| Background:Because of the rapid developing in economy and changing food habits, there was an increasing trend yearly at colorectal cancer (CRC) incidence in our country. It is an effectively preventive method to tumor by ingesting natural food that has anticancer effect. At present, the relationship of resistant starch (RS) and colon cancer is being paid close attention at home and abroad. A number of food, such as cereals, legumes, tubers, green banana have been shown to contain various amounts of RS. Few animal experiments is reported about RS preventing colorectal cancer, especially concerning the effect of RS on the precancerous leisions of colorectal cancer.Objective:1. To study the effect of processing to RS content by comparing the results of RS content in raw food materials and cooked food products.2. To report the RS contents of typical starchy foods used frequently by Chinese population.3. Based on the RS values measured in the present study and food consumption data obtained from the dietary survey, RS intake (g/per capita/day) was estimated by individual.4. The key sources of RS in Chinese foods were also identified. This will supply the basic science information for the study of dietary guidance and colon cancer prevention.5. We use the model of AOM-induced colorectal cancer in Wistar rat. This study was designed to determine the potential inhibitory influence of RS at different doses through the observation of the rat growth test, the small intestine movement test and metabolic test. And it was also designed to investigate the influence of a premalignant lesion by testing the number of ACF, one of the early landmarks of colorectal cancer, and some other biomarkers, such as PCNA, AgNORs, Bcl-2 and Bax. The results will provide evidence for early diagnosis and treatment of colorectal cancer, and be of great importance for guiding Chinese diets and early preventing colorectal cancer.Methods:1. Determining of resistant starch:to collect the typical starchy foods (raw material and cooked food) used frequently by Chinese population and to determine RS content of before and after processing by an in vitro method that mimicked gastrointestinal conditions.2. Dietary survey:in order to estimate the daily RS intake in the Chinese population, food consumption data from a dietary survey was used. Population was Choosed randomly from city and country (Shandong, Henan, China) by the method of cluster sampling. All subjects were asked to complete an estimated dietary record for 3 consecutive days of food items and liquids each individual consumed.3. Estimating the Chinese intake of RS:based on the RS values measured in the present study and food consumption data (g/per capita/day) obtained from the dietary survey, RS intake was estimated by individual. The main sources of RS in foods were also explained. RS intake was also divided according to the food from which it originated (i.e. tuber, legume or cereals). In addition, data reported from references were used for estamiting the intake of different area.4. Effects of resistant starch on intestine function of AOM-induced colorectal cancer in ratsMale Wistar rats were randomly divided into 5 groups:negative control group, positive control group and three different resistant starch (RS) added groups with the ratio of 7.6%,15.2% and 22.8% (n= 10 each group). For inducing colonic tumorigenesis, rats received chemical carcinogen (AOM) injections by intraperitoneal once a week for two consecutive weeks. Animals in different RS added groups received the corresponding doses of RS while rats in the positive control group and the negative control group received ordinary feed during the whole study. This study was designed to determine the protective effect on intestine of RS at different doses through the observation of the small intestine movement test and metabolic test. Fecal pH and fecal output were measured in the 5th weekend and the intestinal transit time measurement was carried out in the7th weekend.Then comparing them among groups. And it was also designed to investigate the influence of weight by testing the rat growth.5. The study of prophylactic effect of resistant starch on precancerous lesions of AOM-induced colorectal cancer in ratsMale Wistar rats were randomly divided into 5 groups:negative control group, positive control group and three different resistant starch (RS) added groups with the ratio of 7.6%,15.2% and 22.8%.For inducing colonic tumorigenesis, rats received chemical carcinogen (AOM) injections by intraperitoneal once a week for two consecutive weeks. Animals in different RS added groups received the corresponding doses of RS while rats in the positive control group and the negative control group received ordinary feed during the whole study. At 13 weeks after the first injection, all of the rats were killed. Colons of rats were removed and the total numbers of ACF as well as the number of crypts per focus were scored using a light microscope at low magnification after staining with 0.5% methylene blue solution. Some other biomarkers (such as PCNA, AgNORs, Bcl-2 and Bax) were also determined by immunohistochemical method and compared among groups to investigate the effect of RS on the precancerous lesion of colon cancer.Results:1. Resistant starch content in food:the contents of RS largely varied with plant source. In general, tubers, tuber vegetables and legumes had more RS than cereals. Among the selected samples of raw material, potato showed high amount of RS. The pure potato starch and cassava starch had the highest RS content. Among the food products, roasted and cooled potato had the highest RS content. The following were sweet potato starch noodle, mung bean starch noodle and Sweet potato starch jelly sheet. The content of RS in cooked foods was lower than raw food material in general. Deep-fried and roasted foods have more RS than general braised foods. The content of RS in cold noodle was the highest in finely processed food.The following were cake, walnut cake, rice noodle, potatochips, Lima bean with special hot sauce and fried dough twist.2. Chinese intake of resistant starch:the daily RS intake of Shandong population was estimated to be 14.9±9.1g, which were 16.2±8.8g/d in male and 14.2±9.3g/d in female. And the value was 14.1±7.5g/d in population who live in the city and 15.9±10.7g/d in population who live in the country. The range of intake was 2.9g～49.0g/d. The daily RS intake of Henan population was estimated to be 17.1±6.5g, which were 18.0±7.8g/d in male and 15.4±8.8g/d in female. And the value was 16.1±5.3g/d in population who live in the city and 17.9±9.6g/d in population who live in the country. The range of intake was 4.1g～63.2g/d. Obviously, RS intake varied considerably among individuals. The average intake of Chinese population was 9.7g/d.3. The key RS sources in the Chinese diet:the key RS sources in the Chinese diet are wheat products, rice products and starch products. Other important sources of RS are tubers and legume products, followed by corn products and other cereals, nuts and seeds. Vegetables and banana contribute fewer amounts of RS to the Chinese diet.4. Body weight alteration of rat:During the whole study, each group of rats grew well and weight was increasing. The body weights of rats in RS groups were lower than control and there was a dose-dependent in lower weight and high RS level. Compared with the positive control group,the body weights of rats in the high dose group was significantly decreased (P＜0.05).5. The fecal excreting habit:Addition of RS to the diet significantly increased fecal output and water content. In the three groups RS fed rats, further increase in fecal output and water content were seen with the concentration of RS. The intestinal transit time was shortened by addition of RS and high-RS group had a more markedly effect.6. Intestinal tract pH:Fecal pH in the three RS added groups were significantly decreased than the positive control group (P＜0.05).7. The number of colonic ACF:ACF were discovered in both of the positive control group and in RS groups. But the negative control was no ACF formation. Compared with the positive control group, the number of ACF and AC in the three RS added groups were both significantly reduced (P<0.01).8. Colonic PCNA-LI and the number of AgNORs:The number of AgNORs and the PCNA-LI in the positive control group were more than the negative control group. Compared with the positive control group, the number of AgNORs and the PCNA-LI in the three RS added groups were both reduced and high-RS group had a more markedly effect.9. Expression of colonic Bcl-2 and Bax protein:There was a low expression of Bcl-2 and Bax in the negative control group. In addition, RS inhibited the expression of Bcl-2 and enhanced the expression of Bax.Conclusion:1. This method mimics physiologic conditions preferably adopted by the overseas laboratory widely these days. Not only is it usable for low-content (＜1%) starchy foods but also for high-content starchy foods, with its good recurrence, low price and high analytic efficiency.2. The contents and pattern of resistant starch in different kinds of food were largely varied. Different processing of food material was another important factor influencing the content of RS. The content of RS in cooked foods decreased obviously. Deep-fried and roasted foods have more RS than general braised foods. It could come to the conclusion that rough processing is propitious to retain high amount of RS.Compared with cereal processed food, the content of RS of porridges and cereal drinks. In addition, the content of RS increased generally in the cooked food after preservation.3. Findings from this study suggest that the estimated intake of RS by Chinese population is approximately 9.7g/d on average. Comparing with the values reported abroad, the level was higher than American and European but lower than African. It might be a factor that CRC incidence was lower in China than European. The estimate of RS intake provides a valuable reference for researchers and food and nutrition professionals. 4. Cereals are the Chinese staple food and the most fundamental source of RS. Tubers are the optimal source of RS. It would be prudent to increase consumption of tubers, legumes and cereal grain products.5. Dietary RS retarded the body weight gain to rat. Therefore it may be an effective natural approach to the treatment of obesity.6. Dietary RS shorten the GTT and increase the fecal output and moisture. These effects on excreting habit might be beneficial for eliminating toxic substrates timely. It also decreased intestinal tract pH. All these findings showed that RS favourably influence the colonic luminal environment.7.ACF may be a valuable middle point instead of cancer incidence for colorectal cancer research of chemical prevention.8.The application of some biomarkers (such as PCNA, AgNORs, Bcl-2 and Bax) will provide evidence for early diagnosis and treatment of colorectal cancer, and be of great importance for guiding Chinese diets and early preventing colorectal cancer. Therefore it can be used for the clinical early diagnosis.9.Pre-cancerous model has many advantages such as shorter period, less investment, simple and reliable method and so on. It can be used for reference by other experiments.
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