| Morindae officinalis Radix(MOR), the root of Morinda officinalis How(family Rubiaceae, Bajitian in Chinese), is one of the main medicinal plants in South China and its geo-authentic habitats is Deqing, Guangdong Province. It has the effect of tonifying the kidney yang, strengthening sinew and bone and dispelling wind-dampness, which is recorded in Chinese Pharmacopoeia(2010 edition), has been used for impotence and seminal emission, infertility caused by uterine coldness, menstrual irregularities, cold pain in the lower abdomen, painful impediment caused by wind-dampness, limp wilting sinew and bone. Anthraquinones, oligosaccharides, iridoids are the main chemical components of MOR which have a various of pharmacological activities including antidepressant, antiosteoporotic, antioxidant, anti-inflammatory, promotion on sperm production, promotion of angiogenesis and neuroprotection. There’re 4 clinical medications of MOR, such as raw MOR, steamed MOR, salt steamed MOR and licorice-processed MOR. It has much differences morphological characteristics and chemical composition content between raw materials from different sources and the chemical compositions change a lot during processing, which may cause the difference in medicinal materials quality and curative effect. In consequence, the influence factors like different habitats, growth years, the differences among raw and processed products of MOR must be taken into consideration in its quality investigation and specification establishment.The chemical components and pharmacological activities of MOR may change during processing. The difference of HPLC or chemical constitute content among raw and processed products of MOR were investigated widely in the past few years, nevertheless, how the chemical components would change and its changing process were lack of systematic research. In order to control the quality of the processed products of MOR, a reversed-phase high performance liquid chromatography method was developed to establish chromatographic fingerprint of anthraquinones from different habitats, growth years and varieties, and the small leaves species from Deqing, harvested after 3-4 years’ plant were supposed to be items in good quality. As the raw materials, the changes of chemical anthraquinones and oligosaccharides in Morindae officinalis Radix during processing were investigated.The main studies were summarized as follows:(1) Study on HPLC fingerprint of anthraquinones in Morindae officinalis RadixThe HPLC fingerprint’s chromatographic conditions were determined through method validation. The chromatographic separation were perform on DiamonsilTM C18 reversed-phase column (4.6 mm×250 mm,5μm), with acetonitrile(A) and 0.4% phosphoric acid aqueous(B) as the mobile phase, using a gradient program of 20% to 95%A in total 55 min(05 min,20%A,5～10 min,20%～44%A,10～40 min,44%A,40～50 min,44%～95%A, 50～55 min,95%A). The flow rate was 1.0 mL/min, the injection volumn was 20μL, with the UV detection wavelength was set at 277nm and column temperature at 30℃.20 distinct characteristic peaks of anthraquinones were obtained in the HPLC fingerprint common pattern, and peak 13 were identified as rubiadin-1-methyl ether. The chromatographic peaks were rich with good resolution and less analysis time. A total of 32 samples of MOR from different collected locations, different growth years and different species were analyzed and chemical pattern recognition methods, like similarity analysis and hierarchical cluster analysis were used for their quality evalution. It indicated that Morinda officinalis native to the town Gaoliang possess good quality and the optimal harvesting year of is 3-4 after planting. The small leaves species were proven to be fine varieties due to its high content of anthraquinones. By studying the effect of post-harvest processing, we found that Morinda officinalis is liable to mildew after removing the xylem. The chemical constituents change a lot when it got mildew which may be unhealthy to human beings, so we suggest that the herb should be dried promptly with xylem.(2) The changes of anthraquinones in Morindae officinalis Radix during processingRubiadin-1-methyl ether and total anthraquinones were selected as quality index by HPLC fingerprint to discuss its changes under different condition such as the steamed time, the dosage of the salt, salt processing time, the dosage of licorice and decoction time. The results showed that anthraquinones in processed products of Morindae officinalis Radix change with the heating time and the addition of excipients.There is a solubilization for rubiadin-1-methyl ether under the heating process and the addition of excipients such as salt, while the content was gradually decreased with the extension of heating time. The content of rubiadin-1-methyl ether reduced with the influence of licorice in licorice-processed products. When the heating time extended and the excipients addition increased, the peak area of total anthraquinones increased first and then decreased. The processing technology of Morindae officinalis Radix were established and optimized on the basis of the research above.(3) The changes of oligosaccharides in Morindae officinalis Radix during processingThe chromatographic separation of oligosaccharides in MOR were perform on the HPLC-ELSD system with Waters XBridgeTM Amide column (4.6 mm×250 mm,3.5μm), the mobile phase consisted of 0.2% triethylamine acetonitrile(A) and 0.2% triethylamine aqueous(B), using a gradient program of 75% to 60%A in total 70 min(0～10min,75～70%A, 10～20 min,70%A,20～45 min,70～60%A,45～70 min,60%A). The flow rate was 0.8 mL/min, the injection volumn was 20μL, the column temperature was set at 30℃, with the gas flow of 2.0 mL/min and draft tube temperature of ELSD was set at 75℃.Simultaneous determination of six oligosaccharides(1-5 saccharides chain), namely D-fructose, D-glucose, sucrose,1-kestose, nystose, 1F-fructofuranosylnystose as well as the peak areas of oligosaccharides with 6～10 chain with an HPLC-ELSD method in different processed MOR during processing. The content of oligosaccharides increased first and then decreased with the change of moistening time, the steamed temperature during steaming and the dosage of the salt during salt steaming. With the extension of steaming time, the oligosaccharides with 1-3 chain increased while oligosaccharides with saccharides chain> 4 decreased, furthermore, some unknown constituents appeared and grew. So we draw conclusions that in steamed MOR, the water for moistening was 70 mL/100g, steaming temperature was 160℃ and processing time was 1hour, in salt steamed MOR, salt concentration was 3% and 1 hour was taken as the more appropriate processing time. |
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