while the existing acupuncture treatment incorporates physical stimulation of associated meridians and acupoints , pharmacopuncture adds chemical stimulation to the existing acupuncture treatment . hominis placenta is a rich resource of various bioactive substances , such as polydeoxyribonucleotides , ribonucleic acid ( rna ) , deoxyribonucleic acid ( dna ) , peptides , amino acids , enzymes and trace elements . hominis placenta pharmacopuncture is generally used these days , but toxicity tests have never been done . all experiments in this research were conducted at biotoxtech ( chungwon , korea ) , an authorized institution for non clinical studies , under the regulations of good laboratory practice ( glp ) . the hominis placenta pharmacopuncture extract was prepared by adhering to korea - good manufacturing practice ( k - gmp ) in a clean room in a laboratory at the korean pharmacopuncture institute . the animals used in this study were 6 week old sprague - dawley ( sd ) rats . the reason sd rats were chosen is that they have generally been used in stability tests of medicine , so the data obtained in this study should be easily compared with many other databases . the mean weights of the rats were 185.1 - 205.9 g and 149.7 - 167.2 g for the male and the female rats , respectively . for all animals , a visual inspection was done , and all animals were weighed using a cp3202s system scale ( sartorius , germany ) . after 7 days of acclimatization , the rats general symptoms and changes in weight were observed . the temperature of the lab was 21.1 - 24.1c , and the humidity was 40.7% - 64.5% . sufficient food ( teklad certified irradiated global 18% protein rodent diet 2918c ) and ultra violet ( uv)-filtered water were provided . the animals were randomly distributed into 4 groups of 5 male and 5 female rats per group ( table 1 ) : the control , low dose , mid dose and high dose groups . the expected dose of hominis placenta pharmacopuncture for clinical applications is 1.0 ml . in a pilot study b13473p ) , 0.5 ml / animal of hominis placenta pharmacopuncture was injected into each male and female rat ; no deaths were observed . from this result , the doses for hominis placenta pharmacopuncture were set as follows : animals in group 1 ( g1 , the control group ) were injected with 0 ml / animal of pharmacopuncture and 0.5 ml / animal of normal saline solution ( choongwae pharma corp . , korea ) , animals in group 2 ( g2 , the low dose group ) were injected with 0.125 ml / animal of pharmacopuncture , animals in group 3 ( g3 , the mid dose group ) were injected with 0.25 ml / animal of pharmacopuncture , and animals in group 4 ( g4 , the high dose group ) were injected with 0.5 ml / animal of pharmacopuncture . using a disposable syringe ( 1 ml , 26 g ) , this study was conducted under the approval of the institutional animal ethics committee of biotoxtech . the general symptoms ( types of toxic symptoms , revealing times , recovery times , etc . ) and the mortality were examined 30 minutes , and 1 , 2 , 4 and 6 hours after the injection on the day of dosing ( day 0 ) . from the 1 day to the 14 day of treatment the body weights were measured immediately before treatment and at 3 , 7 and 14 days after injections . the rats were anesthetized by using isoflurane , and blood samples were taken from the abdominal aorta ( 15 days after injection ) . an automatic hematology analyzer ( advia 2120i , siemens , germany ) two ml blood samples were centrifuged for the blood coagulation test ( 3,000 rpm , 10 minutes ) . coagulation test results were measured by using an automated coagulation analyzer ( coapresta 2000 , sekisui , japan ) ( table 3 ) . for the biochemical tests , the blood remaining after carrying out the hematological tests was centrifuged at 3,000 rpm for 10 minutes , and the serum was collected . biochemical test results were measured by using an automatic analyzer ( 7180 , hitachi , japan ) and an electrolyte analyzer ( avl9181 , roche , germany ) ( table 4 ) . after the observations , organs and tissues from the entire bodies of all surviving animals were visually inspected . routine histological methods , like trimming , dehydration , and paraffin embedding , were conducted on the fixed organs and tissues . all the results from the experiments were analyzed by using statistical analysis system ( sas ) software ( version 9.3 , sas institute inc . a bartlett test was conducted to evaluate the homogeneity of the variance and the significance . the one - way analysis of variation ( anova ) test was conducted when homogeneity of the variance was recognized , and the dunnett 's t - test was conducted posthoc . the kruskal - wallis test was conducted when heterogeneity of the variance was recognized , and the steel test was conducted post - hoc . during the observation , no deaths or abnormal symptoms occurred in any of the rats in the experimental ( 0.125 , 0.25 , 0.5 ml / animal ) and the control groups . no significant changes in body weight were observed in any of the groups ( table 5 , figs . no significant changes in the hematological test results were observed in any of the groups , except that the blood urea nitrogen ( bun ) of g2 males and the total cholesterol ( t - chol ) of g3 males showed statistically significant changes ; however , those results had no toxicological meaning because they were not dose dependent ( table 6 ) . no significant changes in the biochemical test results were observed in any of the groups ( table 7 ) , and no abnormalities were noted when visual inspections of all the animals were conducted . no significant changes were noted in the local tolerance test on the injection sites , except for g3 males ; however , that result was due to spontaneous generation and had no toxicological meaning because it was not dose dependent . pharmacopuncture and herbal acupuncture were introduced to optimize the benefits of acupuncture and herbal medicine by injecting minute quantities of herbs , medicines , blood , oxygen , and allergens into acupoints . hominis placenta pharmacopuncture is used generally in clinics , and many studies on the effects of hominis placenta pharmacopuncture have been done . it has been reported to have effects on sleep pattern disturbance , bell s palsy [ 9 , 10 ] , and dysmenorrheal [ 11 , 12 ] . in addition , it has been reported to have effects on kidneys and livers in rats intoxicated by using hgcl2 . although it is used in clinics , safety studies on hominis placenta pharmacopuncture are insufficient , sd rats have been widely used in safety tests of drugs because they have consistent reactions to drugs . this study was performed to provide objective safety data for hominis placenta pharmacopuncture . during the observation , hematologically , some changes in the g2 and the g3 male group were observed , but those changes had no clinical or toxicological meaning . no significant change in the biochemical test results were observed in any of the groups . visual inspection after necropsy showed no visual abnormalities in the male and the female animals in any groups . histopathological tests on the injected parts showed a cell infiltration in the g3 male group ; however , the result was due to spontaneous generation and had no toxicological meaning . we need to study the acute and the chronic side effects of hominis placenta pharmacopuncture and its relations to the capacity reaction more . also , the toxicity and the safety of hominis placenta pharmacopuncture for use on humans must be investigated . the objective of this study was to analyze the single - dose toxicity of hominis placenta pharmacopuncture extracts . all experiments were conducted under the regulations of glp at biotoxtech , an institution authorized to perform non clinical studies . the results showed that administration of 0.5 ml / animal of hominis placenta pharmacopuncture extracts did not cause any changes in weight or any mortalities in sd rats . ; hct , hematocrit ; mcv , mean corpuscular cell volume ; mch , mean corpuscular hemoglobin ; mchc , mean corpuscular cell hemoglobin concentration ; plt , platelet ; reti , reticulocytes ; wbc , white blood cell ; neu , neutrophils ; lym , lymphocytes ; mono , monocytes ; eos , eosinophils ; baso , basophils ; pt , prothrombin time ; aptt , active partial thromboplastin time .
objectives : this study was performed to analyze the toxicity and to find the lethal dose of the test substance hominis placenta pharmacopuncture when used as a single - dose in 6 week old , male and female sprague - dawley ( sd ) rats.methods:all experiments were conducted at biotoxtech ( chungwon , korea ) , an institution authorized to perform non clinical studies , under the regulations of good laboratory practice ( glp ) . 
 sd rats were chosen for the pilot study . 
 doses of hominis placenta pharmacopuncture extracts , 0.125 , 0.25 and 0.5 ml , were administered to the experimental group , and 0.5 ml doses of normal saline solution were administered to the control group . 
 this study was conducted under the approval of the institutional animal ethics committee.results:no deaths or abnormalities occurred in any of the groups . 
 also , no significant changes in body weights were observed among the groups , and no significant differences in hematology / biochemistry , necropsy , and histopathology results were noted . 
 hematologically , some changes in the male rats in two experimental groups were observed , but those changes had no clinical or toxicological meaning because they were not dose dependent . 
 histopathological tests on the injected parts showed cell infiltration in the male rats in one of the experimental groups ; however , that result was due to spontaneous generation and had no toxicological meaning . therefore , this study showed that hominis placenta pharmacopuncture had no effect on the injected parts in terms of clinical signs , body weight , hematology , clinical chemistry , and necropsy.conclusion:as a result of single - dose tests of the test substance hominis placenta pharmacopuncture in 4 groups of rats , the lethal dose for both males and females exceeded 0.5 ml / animal . therefore , the above findings suggest that treatment with hominis placenta pharmacopuncture is relatively safe . 
 further studies on this subject are needed .