LIGUSTRI FRUCTUS is the fruits of Ligustrum lucidum Aiton (Oleaceae), traditionally used as tonic in China. Phytochemical analyses revealed triterpenoid compounds such as oleanolic and ursolic acids were principal ingredients of the drug. A method for quantitative analysis of oleanolic and ursolic acids using HPLC was developed, and the market samples were analyzed using that method. In order to analyze quality variation caused by processing, fresh fruits of L. lucidum and L. japonicum were collected and processed in several different ways. Compositions of oleanolic and ursolic acids of processed samples were not very different among processed samples. Two origin species, L. lucidum and L. japonicum could be distinguished by the content ratio of these compounds.

Saffron is the stigma of Crocus sativus L. and widely used as a spice and a crude drug. Because the price of saffron is very high, it suffers from various adulterations including coloration with synthetic dyes. In the Japanese Pharmacopoeia 17th edition (JP17), a purity test for aniline dyes is described for saffron. However, the target aniline dyes are not specified, and chloroform, a hazardous solvent which can be used only if no alternative method is available in newly listed JP monographs, is used in this test. Therefore, a new method using thin layer chromatography (TLC) to replace the purity test was examined. From a literature search for the synthetic pigments which were reported to be detected from saffron, five pigments, sudan III (1), sudan red G (2), methyl orange (3), auramine (4) and sunset yellow FCF (5), were selected as target pigments. In addition, tartrazine (6) and naphthol yellow (7), which are used in the purity test of saffron in European Pharmacopoeia, were also included as target pigments. TLC conditions to separate these pigments from the yellow pigments (crocins) contained in saffron, together with suitable detection methods to distinguish the synthetic dyes from crocins, were investigated and a TLC method suitable to replace the current purity test of saffron for aniline dyes in JP was established. In addition, as three yellow spots of crocins are clearly observed under this condition, this TLC method was also suitable for an identification test of saffron for JP.

In a previous study (Horii, C., et al., Shoyakugaku Zasshi, 68 (2), 65-69, 2014), the current authors examined the bioequivalence of Shoseiryuto decoction and its extract preparation, and findings from that study indicated that ephedrine and pseudoephedrine from plants in the genus Ephedra could serve as characteristic constituents with which to evaluate the bioequivalence of preparations. As in the previous study, we examined the bioequivalence, the current study used the Shoseiryuto formula of the decoction and the product. The change in concentration of the 9 constituents, paeoniflorin, gomisin A, scizandrin, glycyrrhizicacid, liquiritin, liquiritigenin, asarinin, [6]-shogaol, and zingerone, was observed. These characteristic constituents can be used to evaluate the bioequivalence of preparations after their oral administration.

A cross-over study was conducted by randomly dividing 6 healthy adult men into 2 groups and then orally administering the preparations. Results revealed variations in the plasma concentration of each constituent depending on when blood samples were taken, and this result was true for both the decoction and the extract. Analysis of variance did not reveal significant differences in constituents (except for zingerone) in the decoction or extract. Analysis of variance indicated that the preparation was a significant factor for variability in the C_max of zingerone. The plasma concentration of zingerone was low and measurements were not obtained with sufficient sensitivity, which presumably explains the results obtained. Analysis of variance indicated that the subject was a significant factor for variability in the peak plasma concentration (C_max) and the area under the curve for the plasma concentration (AUC_0-8) of paeoniflorin, and in the peak plasma concentration (C_max) of gomisin A, schizandrin, and [6]-shogaol.

The statistical power (1-β) of the C_max and the AUC_0-8 was deemed to be insufficient (less than 80%) for all of the constituents. Therefore, based on the data obtained in this study, we estimated the sample size needed to obtain sufficient power. For liquiritin, a sample size of 9 or more subjects per group would yield a C_max and AUC_0-8 with sufficient power (80% or more). For [6]-shogaol, a sample size of 5 or more subjects per group would do so. For gomisin A, a sample size of 18 or more subjects per group would do so. For schizandrin, a sample size of 15 or more subjects per group would suffice. However, a sample size of 61 or more subjects per group would not yield sufficient power for paeoniflorin, glycyrrhizic acid, or liquiritigenin.

The 9 constituents of Shoseiryuto are known to be representative compounds with active ingredients. Results suggested that increasing the sample size for gomisin A, schizandrin, liquiritin, asarinin and [6]-shogaol might allow those 5 compounds to serve as characteristic constituents with which to evaluate the equivalence of the prescribed preparations. The current results indicated that 4 compounds of zingerone paeoniflorin, glycyrrhizic acid and liquiritigenin could not, at the current point in time, acceptably serve as characteristic constituents with which to evaluate the equivalence of preparations.

Ligustrum Fruit is a crude drug derived from the fruit of Ligustrum lucidum W.T. Aiton or L. japonicum Thunb. (Oleaceae). It has several pharmaceutical activities including hepatoprotection, antioxidation and improvement of bone turnover and therefore, it is used as an ingredient in many crude drug products for nutrition fortification. In preparation for the listing of the drug to Non-JP Crude Drug Standards (Non-JPS), we designed an identification test using TLC and identified the marker spot as nuzhenide based on the spectroscopic data including ¹H-, ¹³C-NMR and MS together with the comparison of TLC, LC-MS and NMR with those of the authentic compound.

The established TLC conditions are as follows: chromatographic support, silica gel; developing solvent, EtOAc/MeOH/H₂O (7/2/1); developing length, 7 cm; detection, UV (254 nm) and 1-naphthol-sulphuric acid reagent; R_f value, 0.4 (nuzhenide).

7α-Hydroxystigmasterol (1), stigmasterol (2), 4-hydroxybenzoic acid (3), sitosterol-β-D-glucoside (4), adenosine (5) and 1-methyl-3-carboxy-6,7-dihydroxy-1,2,3,4- tetrahydroisoquinoline (6) were isolated from the seeds of Mucuna pruriens (L.) DC. var. utilis (Wall. ex Wight) Baker ex Burck cv. cinza. The structures were determined by comparisons of their spectral data with literature values and with those of commercially available samples and a synthetic compound. Compounds 1–3 and 5 were isolated from the seeds of M. pruriens for the first time.

The MeOH extract prepared from the aerial parts of Achillea millefolium showed potent aldose reductase (AR) inhibitory activity. Bioassay-guided fractionation of the extract resulted in the isolation of a previously undescribed flavonoid glycoside (1), as well as eight known flavonoid derivatives (2－9), caffeic acid (10), two known quinic acid derivatives (11 and 12), and two known sesquiterpene derivatives (13 and 14). Compounds 1－3, 8, 9, 11, and 12 showed AR inhibitory activity with IC₅₀ values ranging from 3.4 μM to 29.2 MM.