM.) extract on bone metabolism in a tissue culture system using mouse calvaria in vitro was investigated. The calvaria tissues obtained from normal mice were cultured for 48 h at 37°C in 5% CO₂/95% air in Dulbecco's modified Eagle's medium (high glucose, 4.5g/dl) containing either vehicle or leafstalk extract (10, 50 and 250 μg/ml of medium). leafstalk extract was obtained from a homogenate with 20% ethanol. The presence of leafstalk extract (10 μg/ml) caused a significant increase in calcium content and alkaline phosphatase activity in the bone tissues. With higher concentrations (50 and 250 μg/ml), however, the effect was weakened. The bone deoxyribonucleic acid (DNA) content was not significantly altered by the presence of leafstalk extract (10 and 50 μg/ml). The leafstalk extract-induced increase in bone calcium content was completely prevented by the coexistence of cycloheximide (10^-6 M), an inhibitor of protein synthesis, suggesting that the effect of leafstalk extract is based on a newly synthesized protein component. Meanwhile, the anabolic effect on bone calcium content was not seen in the presence of the ethanol extract (50 μg/ml) from loquat leaf, cherry leaf, dried shiitake, gabaron tea, green tea (sencha), muskmelon, satsuma mandarin, tomato, blueberry, and soy bean. The present study demonstrates that leafstalk extract has an anabolic effect on bone calcification, in vitro.

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Long-term storage of in vitro-grown (Wasabia japonica MATSUMURA) meristems at low temperature using dimethylsulfoxide (DMSO) were investigated. Meristems were plated on solidified 1/2MS medium supplemented with 3% sucrose and 1% DMSO for a day, and then they were successfully stored at -5°C for 24 months. After the storage, meristems were recultured on solidified 1/2MS medium supplemented with 3% sucrose and 0.1mgl^-1 6-benzyladenine (BA). The survival rate was about 60%. Although many meristems appeared water-soaked after storage, most recovered to normal shoots.

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