| OBJECTSTo deeply study effects of corn germ oil on reducing lipid auxiliarily and enhancing immunity, as well as it's safety. To provide a reliable theoretical basis for developing of corn germ oil in new functional food and other related industries such as drugs, foods and health products.METHODS1 Toxicology experiments(1)Acute toxicity test 20mice weighting 18～22g, male and female each half, were given 20ml/kg·bw (density 0.500 g/ml) corn germ oil for twice, total infected dose was 20.0g/Kg·bw, and then they were observed for 14days, the performance and mortality were recorded.(2)Hereditary toxicity tests①The mutation of salmonella test (Ames test) Experimental strains are histidine defected rat typhoid salmonella,TA97,TA98, TA100,TA102, which meet the requirements. The vitro activating systems was S-9mixture which was rat liver homogenate that induced by polychlorinated biphenyls(PCBs). According to the toxicity testing results, five doses (8,40,200, 1000,5000μg/dish) groups were set up, and also the spontaneous back to become group, solvent control group and positive control group were set up. According to the flat mixing method, experiments were done with and without S-9 mixture, and three dishes were set up in each group. If back to become number of bacterial colonies about test sample is twice more than the spontaneous number, and there was a dose -response relationship, it was judged to be positive. (②Micronucleus Test of Mice Bone Marrow Polychromatic Erythrocytes 50 mice weighting 25～30g, male and female each half, were randomly divided into 5 groups. Low, middle, high dose groups were given different doses of 0.167,0.333,0.667g/ml (infected doses were 2.5, 5.0,10.0 g/Kg·bw). The negative control group were given peanut oil, and the positive control group were given cyclophosphamide (40mg/Kg·bw). Mice were gavage fed twice with an interval of 24h between each time, and the feeding amount was 15ml/Kg·bw.6hs after the second fed, mice were killed and the conventional production was made. Each animal was found 1000 polychromatic erythrocytes (PCE) under microscope, the number of micronucleus cells was recorded, and then micronucleus rate was calculated (micronucleus cells/PCE cell, expressed in thousands of points).200 PCE were observed, the number of normochromatic erythrocytes (NCE) and the PCE/NCE ratio was calculated.③Mouse sperm deformity experiment 25 male mice weighing 27～31g, grouping and processing like②, gastric irrigated once a day continuously for five days.35d after the first gastric irrigation, animals were killed and conventional production was made. Each animal was counted 1,000 sperms with structural integrity, the type and amount of distortion recorded, sperm abnormality rate calculated (expressed in percentage).(3)30d feeding test in rats 80 weaning rats, weighting 70～90g, male and female each half, were randomly divided into negative control group with peanut oil and high, medium and low dose groups (20,10,5 ml/Kg·bw, equivalent to the recommended dose of 60,30,15 times). Gastric gavage volume was 20ml/Kg·bw, once a day for 30days. Tail vein blood was taken on the thirtieth day, and then blood test was examined. On the thirty-first day, the abdominal arterial blood was taken after fasting for 16 hours, and then biochemical tests was examined. The pathological examination on organs was also conducted at the same time.2 Enhancing immunity experiments 192 female KM mice were randomly divided into 4 immune groups, and the 48 mice in each group was then divided into 4 groups (peanut oil negative control group and low, medium and high dose groups),each small group has 12 mice. Low, medium and high dose groups were given different doses of 1.67,3.33,10.00 mL/kg-bw, by gastric gavage for a consecutive 30 days, and then the indexes were tested.3 Reducing Lipid auxiliarily experiments3.1 Animal experiment According to the lipid level,60 male adult SD rats were randomly divided into negative control group, high fat control group, positive control group, low, medium and high dose groups(1.67,3.33,10.00ml/Kg·bw). Negative control group were given normal standard diet, high fat control group was given high fat diet and peanut oil, positive control group was given high fat diet while fed 0.05g/Kg·bw fenofibrate solution, and the other three groups were given high fat diet and different doses of corn germ oil every day. The changes of lipid levels in rats were observed after 35 days.3.2 Crowd experiment (1)Human feeding experiment 108 volunteers who met the requirements such as pure hyperlipidemia and signed the informed consent certificates were chosen to take part in this experiment. They were randomly divided into experimental group (56) and negative control group (52), experimental group was given corn germ oil instead of lunch and dinner cooking oil (10ml per meal), negative control group was given nothing. All volunteers keep normal diet and lifestyle for 45days. Related indexes of all volunteers were tested before and after this experiment.(2)Dietary and lifestyle questionnaire Dietary survey was conducted using the 24h review questionnaire continuously for 3days. Dietary and lifestyle habits survey was conducted using self-designed questionnaire, by systematically trained inspectors. The basic information, eating behavior and life style of the participants were recorded.RESULTS1 Toxicology Experiments(1)Acute toxicity test The animals have no poisoning performance or death in 14days. MTD results showed that the biooxyrate content on both male and female mice were more than 20.0g/Kg·bw. According to acute toxicity grading criteria, this sample is non-toxic substance actually.(2)Hereditary toxicity test①The mutation of salmonella test (Ames test) In two experiments, the back to become number of bacterial colonies in each dose group was no more than the spontaneous number, and no dose-response relationship was observed either. This indicates that samples with and without S-9 have no genetic toxicity effect on rat typhoid salmonella TA97, TA98, TA100, TA102.②Micronucleus Test of Mice Bone Marrow Polychromatic Erythrocytes Compared with the negative control group, the micronucleus rate in each experimental group has no significant difference (P> 0.05); but positive control group was significantly different (P<0.01). This illustrates that this sample does not induce the micronucleus mutation of rat's bone marrow polychromatic erythrocytes.③Mouse sperm deformity experiment Compared with the negative control group, mice sperm deformity rate in each experimental group has no significant difference (P>0.05);but positive control group has significantly different(P<0.01).That shows this sample does not induce sperm abnormality in mice.(3)30d feeding test in rats The liver, kidney, stomach, intestine, spleen and ovary/ testis of rats were generally observed in experimental groups, no obvious abnormalities was observed. In histological examination, a few animal specimens in the control group and high dose group appeared light pathological change, but the distribution have no differences between groups. Considering the quality of the animals, the experimental group has significant pathological changes.2 Enhancing immunity experiments Compared with the negative control group, low and high dose groups can increase delayed anaphylactic reaction of mice(P<0.05, P<0.01); low, medium and high dose groups all can increase the number of lymphocyte B and phagocytes in abdominal cavity of mice(P<0.01); high dose group can increase mononuclear-macrophages carbon clearance capacity of mice (P<0.01); and medium and high dose groups can intensify the activity of NK cells of mice(P<0.01,P<0.05)3 Reducing Lipid auxiliarily experiments3.1 Animal experiment In the whole experiment, the growth and activities of animals were normal in all groups. Compared with the negative control group, the body weight of animals in each group has no significant difference (p> 0.05).Compared with model group, total cholesterol(TC), triglyceride (TG) level in positive control group, low, medium and high dose groups are lower (p<0.01), and high density lipoprotein cholesterol(HDL-C) level are higher (p<0.01). Compared with model group, the declining percentage of TC in positive control group, low, medium and high dose groups are 18.77%,10.77%,16.92%,17.54%, respectively, the declining percentage of TG are 27.54%,9.84%,14.10%,22.62% respectively. The rising vale of HDL-C are 0.66,0.28,0.34,0.38 mmol/L, respectively.3.2 Crowd experiment(1)Human feeding experiment During the experiment, the mental state, diet, sleep, feces and urine condition of all participants did not show any abnormality. Before and after trial, routine blood test, blood biochemistry indexes and liver, kidney function indexes are all in normal range. Compared to the level before test or in control group, serum TC level in experimental group were decreased by 14.41% and 14.407% respectively, and the decreases were statistically significant (P<0.01); the serum TG level were decreased by 26.50% and 37.61% respectively, which is also significantly (P<0.01). Efficiency of serum TC, TG in experimental group were 69.64% and 58.93% respectively, total effective rate was 64.29%, which were significantly different from the control group (P<0.05). The increase of HDL-C level in experimental group and control group were 0.07 and 0.05 mmol/L respectively, but the difference was not significant (P>0.05).(2)Questionnaire survey①generally information:The highest proportion of education is undergraduate (31%); the largest proportion of career is professional and technical staff (53%); most family income are within 2500-5000 Yuan. Compared with the control group, the distribution of education, occupation, family income, food expenditure per month in experimental group has no difference (P>0.05). The minimum value is of BMI is 16.73, the maximum value was 36.73, mean value was 25.50±36.22. Thin people accounts for 2%, overweight 43%, and obese 12%.②Habits of life:81.9% people consider their life orderly; 76.0% people sleep five to eight hours every day; 81.9% people consider their sleep orderly; 54.5% people almost has no healthy products intake.③Diets information:71.6% people eat breakfast everyday; 76.2% people take meals with the same quantity; 82.9% people doesn't picky on food; 50.4% people have over-eating habit. Compared to food Pagoda, the per capita consumption of oil, salt were 32.71g,8.02g respectively, which is higher than recommended; the per capita consumption of vegetables, fruits, fish, milk and milk products were243.83g,107 g,25.95 g,36.11 g respectively, which was lower than recommended. There is 17.76% participants smoking; 71.23% drinking wines; 50.93% drinking tea; and limited participants drinking coffee or soft drinks. Mean intake of energy was 2037.66±1084.80kcal; the energy production ratios of fat, carbohydrate, protein were 23.69%,60.84%,16.37respectively. Compared with RNI/AI, the intake of zinc(Zn), calcium(Ca), selenium(Se), chromium(Cr),Vitamin A(VA),Vitamin B-1(VB1), Vitamin B-2(VB2), Vitamin (VE) were lower; the level of ferrum(Fe) was 19.5±5.63 mg/d, which was slightly higher.④The effect of diets and habits for TG:The shorter two meal time intervals, the more taking of snack, the higher of BMI, the more overeat and overdrink, the higher of TG is.CONCLUSIONS1. The results of toxicity tests show that all indexes were within normal range in this experiment, which can prove that this corn germ oil is safe.2. Corn germ oil can increase delayed anaphylactic reaction, the activity of NK cells, the number of lymphocyte B and the phagocytes in abdominal cavity, mononuclear-macrophages carbon clearance capacity of mice. According to "technical specifications for health food inspection and evaluation ", this corn germ oil can enhance immunity.3. Different doses of corn germ oil can reduce TC and TG levels, and increase HDL-C level in rats. Human feeding test experiment showed that corn germ oil can reduce TC and TG levels, and increase HDL-C levels. According to "technical specifications for health food inspection and evaluation ", corn germ oil can reduce lipid auxiliarily. 4. The results of the questionnaire survey showed that the hyperlipidemia crowd generally has higher BMI, normal drinking amount; and higher intakes of oil and salt than recommended; and lower intake of vegetables, fruits, fish, milk and milk products than recommended. Compared with RNI/AI, the intake of zinc, calcium, selenium, chromium, VA, VB-1, VB-2, VE are lower, the intake of iron is slightly higher. The shorter two meal time intervals, the more intake of snack, the higher BMI, the more overeat and overdrink, the higher level of TG is. |
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