| ObjectiveCuscutae Semen was derived from Cuscuta australis R.Br.and Cuscuta chinensis Lam.,Cuscutae Japonicae Semen was derived from Cuscuta japonica Choisy.There was confusion.This project studies Cuscutae Semen and Cuscutae Japonicae Semen from the aspects of resource distribution,morphological organization,component content,and efficacy,in order to provide the basis for the corresponding quality improvement in Chinese Pharmacopoeia and local standards of Chinese medicinal materials and whether Cuscutae Japonicae Semen can be used as a substitute variety.Methods1.By investigating the collections and existing documents of major digital herbarium at home and abroad,geographical distribution pattern and resource characteristicsof main distribution areas of Cuscuta australis R.Br.,Cuscuta chinensis Lam.,Cuscuta japonica Choisy.were analyzed.2.61 batches of Cuscutae Semen and 12 batches of Cuscutae Japonicae Semen were directly observed.Cuscutae Semen and Cuscutae Japonicae Semen were observed by stereopicroscope as well as the whole body shape and size of the seed umbilicus,the four-sided view [ventral view(the face of the seed umbilicus),the back view(the face opposite to the ventral face),the left view(the left side of the ventral face),the right view(the right side of the ventral face)];The section features of seed coat grid cells were observed by optical microscope;The characteristics of species umbilicus were observed by scanning electron microscopy.3.(1)Polysaccharide content of 61 batches of Cuscutae Semen and 12 batches of Cuscutae Japonicae Semen was determined by phenol-sulfuric acid method,and the effects of solid-liquid ratio,extraction temperature and extraction time on polysaccharide content were investigated by single factor test.The Box-Behnken Design test(BBD test)design principle was used to optimize the extraction process of polysaccharide.(2)HPLC fingerprints were established for Cuscutae Semen and Cuscutae Japonicae Semen,respectively,and SPSS 21.0,SIMCA 14.1 were used for cluster analysis(CA),principal component analysis(PCA),and orthogonal partial least squares discriminant analysis(OPLS-DA).(3)Quantitative analysis of multi-components with a single-marker(QAMS)was used to determine 8 components in 61 batches of Cuscutae Semen and 6 components in 12 batches of of Cuscutae Japonicae Semen.Cuscutae Semen used hypericin as the internal reference to determine 7 components of chlorogenic acid,caffeic acid,p-hydroxycinnamic acid,isoquercetin,isochlorogenic acid C,quercetin and kauniol.Cuscutae Japonicae Semen used chlorogenic acid as the reference to determine the content of 5 components of caffeic acid,p-coumaric acid,rutin,isoquercetin and isochlorogenic acid C.Compared with external standard method(ESM),the accuracy and feasibility of QAMS method are verified.4.108 SD rats were divided into blank group(group K),model group(group M),Jinkui kidney Qi Wan group(group J),Cuscutae Semen high-dose,medium-dose and low-dose groups(group TH,group TM,group TL),Cuscutae Japonicae Semen high-dose,medium-dose and low-dose groups(group DH,group DM,group DL),with 12 rats in each group and half male and half female.The rat model of kidney-yang deficiency was established by intramuscular injection of hydrocortisone(25 mg/kg),which was given at the same time,Cuscutae Semen and Cuscutae Japonicae Semen high-dose,medium-dose and low-dose groups were intragastricated with 3.75 g/kg,1.25 g/kg and 0.4167 g/kg water decoction,group J was given intragastric administration of 0.9375 g/kg,once a day for 14 consecutive days.The indexes of body weight,anal temperature,fecal water content and urine volume were determined;the related glandular organ indexes of hypothalamic-pituitary-adrenal axis(HPA)and hypothalamic-pituitary-gonadal axis(HPG),serum hormone levels and pathological changes of related tissues were detected.Results1.C.japonica had the most specimens,C.australis the least.C.chinensis,C.australis and C.japonica mainly grow at altitudes below 1000 m,400 m and 1200 m,respectively.2.The characters of Cuscutae Semen were mainly plump;most are near spherical;the central ventral view is slightly circular,the back view is slightly elliptical,the left view and the right view are slightly quasi-circular;The umbilicus are mostly oblate.Cuscutae Japonicae Semen is spherical or triangular;the ventral view and the right view are elliptical,the back view is semi-fan-shaped,and the left view is fan-shaped;umbilical round.The microscopical identification observed that the section of Cuscutae Semen seed coat grating cells had 2 layers,and the radial length of the inner row was about 1～2 times that of the outer row,while the grating cells of Cuscutae Japonicae Semen seed coat only had 1 layer,the radial length is about27.49～42.59 μm.The umbilicus of Cuscutae Semen were lip shaped,Cuscutae Japonicae Semen seed umbilical collar shape,umbilicus longer,umbilical groove wider.3.Composition studies have shown that(1)the optimal extraction parameters were: solid-liquid ratio 1:300 g/m L,extraction temperature 70 ℃,extraction time 8 h.The polysaccharide content range of Cuscutae Semen was 1.39% ～ 3.01%,and Cuscutae Japonicae Semen was 1.94%～4.97%.(2)The fingerprint of Cuscutae Semen was labeled with 16 common peaks,and peak 2(chlorogenic acid),peak 3(caffeic acid),peak 5(p-hydroxycinnamic acid),peak 8(hyperoside),peak 9(isoquercitrin),peak 13(isochlorogenic acid C),peak 14(quercetin),peak 15(kaonferol)were identified.Among them,peak 2(chlorogenic acid)and peak 8(hyperoside)were characteristic difference peaks.A total of 16 common peaks were identified of Cuscutae Japonicae Semen,including peak 3(caffeine acid),peak 5(coumaric acid),peak 8(rutin),peak 9(isoquercitrin),peak 2(chlorogenic acid)and peak 13(isochlorogenic acid C),among which peak 8(rutin)and peak 13(isochlorogenic acid C)were characteristic difference peaks.(3)QAMS method was established for the determination of 8 components in Cuscutae Semen and 6 components in Cuscutae Japonicae Semen,there is no significant difference between QAMS method and ESM,and the content of hypericin in Cuscutae Semen and isochlorogenic acid C in Cuscutae Japonicae Semen was the highest.4.Efficacy results show that,compared with the same dose of different herbs,Cuscutae Semen had better improvement effect on the urine volume and LH level of female rats under high dose(P<0.05,P<0.01);Cuscutae Japonicae Semen had better effect on thyroid index and LH level in male rats(P<0.01,P<0.05).At medium dose,Cuscutae Semen had better improvement effect on kidney index of male rats(P<0.05).At low dose,Cuscutae Japonicae Semen had better improvement effect on total urine volume(P<0.01).Cuscutae Semen had better effect on ACTH and T levels in male rats and E2 levels in female rats(P<0.05,P<0.01).In terms of histopathological changes,Cuscutae Semen ovary and testicle structure were more normal.ConclusionCuscutae Semen and Cuscutae Japonicae Semen codendron are widely distributed in China,There is a co-domain distribution.Cuscutae Semen more full,near spherical,seed coat grid cell section view 2 layers,inner row radial length is about 1～2 times of the outer row.Therefore,it is suggested that Chinese Pharmacopoeia improve its characters and microscopic characteristics.The polysaccharide content of Cuscutae Japonicae Semen was higher by UV spectrophotometry.It was found that Cuscutae Semen was dominated by flavonoids and Cuscutae Japonicae Semen was dominated by phenolic acids.It is suggested that one test and multiple evaluation method should be used to determine the content of cuscuta cuscuta.Cuscutae Semen is better than Cuscutae Japonicae Semen in treating kidney-yang deficiency to a certain extent,so it is suggested that Cuscutae Japonicae Semen should not replace Cuscutae Semen as medicine. |
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