The Association between dietary total antioxidant capacity with the risk of helicobacter pylori infection among adult: a case–control study

Background Dietary total antioxidant capacity (DTAC) has been assumed as a useful tool to assess the relationship between the cumulative antioxidant food capacity and several chronic disorders. The aim of this study was to evaluate the association between DTAC and risk of Helicobacter pylori in a case-control study. Methods This case-control study was carried out among 150 patients with Helicobacter pylori and 302 healthy subjects aged 18–55 years. Dietary data were collected using a validated 168- items quantitative food frequency questionnaire. DTAC was calculated based on the ferric reducing-antioxidant power (FRAP) values that reported by US Department of Agriculture. Results The mean age and BMI of the study participants were 38.70 years and 25.86, respectively. Compared with control subjects, participants with H pylori signicantly were older and had higher BMI and smoking use. Patients with H pylori compared with control individuals had also lower intake of total ber, vitamin E, vitamin C, vitamin D, total dairy, whole grain and vegetables. Compared with participants in the lowest tertile of DTAC, those in the highest terrtile had a signicantly lower OR for H pylori after further adjustment for potential confounders (Model 2: OR, 0.20; 95% CI, 0.10 – 0.40; P for trend, < 0.001). Conclusions Our ndings showed that a high DTAC was associated with a reduced risk of Helicobacter pylori in adult, which reects the increased intake of natural dietary antioxidants to prevent the progression of Helicobacter pylori. However, further studies are needed to investigate the t relation between DTAC and risk of Helicobacter pylori.


Background
Helicobacter pylori (H pylori), which it has been estimated that its worldwide prevalence is approximately 4.4 billion individuals (1), is a gram-negative micro-aerophilic pathogen replicating on the surface of the gastric epithelium that can cause gastritis, gastric cancer (2). Moreover, the recent document showed that the H pylori infection is associated with an increased risk of pancreatic disease(3), lymphoma (4) and atherosclerosis (5). Based on evidence, H pylori infection is affected by genetic and environmental factors (6) such as host diet and nutrition status (7).
On other hand, Studies suggest that antioxidants (eg, vitamins and N-acetylcysteine) play a role in reducing active oxygen species and gastric in ammation caused by H. pylori, which also exacerbates the disease and prevents the growth of bacteria and eliminates bio lm formation (8). So far, numerous studies have shown that vitamin C and E levels are lower in people with Helicobacter pylori than in those without H. pylori (8). Thus, impaired antioxidant and anti-in ammatory defenses may cause or exacerbate a chronic condition of pyloric H infection. On other hand, various aspects of the effect of natural antioxidants found different in foods on the modulation of oxidative stress and disorders in the antioxidant system have been studied. (9,10) However, the evaluation of an antioxidant compound alone cannot re ect the total antioxidant potency of the diet and re ect the synergistic and potential effects of dietary antioxidant interactions. Thus, the term total antioxidant capacity for diet (DTAC)has been developed and is used as a suitable tool to evaluate the effects of dietary antioxidants that is a strong correlation with serum total antioxidant capacity (11) And it is closely related to the quality of the diet to determine the risk of chronic diseases (12).
Several evidences have suggested the potential link between DTAC and reduce the risk of chronic diseases such as diabetes (13), metabolic and oxidative stress markers (14), ulcerative colitis (15), blood pressure (16), and cardiovascular disease (17), which share common metabolic parameters with H pylori.
However, to our knowledge, the association between DTAC and risk of development H pylori have not yet been investigated. Only, a number of studies in vivo and animal studies have shown the ability of nutrients rich in antioxidant to reduced oxidative stress and subsequently reduced risk of H pylori including garlic, ginger, quercetin, green tea and Cranberry (18)(19)(20)(21). However, the synergistic effects of different antioxidant nutrients, which also found in DTAC and other chemical ingredients utilized in the growing or preparation of food have not been discussed. (22).
Considering the lack of convincing evidence regarding the association of DTAC with liver function, in this study we aimed to investigate the association between DTAC and risk of H pylori in Iranian population in order to improve and reduce the burden of diseases.

Participants
This case-control study was performed on 153 patients with H pylori and 306 healthy subjects who had been referred to the Hazrat Rasoul Hospital, Tehran, Iran during the recent years. In this study, patients with H pylori infection were con rmed by one of the diagnostic methods of blood antibody, fecal antigen, UBT respiratory test, endoscopy and gastric biopsy and performed Rapid Urease Test or pathological and microscopic examination. If they meet the inclusion criteria, are selected as case group after obtaining written informed consent. The control group was twice as likely as the case group and among other patients referred to other parts of the hospital such as ophthalmology, orthopedics, maxillofacial surgery, ear, nose, and throat who were not diagnosed H pylori infection with the diagnostic methods and also not having a history of alcohol or alcohol consumption less than 10 mg / day in women and less than 20 mg / day in men. Case and control were matched regarding age and gender. After entering into the study, information about demographic variables and Hematological parameters were obtained by completing the General Information Questionnaire. Also, the Hematological parameters checked after they came to the hospital, again. Then, for completing data on dietary intake and other information we called the patients to invite them to the research center in a special date. Participants with a history of certain diseases (Gastrointestinal malignant diseases, psychiatric psychosis, diseases that result in lack of reminder like Alzheimer's) were excluded. Also, pregnant and lactating women and subjects with an arbitrary special diet were excluded. In this study, nutritionists were used as interviewers in order to reduce information bias as well as to answer completely to the survey questions. Furthermore, written informed consent was completed for each participant. Also, in order to assess physical activity levels of participants, we used the General Practice Physical Activity Questionnaire (GPPAQ), a simple questionnaire re ecting an individual's current physical activity (23). This study was approved by the research council and ethics committee Iran University of Medical Sciences, Tehran, Iran (ethics code: IR.IUMS.REC.1398.667).
Anthropometric assessment Anthropometric measurements were conducted by a trained dietician. Weight was measured using a standard digital seca scale (made in Germany), while participants wore minimum clothes and without shoes and recorded to the nearest 100 g. Height was measured using a mounted tape in a standing relaxed shoulder position with no shoes to the nearest 0.5 cm. Body mass index (BMI) was calculated as weight (kg) divided by height in square meters (m2).

Measurement of Hematological parameters
All stool samples were collected in plastic containers and sent to the laboratory within 2 h. About 5 mL of venous blood were collected from each subject (cases and controls) and divided equally (2.5 mL) in one tube containing K3-ethylenediaminetetraacetic acid (K3-EDTA) to perform a complete blood count (CBC) using a Cell Dyne 1800 electronic counter (Sequoia-Turner Corporation, California, USA) and in a serum tube to determine serum iron using the DiaSys reagent kit. The serum vitamin B12 concentration was determined quantitatively using a solid phase, competitive chemiluminescent enzyme immunoassay (Immulite/Immulite 1000).

Dietary assessment and DTAC calculation
Nutritional status of individuals was assessed by a 168-items FFQ. The questionnaire has a standard size for each food item designed according to the Willett method (24). Study participants were asked to indicate their frequency of consumption of each food item according to their consumption in the past year. Depending on the type of food consumed, the frequency of consumption per day, week or month was questioned. Previous studies using this questionnaire have provided acceptable results and validity of the questionnaire indicates that the questionnaire has acceptable validity. The values listed for each food item were converted to grams daily using the Home Scale Handbook. Then, the amount of nutrients obtained by each individual was obtained using Nutritionist IV software. The total antioxidant capacity of the diet was obtained from previous articles based on the ferric reducing-antioxidant power (FRAP). The FRAP is de ned as the ability of dietary antioxidants to reduce ferric to ferrous ions. The FRAP was expressed in milligrams per 100 grams of food (25). For similar foods in Iranian culture (for example, different types of bread), the average total values will be calculated. Finally, the frequency of consumption of each food item is multiplied by the amount of iron-reducing antioxidant power associated with it, and then the diet is collected for each participant.to obtain the total antioxidant capacity Statistical analysis: All statistical analyses were using SPSS software (version 19.0; SPSS Inc, Chicago IL). The normality of variables was evaluated by Kolmogorov-Smirnov and histogram tests. For the variables that do not have a normal distribution, logarithmic equivalent (Ln transformation) of that variable was used in the analysis. Also, independent sample T test was used for comparing the mean of DTAC as well as the general characteristics and hematological parameters among case and control group. For assessing the relation between DTAC index and H pylori in an adjusted model, multiple logistic regressions were used. The data were presented as mean ± standard deviation and odds ratio with 95% con dence interval, and in all results, the signi cance level was determined as P < 0.05.

Results
The mean ± SD for the age and BMI of the study population (59.5% women) were 38.70 ± 10.65 years and 25.86 ± 4.84 Kg.m2, respectively. Table 1 shows the distribution of cases and controls according to socio-demographic characteristics, smoking habits, alcohol drinking, body mass composition and hematological parameters. Compared with control subjects, participants with H pylori signi cantly were older and had higher BMI and smoking use. The mean levels of hemoglobin, RBC, WBC, hematocrit, and mean corpuscular hemoglobin concentration (MCHC) were signi cantly lower among cases compared to controls individuals. Furthermore, no signi cant differences were found for mean corpuscular volume (MCV) and mean corpuscular hemoglobin (MCH) cases and controls subjects. On comparing cases with controls, the mean levels of serum vitamin B12 and iron were signi cantly lower in the patients. Table 2 illustrates the dietary intake of H pylori patients compared to controls. Patients with H pylori compared with control individuals had also higher intake of energy, protein, carbohydrate, saturated fatty acids, fructose, iron, whole and re ne grain, and red and processed meat and lower intake of total ber, vitamin E, vitamin C, vitamin D, total dairy, Whole grain and vegetables. There were no significant differences in all other characteristics and dietary intake among case and control subjects.
The ORs and 95% CIs for H pylori in the terrtiles of DTAC are shown in Table 3. Compared with participants in the lowest tertile of DTAC, those in the highest terrtile had a signi cantly lower OR for H pylori ( crude model: OR, 0.29; 95% CI, 0.14 -0.6; P for trend, 0.001), which remained signi cant after further adjustment for BMI, waist circumference, physical activity, smoking, dietary intake of energy and fat. (Model 2: OR, 0.20; 95% CI, 0.10 -0.40; P for trend, < 0.001).

Discussion
The study investigated the association between DTAC and H pylori in a case-control study. We found that DTAC had an inverse association with risk of H pylori infection. Participants with higher DTAC score had a signi cantly lower OR for H pylori infection compared with low DTAC score, after adjustment for potential factors. Our study suggests that DTAC may potentially reduce the risk of NAFLD.
According to our information, this is the rst time that the inverse relationship between DTAC and risk of H pylori infection has been proven H pylori infection has been documented. In the previous evidences, dietary TAC has been inversely associated with several chronic diseases such as cardiovascular diseases (17), cancer (26)diabetes (13), and metabolic disorders (14), which share common metabolic parameters with H pylori infection.
Overall, an inverse associations between intake of antioxidant micronutrients and antioxidant-rich foods with risk of H pylori infection was registered. For example, The DASH diet (Dietary Approaches to Stop Hypertension), dietary patterns with antioxidant-rich component based on fruit and vegetables, low-fat dairy products, and whole grains, has been shown to eradicate Helicobacter pylori and its gastric cancer (27), as well as an inverse association in chronic diseases associated with H pylori infection (28). Furthermore, coffee has been investigated as an antioxidant-rich nutrient as a risk reduction for coffee.
So that, Coffee showed a strong antibacterial action against Helicobacter pylori (H. pylori) (29).This action against H. pylon suggests that coffee may have action as a useful natural inhibitor of gastritis and gastric ulcers. Also, the ability of other antioxidant-rich nutrient to increase plasma TAC and nally reduced risk of H pylori infection through several mechanism, garlic, ginger, quercetin, green tea and Cranberry (19)(20)(21)(22) was well revealed. The ndings were con rmed even in complementary interventions with antioxidants. Besides, the bene cial effects of the combination of antioxidant compounds were also reported, as demonstrated the synergistic effect of Simultaneous administration and combination of one or more antioxidant supplements reducing risk of H pylori infection (19).
The mechanism of action of dietary antioxidant compounds on H. pylori is not clear. However, the intake of antioxidant-rich nutrients in animals with Helicobacter pylori has been shown to reduce both the number of bacteria in the gastric mucosa and the in ammatory process (8,20). On the other hand, it is possible that these cultivations, by destroying the Iklova factor, have direct antiseptic properties and prevent NF-κβ nuclear transfer, thereby reducing genetic expression and producing mediators associated with relevant in ammation such as TNF-α, IL-1β, IL-6 and IL-8.It is possible that antioxidant-rich nutrient, by controlling some of the toxins that cause enough bacteria to grow by creating special anionic channels in the plasma membrane associated with the release of bicarbonate and organic anions in gastric cells, they have a bene cial effect (30). On the other hand, it is possible that these compounds reduce the action of urease on H. pylori, because it has been shown that several polyphenolic compounds can alter the activity of this enzyme, which is responsible for the survival of bacteria in acid. (31) In addition, ber, as part of a healthy diet in many high-antioxidant plant foods, may have health bene ts through protection against oxidative damage, as well as improving glucose metabolism and serum lipids (32). Also, some prior studies have declared that dietary bers can alter abundances of speci c gut microbes and change the pro le of human gut microbiota (33)(34)(35)(36). Fiber fermentation is dependent on microbiota (37) and it has been seen that ber may increase gram-positive bacteria (38), by decreasing the gut transit time and pH (39,40). Moreover, we know that H pylori is the gram-negative bacteria so it may be negatively affected by ber. Also, a high ber diet in rich of vegetables, fruits, whole grains and beans (41) can exert a protective effect against H pylori infection (42,43). This may be due to their prebiotic properties which is con rmed in a meta-analysis (44). Besides, fresh vegetables are the source of vitamin C which has been mentioned as a chemopreventive factor against digestive disorders caused by H pylori infection (42). As far as we know, vitamin C can lessen stomach cancer risk and exert an effect on the cycle of H pylori infection (45). Vitamin C is highly concentrated in stomach mucosa and gastric juice. it has a positive effect on the production and function of immune cells and immunoglobulin (46). In our study, dietary vitamin c consumption was signi cantly lesser in case group. Also, Dietary antioxidants may reduce the risk of developing this disease by reducing the body's obesity and visceral fat, which are known to be one of the risk factors for Helicobacter pylori infection (47,48). This is the rst study to assess the relationships between DTAC with risk of H pylori Our sample size was su cient and we tried to eliminate the effect of confounders as far as possible, by adjusting wide range of variables and a validated questionnaire has been used. As for most case-control studies, the causal relationship can be determined but it cannot be quite de nite. Furthermore, some conditions of foods such as the growing condition, cultivation method, storage, processing and cooking condition, and assay method might affect the antioxidants content of the foods (25) Also, information collecting in these kinds of studies are based on individual's reports and memory which is not totally reliable. We have tried to minimize information bias by administrating a questionnaire by trained interviewers.

Conclusion
Our ndings showed that a high DTAC was associated with a reduced risk of H pylori infection in adult, which re ects the increased intake of natural dietary antioxidants to prevent the progression of. However, further observational studies and clinical trials are needed to investigate the exact relation between DTAC and risk of H pylori infection.

Declarations
Abbreviations Not applicable.
Ethics approval and consent to participate Not applicable.

Consent for publication
All authors of this manuscript declared their consent for publication.