| BackgroundThe mechanism of pain has not been explained yet clearly, but it is the main reason for the patient to the hospital. Pain is usually divided into two kinds:one kind is disease signal that the pain is just a symptom of disease, such as cold fever headache brain hot, etc. When the disease was cured, the pain of disease signal would disappear. Another type is that the pain is the disease of itself., such as trigeminal neuralgia, herpes zoster neuralgia, cancer pain (The treatment is invalid for terminal cancer which has the serial of six months to a year, but the pain often make patients survive only two or three months.), limb pain, stubborn blacklegs pain, etc. Overall, the pain is difficult to control, when the pain threaten the patient's life, it will change the symptoms of the disease into disease, and need analgesic therapy.Pain medicine in developed countries has been greatly concerned. In 2000, the world health organization officially defined the "pain" that chronic pain is a kind of disease. And after breathing, temperature and pressure pulse, the pain become the fifth human life indications. The current treatment of chronic in china pain is still in the initial stage. The means of pain management is single, and the drug therapy is the Controllable analgesia, improve the quality of medical profession is always pain one major issue of common concern.Along with the study of traditional Chinese medicine and modern pharmacology, it 'is found that much Chinese herbal medicine has obvious analgesic effect of various pain, definite effect of analgesia. At the same time it have no obvious addiction, tolerance and side effects, etc. It is the analgesic drug research hot spot. Especially in recent years, China use superfine pulverizing technology comparatively mature of the preparation of Chinese medicine balm, micron refers to microns in diameter (average more than 15 mums) the TCM, effective parts, technical and compound preparations. It is the product of traditional Chinese medicine microns, not a new medicine. This drug can not only effectively keep the effective ingredients Chinese was not damaged, and because the systems particles increases, can effective than surface area of effective ingredients of promotional drugs, improve dissolved out biological utilization of drugs. It can increase drug targeting tropism sexist is helpful for controlling the distribution of drugs in the body; Its transmit more convenient in the body, not only can prolong drug interaction time with slow-release drugs, but also can achieve the purpose of targeted conveying, thus greatly improve the curative effect of traditional Chinese medicine, reduce its adverse effect, increase drug stability, and this is linked with the core of the modernization of traditional Chinese medicine "safe, effective and controllable, stable".Transversal drug delivery is a popular international medical research community; strong penetration (through the physical positioning of percutaneous drug) is the fastest growing technology branch. In particular if we combine several physical methods and penetration and synergy together, it does will be the most popular. More recent research methods:porous laser technology (laser microdissection LMD), electroporation technology (electropration), ultrasonic transversal technology (sonophoresis) and modern iontophoresis technology (iontophoresis) and so on. Ultrasound therapy is a targeted drug delivery conductance of the United States and other Western countries in recent years, the rise of new drug therapy. It is through a combination of laser microspore, electroporation, ultrasonic cavitations and ion into the four used in modern biological engineering technology of high-tech means, so that drugs enter the body through the skin lesions of tissues and organs, directly play the role of medicine. First, within a certain range and depth of the skin and tissue greatly increased the permeability of drugs, development of drug into the target tube of artificial biological pathways. Then the kinetic energy driven by a specific, along the biological pathways of drug particles through the skin, tissue and capsule, orientation, quantitative, fixed speed of access to diseased tissue and organs, and ultimately the development of drug in the target tissue infiltration area of high concentration, direct play the role of medicine to achieve the purpose of targeted therapy. In the medical profession it's a third generation of drug delivery methods. The role of advantages:1, through the drug speed, a hundred times higher than the natural infiltration; 2, enabling time, quantitative and fixed-speed delivery, local development of drug in the lesion area of high concentration, and promote the transport of drugs to the cells, increase the role of drugs, thereby enhancing the efficacy; 3, greatly increased the bioavailability of drugs to reduce the total amount of medication to avoid side effects; 4, stable and lasting efficacy than the oral medication long 2-3 times; 5, the adaptation range, In addition to chemical drugs, but also the body through the macromolecular colloidal particles of biological agents and traditional Chinese medicine ingredients; 6, painless, noninvasive, simple and convenient.The U.S. and other developed countries have successfully used this method to antibiotics, hormones, anti-cancer drugs, analgesics and other nine categories of more than 300 kinds of pesticides, especially those with high activity, high toxicity, high risk and in need of local high concentrations of drugs, with this method has played a unique treatment. China is also using this therapy the treatment of focal infections, prostate disease, bone and joint diseases such as clinical reports of pain. In recent years, more domestic and international research is a local analgesic drugs (such as anesthesia, analgesia and other non-steroidal anti-inflammatory drugs) treatment of percutaneous penetration of a variety of focal pain. Large number of experiments (including in vitro experiments, animal experiments and human studies) and clinical application has made gratifying achievements. But Western analgesics addiction, tolerance and other side effects remains a constraint on a wide range of clinical applications. China Medical Technology Co., Ltd. Tongzhou Beijing Noah ultrasound Conductivity Meter, with reference to technological innovation from both the United States for products of chemical drugs and biological drugs, and traditional Chinese medicine preparation for the coupling gel patch, its products have been used in clinical. But the patch to be adding a variety of other drugs, the dose per tablet is only adding 0.5ml, a large man-made factor, there is the local effective concentration of drug loading and easy to control, may affect efficacy, but there is insufficient use of inconvenience. In this study, the current cutting edge of modern medicine innovations-micron medicine technology, proven by modern pharmacological studies of Chinese medicine analgesic microns, the development of a gel containing Chinese drugs micron chip storage library, the gel patch with skeletal characteristics of transversal drug delivery systems, storage library micron medicine in the role of ultrasound conductivity meter, release the drug directly into the skin and spread to the lesion, and maintain steady state plasma concentrations to achieve rapid analgesia purposes. International medical community has concluded that this method will become the new century, the leading drug treatment.Powder for my blood analgesia Traditional Chinese Medicine, Professor Leung ancestral homes for the treatment of acute soft tissue injury experience of foreign rule side by Drynaria, notoginseng, safflower and other herbs composition, with blood stasis, Stretch muscle, consumer swelling and pain, bone injury such as the role continued, mainly for the treatment of sprain pain rheumatalgia, frostbite swelling, soft tissue injury and fractures. The Chinese product coupled micro-gel patch, the first Chinese prescription "blood analgesia San"-micron technology, and by pharmacological experiments confirmed its efficacy, and then refer to the relevant U.S. technology to produce medicine ultrasound conductivity gel micro SMD The placement of the fiber network woven by the improved materials for the filled micro gel matrix hydration medicine storage library, both good carrying capacity and controlled release capabilities, coupled with the ultrasound and electrical conductivity and so on, to meet the conductance through ultrasound skin treatment needs, while allowing the standardization of dosage, standardization, and agent production processes, quality control, standardization and normalization.This study included three items, namely:blood orthogonal design of the extraction of bulk analgesic studies, blood scattered quality standard analgesic control study of percutaneous penetration of ultrasound conductivity micron bulk analgesic mechanism of blood and applied basic research.Orthogonal design of blood scattered extraction of analgesic studies:the reference to the traditional water extraction of blood scattered analgesic efficacy was not obvious, suggesting that the active ingredient may be due to inadequate extraction, the main active drug is Drynaria represented dihydro naringin flavonoids, soluble in ethanol, acetone and other organic solvents, Robinson is the main active ingredient drug Panax notoginseng saponins and ginsenosides (Rbl, Rb2, Rb3, Rc, Rd, Re), etc., soluble in methanol, ethanol and other organic solvents. In this study, we chose the experiment makers were prescription alcohol extract yield and active ingredients of herbs Drynaria naringin content, the use of L9 (34) orthogonal experimental pain bulk alcohol on blood and screening of the extraction process.Casual blood analgesia controls the quality standards:for better internal quality control of drugs, to ensure the safety and effectiveness of clinical medicine, were scattered blood qualities standard of analgesia. In this study, TLC was scattered on the blood of Drynaria, notoginseng, safflower, Eupolyphaga, barbata, qualitative identification of borneol, and the compound in the salt content of heavy metals and arsenic limits for experimental study for the development of quality standards for reference.Percutaneous penetration of ultrasound conductance blood micron bulk analgesic mechanism and application of basic research:percutaneous ultrasound conductance through new technologies and micron Chinese medicine formulations of a combination of new experimental and applied basic research, to observe the conductance of percutaneous ultrasound through scattered into the micro blood analgesia, physical therapy and ultrasound conductance with external application of analgesic effect of micro blood circulation in the rabbit model of soft tissue injury to the extent Panax micron powder, soft tissue swelling at the area of contusion, the number of changes in peripheral blood leukocytes by light microscopy histology observations were assayed. Observed treatment, ultrasound conductivity also investigated the role of transversal delivery method for the conditions and factors, its mechanism and to examine drug safety.ObjectiveFist, optimize and screen the extraction craft of the powder of promoting blood flow and relieve pain, and observe the influence of ethanol-extraction concentration, adding alcohol quantity, duration, frequency and other factors on extraction craft of the powder of promoting blood flow and relieve pain, Second, establish a good quality control standard of the powder of promoting blood flow and relieve pain and complements the reference standard of quality of traditional medicine. Third, observed the influence of using ultrasound to import micron traditional Chinese medicine on the rabbits of acute soft tissue injury and provide the basis for its clinical application and the transformation of achievements. At last, apply for patent.MethodsStudy on extraction of the powder of connecting bones and reducing swelling by orthogonal design method1. Factors and levels of experimental investigation:Investigate three factors (volume fraction of ethanol, add alcohol quantity and extraction time). Meanwhile, select three conventional levels of each factor, which are 85%,65%,50% of ethanol volume fraction.2. Mode of processing:Optimize and screen the extraction craft of the powder of promoting blood flow and relieve pain with L9 (34) orthogonal experimental method.3. Observation index:the ratio of extracted alcohol, the ratio of absorbed alcohol and the content of naringin measured by HPLC.Quality control standard of the powder of promoting blood flow and relieve pain1. Qualitative identification method:Identification Rhizoma Drynariae, pseudo-ginseng,red flower, Sculellaria barbata, ground beeltle and borneol of the powder of promoting blood flow and relieve pain qualitatively with thin-layer chromatography(TLC).2. The method of investigating heavy metal and arsenic salt:Investigate heavy metal with Nuss colorimetric method, while measure the limit quantity of arsenic salt with Gutzeit.3. Observation index:In qualitative identification, comparing supplied experimental chromatograph with contrast medicinal materials and contrast chromatograph, whether there are the same spots. In Heavy metal investigation, according to color depth of sample Nuss colorimetric tube and standard lead Nuss colorimetric tube, determine the limit content of heavy metals in samples. In arsenic salt investigation, comparing sample arsenic spots with standard arsenic spots, determine the limit content of arsenic salt in the samples.The analgesics mechanism and basic application of the microns powder used for invigorating the circulation of blood and analgesia for the treatment by ultrasonic transversal drug delivery targeting conductancel.How to made acute tissue injury of soft model and experimental group:Taking a big white New Zealand rabbits, then choose rabbit hind leg to be marked, About 4 to7cm from the nest, We use self-made staff then caused it in free fall on the marked station, Building a successful representation model if without fracture confirmed. Taking 24 acute tissue injuries of soft model rabbits, random divided into four groups and each group only 6. As follows:Model control (group 1), Ultrasonic conductance blank gel strips into percutaneous penetrate (group2), Ultrasonic conductance microns the powder of promoting blood flow and relieve pain into percutaneous penetrate (group 3), and micron the powder of promoting blood flow and relieve pain topical (group 4).2. the preparation process of Micron promote blood flow analgesia scattered gel strips:①Made all the crude drugs crushing into sifting.②Get prescription effective components from ethanol-extraction, then making into dry paste powder.③With inert atmosphere protection of high-energy ball mill equipment system, we made notoginseng powder into microns.④Add microns notoginseng and borneol into dry powder, mixed uniformity, Join into blank gel strips (gel base within the hydration filling, penetration enhancers agent, PVA, etc.), gel strips made compound Chinese medicine usage of ointment composition evenly mixed fabric woven material suction amend the hydration gel matrix in filling for micron, namely Micron promote blood flow analgesia scattered gel strips3.The therapy of experiment:Each animal (except group 1) after made acute tissue injury of soft model 2h later, Giving related treatment,As follows:Article 2, 3 groups will get strips posted on head, the head was fixed in injury treatment of soft tissue place. With constraints fixed therapeutic part, treatment of strength and parameter selected:the hole 2, ultrasonic 2, intermediate frequency 1, time 15min, After treatment, keep removing treatment head,2 times patch 2h/day and continuous 4d. The fourth groups according to administration dosage topical in trouble place 2 times per day, and fixed reserves, continuous drug 2h,4d.4. Skin medication acute toxicity tests and skin irritation test methods:skin medication acute toxicity tests:randomly take 15 rabbits into five groups, namely: three rabbits in a group of control group only (1mL), twelve rabbits besmear physiological saline in every four groups (whole skin group and damaged skin group, each two doses). Low dosage group for clinical use the dosage of 1 mg, the dosage of quantity high-dosage groups is low dose group 2-3 times. Skin irritation test:take rabbits were randomly divided into two groups, three is complete skin, and the other three is damaged skin. Respectively in hair removal,2days after the biooxyrate content posted on hair removal, and fixed them with no excitant. Each animal is feeding in single cage. Applying medicine after 24h, uncover gauze with warm wash to remove of residual biooxyrate drugs.5.Index measurement methods, sanqi (panax notoginseng) powder microns in scanning electron microscopy (sem) downside of particle before contrasts; Measuring and calculating four groups of daily variation, continuous survey 4d; before Exsanguinations measurement was made, made after die mould 2,3,4, white cell of margin veins, in rabbit ears numerical was changed; After the same strike 4d take animal parts, equal sections of tissue biopsies, paraffin embedding militiamen dyeing, Trypan-blue and scores histologist observation; Observe the tested objects 1,24,48,72 h removal from the daily observation and record the 4d, animal weight, skin and hair, eyes and mucous membrane changes, breathing, central nervous system, limb activity level other poisoning performance.In macroscopic observation,records whether it is erythematic and edema areas such as local reaction after removed 1,24,48,72 h daub tested objects without. And according to skin irritant reactions rating criteria rating, and record the time scoring values.Statistical methodsMeasurement data of the experimental data are expressed by Mean±standard deviation(x±s). Use SPSS 13.0 software to analyze the statistical data. Compare the overall mean of two independent samples with the independent sample t-test. Analyze L9 (34) orthogonal design material with orthogonal design anova. Compare multi-group measurement data with one-way ANOVA. Compare swelling area of soft tissue contusion place in the group at different time points and white blood cells of peripheral blood with repetendum measurement variance analysis. Inspection standard is 0.05ResultStudy on extraction of the powder of connecting bones and reducing swelling by orthogonal design method1.Methodology investigation results:Standard curve equation is Y= 72.008X+18.041,r=0.9994(n=6). The results show that the naringin in the range of 0.135-1.620 mg/L has good linear relationship. RSD value measured by Precision experiment is 1.46%, while by stability test is 1.04%.The result show that the selected product solution within 24 hours is basically stable. The result of recovery rate about add kinds:recovery rate is 102.46% and RSD value is 2.31.2. Investigation results of alcohol absorption rate:The average of alcohol absorption rate is 265.67 mL/g and RSD is 1.48%, which show that alcohol extraction in the first time should be added more than three times of the corresponding volume fraction amount of the ethanol.3. Comparison the results of Permeability Lu method with refluxing extraction: The average naringin level of reflux extraction result is 238.23±7.00mg/g, RSD 5.86%; The average naringin level of Permeability Lu method result is 238.18±5.86 mg/g,RSD 7%; Two groups by two independent sample mean t test results to a statistically significant difference (t=-8.039, P=0.000).Permeability Lu method can be thought to have significant difference significance with permeability refluxing extraction about naringin volume, Reference mean there, the results show that the best methods is reflux method ethanol-extraction.4. Comparison about the results of Ethanol-extraction times:the third of alcohol extraction rate and naringin volume accounted for extraction volume ratio were greater than 10%, If extract drug ingredients completely, we should be ethanol-extraction 3 times.5. The results of orthogonal experimental:The each level factors average of ethanol volume fraction (A) is A1=87.197, A2=91.789, A3=81.367; each respective mean level of add alcohol quantity (factors B) is B,=90.607, B2=91.828, B3=77.917; the results of extracting time (factors C) is C1=86.282, C2=87.241, C3=86.829.The results of variance analysis show ethanol volume fraction, add alcohol quantity have significant meaning (FA=19.738,PA=0.048, FB=19.738,PB=0.023), But extracting time factors do not have significant (Fc=0.167,Pc=0.857). Based on the results, we know the combination of A2B2C2 is the best optimizing selection, The best casual blood extraction pain is:herbs plus ethanol extracted 3 times,1st with 6 times the amount of the volume fraction of 65% ethanol extract of 2h, the 2nd plus 5 times the volume fraction of 65% ethanol extract 1.5h,3rd plus 5 times the volume fraction of 65% alcohol extract 1.5h.The conformity quality standards of the medicinal powder used for invigorating the circulation of blood and analgesia1. Thin Layer Chromatography (TLC) was used to identify the qualitative results of compound ingredients:①Drynariae:With the limulus'chromatography, the control medicinal reference substance appeared the same yellow fluorescent spots in the corresponding position. On the negative control group, we can't see the same fluorescent spots in the corresponding position.②comparing with the chromatography of the reference substance, the limulus'chromatography the same color on the fluorescent spots in the corresponding position. On the negative control group, we can't see the same fluorescent spots in the corresponding position.③saffron:With chromatography in the reference drug of the corresponding position, the limulus'chromatography was the same pink color spots by the daylight.④barbata: With the chromatography in the reference drug of the corresponding position, the limulus'chromatography was the same color on the fluorescent spots by the daylight. On the negative control group, we can't see the same fluorescent spots in the corresponding position.⑤Eupolyphaga:In comparison with the chromatography of the Control medicinal materials, the limulus appeared the same yellow fluorescent spots in the corresponding position.⑥borneol:Along with the chromatography of the Control medicinal materials, the limulus appeared the same red spots in the corresponding position. On the negative control group, we can't see the same fluorescent spots in the corresponding position.2. The results of heavy metals'study:the color of the test samples'Nessler colorimetric tubes was lighter than the standard. So it can predicate the heavy metals were less than lOppm in the samples.3. The results of arsenic salts:comparing with the standard, the color of the tested sample spots was much lighter. So it can predicate the arsenic salts were less than lppm in the samples.The analgesics mechanism and basic application of the microns powder used for invigorating the circulation of blood and analgesia for the treatment by ultrasonic transversal drug delivery targeting conductance1. The SEM results of microns Sanchi Powder:based on the high energy ball milling method preparing for microns Sanchi Powder, we could find the powder keep fresh color and strong smell by observing the powder which got out every half hour. After the powder grinded 8 hours, it gradually turned to black and smell fades. It might be grinded too long to carbonize the Powder. So we chose the best ball-milling time with 8 hours. After the powder grinded 8 hours for turning the large particles to the lamellar shape, it asked the lamellar thickness less than 1.5μm and the large diameter about 13μm for reaching the requirement of micron.2. The injury results of the rabbits with acute swelling of the soft tissues:after modeling, the injury soft tissues where occurred swelling, skin ecchymosed, and subcutaneous bleeding. The most obviously is at 24 hours, but the turgescence then gradually reduced. The four groups all adopted the difference between the daily area of the swelling and the 1st day to do one-way analysis of variance. The results showed differences among the four groups were statistically significant (F=27.793, P<0.01). the group of ultrasonic transdermal the microns powder delivery targeting conductance, the group of posting the microns powder, and the ultrasound group all reduced more swelling area than the soft tissue injury group at 2,3,4 days. The differences were statistically significant (P<0.01), and the group of ultrasonic transdermal the microns powder delivery targeting conductance reduced swelling most obviously.3. The results about the impact of changes in peripheral white blood cell count after the rabbits with acute swelling of the soft tissues:the increasing number of neutrophil suggested the inflammatory response, comparing with the former modeling group, the peripheral white blood cell count in four groups all obviously increased (P<0.01), indicating that the inflammatory response caused by acute soft tissue injury, the four groups measured the peripheral white blood cell count at different time points to do one-way analysis of variance. The results showed differences among the four groups were statistically significant (F=19.984, P<0.01). comparing with the soft tissue injury group, the group of ultrasonic transversal the microns powder delivery targeting conductance, the group of posting the microns powder, and the ultrasound group all obviously reduced the localized white blood cell count (P<0.01), and the group of ultrasonic transversal the microns powder delivery targeting conductance reduced most obviously.4. The outcomes of the pathological examination and scores of the rabbits:after treatment for 4 days based on histological observation, the results of statistical analysis of variance are shown:as a whole, the means between the groups were significantly different (F=14.182, P<0.01). a comparison of the soft tissue injury group, the group of ultrasonic transversal the microns powder delivery targeting conductance, the group of posting the microns powder, and the ultrasound group all obviously reduced the inflammation response(P<0.01), promoting collagen synthesis of local injury, muscle tissue repair. And the results about the group of ultrasonic transdermal the microns powder delivery targeting conductance ameliorated most obviously. the histological of the four groups at four days under microscope(icon):①Group one:the hit place of the rabbits muscles observed a large number of ruptured muscle cells and extensive bleeding. a large localized inflammatory exudates and necrosis can be observed with ill-defined. The rich granulation tissue observed in peripheral, which mainly composed of inflammatory cells, fibroblasts and capillary structure. Multinucleated my oblasts also can be observed around the ruptured muscle cells with a large number of acute and chronic inflammatory cells.②Group two: The inflammatory exudates and necrosis had completely disappeared. The granulation tissue is also reduced, replaced by large collagen fibers. The ruptured places were basically fixed, but the arrangement of the repaired muscle cells at a little disorder.③Group three:The hit places of the muscles were basically fixed with the muscle cells in good order. The bundle of collagen fibers can be examined in the local, with the infiltration of inflammatory cells between the muscle cells.④Group four: Unobserving exudation and necrosis, the area of granulation tissue compared with model group were significantly reduced and proliferation of muscle cells.⑤Group five (normal rabbit muscles):The form of the rabbits was rather consistent. The muscle cells were closely packed bundle by parallel, elongated spindle cells, abundant cytoplasm, and the nuclear was the short rod. 5.the acute toxicity test of the micron Chinese medical gel patch on the skin: after removing the test substance from 1 hour to 7 hours, we could observed that the experimental animals moved without restricting, feeding normal, normal applicator at the skin shiny hair, no change of eyes and mucous membranes, no abnormal breathing, no central nervous system abnormalities, no systemic toxic reactions in rabbits, and no animal deaths occurred, indicating that the micron bulk for blood circulation and analgesic used at low and high doses without any external time acute toxicity.6. The skin irritation test of the micron Chinese medical gel patch:the micron bulk for blood circulation and analgesic posted only a dose. There were not observed erythema, edema, other abnormal conditions, and stimulation index less than 0.4. Shows a micro dose of Chinese medicine gel patch intact skin of rabbits had no significant skin irritation and damage. Although small pieces of broken skin and blood of callus formation has 4 days off, paste dispensing area with no significant difference between control areas, but due to scratches, not drug induced.ConclusionThe best casual blood extraction pain is:herbs plus ethanol extracted 3 times,1st with 6 times the amount of the volume fraction of 65% ethanol extract of 2h, the 2nd plus 5 times the volume fraction of 65% ethanol extract 1.5h,3rd plus 5 times the volume fraction of 65% alcohol extract 1.5h.TLC can be detected Drynaria, notoginseng, safflower, Scutellaria barbata, and borneol Eupolyphaga characteristic spots. Salt content of arsenic in traditional Chinese medicine compound limits≤1ppm, limits of heavy metal≤10ppm, reached the limits of heavy metals and arsenic salts.Percutaneous penetration of ultrasound conductance blood micron bulk analgesic treatment of acute soft tissue injury in rabbits with a good anti-inflammatory swelling, reduce inflammation, promote the inflammatory exudates absorption and the repair of soft tissue injury. Ultrasound was superior to physical therapy and micro blood conductance with external application of pain, and no stimulation to skin safety, safety of the new medicine is a gel patch. |
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