| qPurpose:Radix Aucklandiae is the dry rhizome of Aucklandia lappa Decne.The drug is pungent and bitter in flavour, warm in nature, and acts on the spleen, stomach, large intestine, sanjiao and gallbladder channels. < Compendium of Materia Medica > said" Radix Aucklandiae only uses its crude samples when added into qi-regulating drugs, but no fire, and if solid the intestine, should be roasted with flour to cooked." Pungent and warm for dispersing, aromatic for enlivening the spleen, it is good at promoting circulation of Qi in the spleen, stomach and large intestine to remove stagnated food and alleviate pain. The herb is often used to treat Qi stagnation, gastro-intestinal distention, food retention, diarrhea, dysentery and abdominal pain.“Roasted product can stop the dysentery, both because its property and flavor are heavy, and cooked product has no dispersed property, dry and aromatic to keep, so it can solid large intestine, and often was used for diarrhea.”Thus it can be seen, the knowledge and clinical application of the raw Radix Aucklandiae with effect of regulating qi and roasetd Radix Aucklandiae with effect of antidiarrhea are early known in the ancient. But the depth study of processing principle is still undefined. Under the guidance of TCM theory, this thesis studied the effects of the raw Radix Aucklandiae, bran-roasted materials, costunolide and dehydrocostuslactone on gastro-intestinal motility sysmatically; combined with study of the change of chemical composition after bran roasted, find the therapeutic material basis and study the mechanism of the raw Radix Aucklandiae with effect of regulating qi and roasetd Radix Aucklandiae with effect of antidiarrhea, analysis the processing principles of Radix Aucklandiae, which will regulate processing technology and guide the clinical medicine.Material and method:1. MaterialRadix Aucklandiaes were purchased from the Menghai Research Institute of Agriculture in Xishuangbanna, Yunnan province. Zhai yanjun Professor of Liaoning University of traditional Chinese medicine identified them as the genuine. Costunolide reference substance (batch number: 111524-200503, purity?98%), Dehydrocostuslactone reference substance (batch number: 111525-200706, purity?98%) were purchased from national institutes for food and drug control.2. Method(1) The contents of the costunolide and the dehydrocostuslactone were estimated in raw and bran-roasted Radix Aucklandiae using HPLC.(2) Determinate the constituents of volatile oil in crude and bran-roasted Radix Aucklandia by GC-MS.(3) Established model of reserpine induced mouse spleen deficiency syndrome, to investigate the effect of the raw Radix Aucklandiae, bran-roasted materials on the gastric emptying, intestine propulsion, D-xylose intestinal absorption and the pepsin activity.(4) Establishment of senna and castor oil induced diarrhea model to study the anti-diarrhea action of bran–roasted materials and raw products; optimization of costunolide and dehydrocostuslactone effect on the experimental diarrhea with increase-decrease baseline geometric proportion design method.(5) By double-antibody sandwich enzyme-linked immunosorbent assay (ELISA) measured levels of gastrointestinal hormones in spleen deficiency rats and in serum cytokine TNF-?and IL-10 expression of in spleen deficiency mice.(6) Investigate the effect of the raw Radix Aucklandiae, bran-roasted materials costunolide and dehydrocostuslactone on the vitro intestinal smooth muscle contraction of normal and deficiency rats.(7) Determinate the effect of costunolide and dehydrocostuslactone on the smooth muscle myosin light chain kinase expression of intestine tissue of mice induced by reserpine with the western-blot experiments.(8) Optimization of the cutting technology and the bran-roased technology associated with pharmacodynamic index for Radix Aucklandiae by orthogonal design; using HPLC to determinate the stability of Radix Aucklandiae pieces. Results:1. In the raw and bran-roasted Radix Aucklandiae, the contents of dehydrocostuslactone were higher than that of costunolide, after bran roasted, the contents of the two index components were reduced, costunolide content declined more than dehydrocostuslactone, which leads to significantly lower the ratio of the two index components (P <0.01).2. The content of volatile oil, costunolide and dehydrocostuslactone in crude drug were higher than that of bran-roasted Aucklandia lappa Decne. The crude products essential oil were isolated 76 peaks, of which 34 compounds were identified; 63 peaks of the bran-roasted products essential oil were separated, identified 22 compounds. In addition, 12 low boiling point components in the bran-roasted material are not detected in the essential oil, there are three higher levels of compounds are not identified, and the relative contents of them increase significantly after roasted.3.?The results of gastric emptying and intestine propulsion experments in normal mice suggested that the effect of the raw and bran-roasted Radix Aucklandiae on gastric emptying had no effect (P> 0.05); Raw materials of the secondary dose and high dose promoted the intestine propulsion. Spleen reserpine gastric emptying results show that: the roasted materials of low dose, secondary dose can significantly contribute to gastric emptying; small intestine results show that: the raw materials of low, secondary and high doses of the small intestine porpulsion significantly promoted.?Normal rat intestinal absorption of D-xylose showed that low doses of bran-roasted materials promoted the intestinal absorption and increaseed pepsin activity significantly.4. In Anti-diarrhea experment, diarrhea index results showed that the Anti-diarrhea effect of bran-roasted materials are stronger than paper-roasted products and raw materials, and the Anti-diarrhea effect for senna of Radix Aucklandiae was better than that of castor oil. The ratio 0.43-1.00 of costunolide and dehydrocostuslactone effect on the experimental diarrhea is better by increase-decrease baseline geometric proportion design method.5. Gastrointestinal hormone and cytokine ELISA, the results showed that:?the raw and bran-roasted Radix Aucklandiae on the motilin's fallen has inhibited action, in which raw materials of the middle dose for motilin regulation is strongest. Raw products of middle dose, high dose of bran roasted materials can significantly increase serum gastrin levels, lower plasma vasoactive intestinal peptide, and each group showed no significant difference in plasma somatostatin.?Costunolide high doses can increase serum tumor necrosis factor TNF-?level, while reducing interleukin IL-10 levels of the deficency mice, while the dyhydrocustuslactone's regulation of these two cytokines was not obvious.6. The results of isolated intestine experments showed that:?bran-roasted materials and raw products on the natural state of normal rat intestinal smooth muscle in vitro showed a slight inhibition of the first after a strong stimulant, and a dose-dependent. Excitatory effects of raw materials are stronger than that of bran-roasted products.?Bran-roasted materials and raw products on the spleen rat intestinal smooth muscle stimulant were first inhibited, the overall performance is stimulant, raw materials are stronger than bran-roasted products.?Bran-roasted materials and raw products on the atropine, propranolol, high calcium and low calcium solution caused both abnormal regulation of intestinal contractions, and raw products Regulation of intestinal contractions are stronger than the bran-roasted materials.?costunolide for the natural state of normal rat intestine and spleen deficiency rat intestine showed excitatory effects, while the dehydrocostuslactone for the natural state of normal rats in vitro rat intestine and spleen deficiency intestine demonstrated inhibition in a dose dependent manner.7. The result of Western-blot experment showed that: the high dose costunolide for the deficiency mice can significantly increase the expression of intestinal smooth muscle MLCK, and the weak action of dehydrocostuslactone.8. Processing technology results:?the best cutting technology of Radix Aucklandiae is cutting 1-2mm thin pieces after soaking 10h with 0.3 times water.?The optimum processing technology was Radix Aucklandiae mixed with 30% wheat brans stir-frying for 10 minutes at 110?120?.?The index components of pieces of different thickness increased with the storage time firstly and then decreased. Conclusion:1. After bran-roasted, the chemical composition of Radix Aucklandiae occurred qualitative changes and quantitative changes, the ratio of the two index components costunolide and dehydrocostuslactone lowering significantly can be considered as to one of the main reason for enhancing the antidiarrheal effect of bran-roasted Radix Aucklandiae. In addition, in the essential oil of the raw materials and bran-roasted products, there are three unidentified compounds which maybe have relative with the antidiarrheal effect of bran-roasted Radix Aucklandiae, yet to be further studied.2. Regulating-qi mechanism of the raw Radix Aucklandiae: the content ratio of costunolide and dehydrocostuslactone is large in raw materials, and costunolide has significance act on promoting intestine, but the dehydrocostuslactone has weak inhibition for intestine movement. the synergistic reaction of these two components exhibite the stronger promoting gastric emptying and intestinal propulsion, promote the secretion of motilin and gastrin, reduce the secretion of vasoactive intestinal peptide to increase gastrointestinal smooth muscle contraction frequency, and promote blood circulation of the stomach to help digestion and absorption; and by regulating the serum cytokine TNF-?and IL-10 levels and adjust the network back to the Th1/Th2 cell balance and regulate the body's immune mouse spleen Functions, improve intestinal smooth muscle myosin light chain kinase expression and promote expression of bowel movements, regulate the spleen and stomach qi disorders.3. Antidiarrheal mechanism of the bran-roasted Radix Aucklandiae: the content ratio of costunolide and dehydrocostuslactone decreased significantly compared with the raw materials, namely the descending scope of costunolide content is rather obvious, while the content of dehydrocostuslactone relative to costunolide increase, resulting in synergistic reaction between the two has changed, showing reduced the action of promoting the small bowel, enhanced the action of the intestinal absorption. The content ratio of costunolide and dehydrocostuslactone in a certain proportion treated large intestine diarrhea well.4. The raw and bran-roasted Radix Aucklandiae act on the contraction of isolated intestine with two-way adjustment.5. Investigated process parameters should be integrated with the indicators to measure efficacy. Radix Aucklandiae is suitable for current use, and easy dissolution of active ingredients cut thin slices, which consistent with historical records "broken, thin slice". 6. One of the processing principle of Radix Aucklandiae is that the larger the content ratio of costunolide and dehydrocostuslactone, the stronger regulating-qi; after bran-roasted, the contentration of them decrease and the antidiarrheal effect enhanced. |
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