Impacts of Antioxidant Vitamins, Curry Consumption and Heavy Metal Levels on The Metabolic Syndrome With Comorbidities: A National Cross-Sectional Study

The burden of metabolic syndrome (MetS) is increasing worldwide especially in the coronavirus disease 2019 (COVID-19). This phenomenon can be related to environmental, dietary, and lifestyle risk factors. We aimed to determine the association between the levels of serum heavy metals, vitamins, and curry intake, subsequently predict the risks for MetS by margin effects. Daily intake of vitamins was measured by 24-h recall was calculated using a food frequency questionnaire. Heavy metals were quantied by graphite furnace atomic absorption spectrometry, and mercury analyzer. The risk of MetS was signicantly lower in the high curry consumption than in the low curry consumption, risks of Mets were reduced by 7%, 13%, 1%, and 1%, when the levels of vitamin B1, B2, B3, and C intake increased by one mg, respectively. However, risks of Mets were increased by 9%, 3%, 5%, when the levels of serum Pb, Hg, and CRP increased by one unit. The potential health benets resulting from vitamin and curry supplementation could guard the public against the dual burden of communicable and non-communicable diseases. Further works are required to thwart risk factors related to heavy metals and determine the mechanistic dual effects of vitamins and curry in MetS. (HDL-C),


Introduction
Food is emerging as a signi cant modi able contributor to chronic disease with empirical evidence that dietary modi cations have clear positive as well as negative impacts on the lifelong health 1 . In fact, consumption of fatty, high-saturated, and low-energy diets as well as inactive physical activity, overuse of tobacco and alcohol has been increasing, which was contributing to the growth of non-communicable diseases (NCDs), especially Metabolic syndrome (MetS) 1,2 . MetS is commonly known as a collection of metabolic disorders, including resistance to insulin, dyslipidemia, central obesity, and hypertension.
Besides, MetS was the risk factor for the development of type 2 diabetes and cardiovascular diseases 3 .
Of note, recent evidence suggested that MetS affects the progression and prognosis of coronavirus disease 2019 (COVID- 19), and increased metabolic severity has been correlated with worse COVID-19 consequences 4,5 . It makes the situation get worse once COVID-19 has spread around the world.
In addition to lifestyle and genetic factors, the effects of heavy metal on the risk factors of MetS 6-9 . Remarkedly, the risk of exposure to heavy metals released by vehicles, factories, or contaminated seafood is increasing, especially lead (Pb), mercury (Hg), and cadmium (Cd) without biological roles in human body systems, but accumulate in bones, kidney cortex, and lungs 10 . Heavy metals catalyze the release of reactive oxygen species as well as in ammatory mediators and antithrombotic substances that cause damage to vascular endothelial cells and exacerbate hypertension 7,11 . Therefore, Pb and Cd disrupt blood clotting and increase the risk of CVD, while Hg accelerated the process of carotid atherosclerosis 6,12 .
Increasing evidence shows that vitamin supplementation could reverse CVDs, and diabetes, and mental illness 13 . Additionally, curcumin, curcuma longa dried rhizome, is also supportive for the prevention and/or treatment of CVDs due to its anti-oxidant and anti-in ammatory properties 14 . In this study, we present tentative evidence that increased daily intake of vitamins and curry is related to better performance of MetS among the Korean population with various non-communicable diseases. On the other hand, the association of heavy metal levels with MetS is also provided.

Methods
Study population. We used a strati ed, multi-stage, cluster-sampling method that regarded the geographic zone, level of urbanization, economic development condition, gender, and age distribution, conducted by  Parameters. Information on sociodemographic characteristics, lifestyle, current medications, medical, and family history was collected during the health interview. Alcohol intakes were classi ed as low and high (high-risk drinking was de ned as > 5 drinks per day and ≥ 1 month). Subjects with a lifetime history of smoking of >100 cigarettes in their lifetime and still smoked daily or occasionally were classi ed as current smokers; others were classi ed as ex/non-smokers. Physical activity was dichotomized as regular or irregular. Regular physical activity was de ned as: (1) vigorous physical activity, ≥20 minutes per session ≥3 days a week (2) moderate physical activity; ≥30 minutes per session ≥5 days per week, and (3) walking; ≥30 minutes per session ≥5 days a week.
Dyslipidemia was de ned as one or more of the following: LDL-C ≥160 mg/dL, triglyceride ≥200 mg/dL, HDL-C <40 mg/dL. Hypertension was de ned as having either systolic blood pressure (SBP) ≥ 140 mmHg or diastolic blood pressure ≥ 90 mmHg or on anti-hypertensive medication. Type 2 diabetes mellitus was de ned as having a fasting plasma glucose of ≥126 mg/dl or on anti-diabetic medication, or HbA1c≥6·5%. Stroke, angina, myocardial infarction (MI), MI or angina, and other diseases were de ned as physician diagnosis, the current presence or treatment for stroke, angina, MI, MI or angina, and other diseases. Depression was de ned as physician diagnosis, the current presence or treatment for depression, or if participants have experienced depression in the past year or despair to the point where it disturbs their daily routine for 2 weeks in a row or longer 59 . Comorbidities have de ned any diseases such as CVDs, hypertension, hyperlipidemia, type 2 diabetes, cancers, thyroid, kidney, arthritis, osteoarthritis, rheumatoid arthritis, depression are present in the participants with MetS. A family history of cardiovascular disease was de ned as having at least one parent or sibling with a diagnosis of hypertension, ischemic heart disease, or stroke. A family history of type 2 diabetes or hyperlipidemia was de ned as having at least one parent or sibling with a diagnosis of type 2 diabetes or hyperlipidemia.
Laboratory measurements. The height, weight, waist circumference, and blood pressure were measured during medical checkups using the standard procedure. BMI (kg/m 2 ) was estimated using the formula: BM= weight (kg)/ height 2 (m 2 ). Waist circumstance (cm) was measured at the midpoint between the bottom of the rib cage and the iliac crest of the mid-axillary line when exhaling. Blood pressure was calculated three times with intervals of 5 minutes using a mercury sphygmomanometer with a subject seated after a 5-minute stabilization period. Final blood pressure was the average of the second and third measurements. Blood samples after ≥ 8 hours of fasting were collected and analyzed at the Neodin Medical Institute in Korea. An enzymatic assay was then used to determine levels of total cholesterol, high-density lipoprotein cholesterol (HDL-C), triglycerides, low-density lipoprotein cholesterol (LDL-C), and fasting glucose using the Hitachi automated analyzer 7600 (Hitachi, Japan).
Determination of Pb, Hg, and Cd in blood. Pb, Hg, and Cd analyzes were decribed in the previous study 9 . In brief, these tests were performed by the Neodin Medical Institute, which was approved by the Korean Ministry of Labor for Heavy Metal Analysis. Furthermore, these tests also were met the criteria of the Korea Occupational Safety and Health Administration, the German External Quality Assessment Scheme, and the U.S. CDC. Pb and Cd were measured by graphite furnace atomic absorption spectrometry (model AAnalyst 600; Perkin Elmer, Turku, Finland) using Zeeman background correction, and total Hg was calculated by a direct mercury analyzer (model DMA-80 Analyzer; Bergamo, Italy) and gold amalgam (KCDC 2013). Limits of detection (LODs) were 0.223 µg/dL, 0.05 µg/L, 0.087 µg/L for Pb, Hg, and Cd, respectively. Commercial standard reference materials purchased from Bio-Rad for internal quality assurance and control (Lyphocheck Whole Blood Metals Control; Bio-Rad, Hercules, CA, USA).
Metabolic syndrome. MetS were de ned using American Heart Association/National Heart, Lung, and Blood Institute criteria for clinical diagnosis that included abdominal obesity, elevated triglycerides, increased waist circumference, decreased HDL, elevated blood pressure, and elevated plasma glucose 60 .
Participants with three or more of the following ve risk factors were de ned with metabolic syndrome.
Vitamin intake. Daily food intake was calculated using the 24-h recall method. Before evaluating the food intake, all participants were instructed to uphold their normal dietary habits. A semi-quantitative questionnaire on food frequency, which addressed the intakes of 63 food products, was completed by each participant. The levels of participants of food consumption were calculated using nine categories: never or rarely," "once a month," "two to three times a month". Often a week," "three or four times a week," " ve to six times a week," "once a week," " ve to six times a week. Day," "twice a day," and "every day, three or more times. The daily intake of thiamine was determined by summing the mean of the 24-hour dietary intakes using the Can-Pro 3.0 nutrient intake assessment software developed by the Korean Nutrition Society. The daily total energy intakes were measured using the Estimated Energy Requirement (EER) in Korea 62 .
The curry consumption was estimated using the KNHANES food frequency questionnaire. Curry rice was the only food in the surveyed foods related to curry consumption. According to the frequency of their curry consumption, subjects were divided into two groups: the low curry consumption group ("almost never", or "once a month"), and the high curry consumption group ("2-3 times a month" or "once a week" or "2-6 times per week").
Statistical analysis. All statistical analyses were undertaken using STATA software (version 16.0; StataCorp, Texas, USA). The baseline characteristics of participants were summarized using frequency and proportion for categorical variables; mean and standard deviation for continuous variables. Student's t-test for continuous variables and χ² test for categorical variables.
After adjustment for comorbidities, the risk of MetS was signi cantly lower in subjects with high curry consumption than subjects with low curry consumption, risks of Mets were reduced by 44% ( 1.09, p<0.001), when the levels of serum Pb, Hg, Cd, and CRP increased by one unit. The marginal effect of the levels of vitamin intake, curry consumption, heavy metals and serum CRP on MetS by comorbidities was shown in Figure 2.

Discussion
Our ndings include epidemiological evidence that draws upon an important volume of earlier experimental knowledge to support the association between vitamin, curry consumption, heavy metals, and effectiveness for MetS among subjects with comorbidities. We also identi ed the association between MetS, vitamin heavy metals, and comorbidities on a national level in Korea. We found that levels of vitamin intake and high curry consumption showed an inverse correlation with MetS, while levels of serum heavy metals and CRP showed a positive correlation with MetS. These ndings partly contribute to the dual effects of vitamin, curry, and heavy metal consumption in Korea.
The dramatic global rise of urbanization and industrialization has increased the risk of exposure to pollutants, particularly heavy metals 15 . Besides, the growing global burden of NCDs has made the prevention and management of NCDs a priority, especially in the context of COVID-19 pandemic. Because the COVID-19 pandemic has also been connected with NCDs related to disease-induced morbidity and mortality, especially preventive measures such as the impact of social distancing and stay-at-home orders 16 .
In this study, we found that the levels of heavy metals and cardiometabolic risk factors have positively correlated with MetS. These ndings are in agreement with the previous study 9,17 . Heavy metals such as Pb, Hg, and Cd are toxic to the human body and can trigger different diseases, especially CVDs 18 . Heavy metals can increase the levels of reactive oxygen species and reactive nitrogen species leading to increased oxidative stress, consequently, cause DNA damage and oxidize protein thiol groups 19 .
Furthermore, heavy metals destroy blood clotting and provoke the production of in ammatory cytokines and anti-thrombotic agents 6,7 . Recent data also indicate that, increased risk of the development of MetS with higher quartiles of CRP level in obese and non-obese women 20 . Therefore, the special concern should be given to the harmful impacts of multiple environmental pollutants, along with heavy metals on MetS. It is crucial to developing a prevention strategy targeting the high-risk population to slow down this progression to postpone risk factors related to heavy metals and reduce prevalence.
MetS is a risk factor for the formation of type 2 diabetes and CVDs 21 . While vitamin B1 plays an important role in intracellular glucose metabolism since it acts as a coenzyme for the α-ketoglutarate dehydrogenase complexes, transketolase, and pyruvate dehydrogenase 22 . Our study showed the levels of vitamin intake were signi cantly lower in subjects with MetS compared with subjects without MetS. Additionally, levels of HbA1c, fasting glucose were signi cantly higher in participants with MetS compared with those without MetS. It has been shown that reduced vitamin B1 in diabetic vascular cells exacerbates metabolic dysfunction in hyperglycemia 23 . The relationship between diabetes and vitamin B1 has been reported by genetic studies such as Tk, α-1-antitrypsin, SLC19A2 gene, and p53 [24][25][26][27] . Vitamin B1 and its derivatives could thwart the biochemical pathways of caspase activation like improved ux via the polyol or hexosamine biosynthesis pathway, the making of advanced glycation end-products, induction of protein kinase C activity produced by hyperglycemia [28][29][30][31] . These data support our nding for the consumption of su cient vitamin B1 presented an inverse correlation with MetS. Our ndings were in line with the previous results of a signi cant reduction in plasma fasting glucose concentrations at a daily dose of 150 mg thiamine per 1 month in patients with drug-naïve type 2 diabetes 32 . Besides, the consumption of su cient vitamin B1 also reported an inverse related to dyslipidemia 33 . Vitamin B1 thwarts the adverse consequence of high endothelial glucose levels by dropping the glycation of intracellular proteins 34 ; vitamin B1 also plays a vital role in averting the development of atherosclerotic plaque as it had a positive effect against glucose-and insulin-mediated development of human infragenicular arterial smooth muscle cells 35 . Several studies also showed the regular vitamin B1 administration increases the functions of endothelial cells and slows the development of atherosclerosis 36 , and short-term vitamin B1 treatment has regenerated the function of endothelial cells in healthy smokers with endothelial dysfunction caused by smoking 37 . Our ndings long-established the role of vitamin B1 in the reversal of MetS, which is a risk factor that impacts the progression and prognosis of COVID-19.
Oxidative stress could play a signi cant part in MetS etiology 38 . Subjects with MetS have increased systemic oxidative damage due to upregulation of reactive oxygen species (ROS) and/or reduced antioxidant protection due to antioxidant enzymes 39 . Our ndings show that intake of vitamin B2, B3, and C can reduce MetS, which was in line with the previous studies 40-42 . It could be explained that vitamin B2 could prevent the pro-in ammatory activity of adipocyte and macrophage co-cultures, thus decreasing the possibility of mild in ammation associated with obesity 40 . While vitamin B2 de ciency could increase the proin ammatory activity of adipocyte cells, resulting in chronic in ammation in obesity 43 . Vitamin B3 has also been found to be an e cient antioxidant in ROS production and to prevent DNA damage in lymphocytes 44 . It supports the previous study about vitamin B3 treatment has favored the normalization of low HDL-C atherogenic dyslipidemia 45 . Furthermore, a positive correlation has been reported between diets enriched with speci c antioxidants such as vitamin C for oxidative stress 46 . These ndings highlight the value of urgent efforts to establish targeted vitamin supplementation in Korea. We believe these strategies would effectively diminish the prevalence of MetS; therefore, it could support patients through the COVID-19 pandemic and reduce the morbidity and mortality of this syndrome.
Our analyses revealed that the risk of MetS was signi cantly lower among the high curry consumption group. These ndings support our hypothesis that high curry consumption is related to reducing the risk of type 2 diabetes, which was in agreement with previous studies 47 . Curcumin could improve endothelial function and reducing oxidative stress and in ammatory markers (IL-6, TNF alpha, endothelin-1) in type 2 diabetes patients, and it also helps to enhance the function of β cells. 48,49 . Of note, curcumin has also an impact on insulin secretion in healthy participants 50 . On the other hand, Curcumin has the potential bene t of reducing triglyceride, total cholesterol, LDL-C, and increasing HDL-C in patients with acute coronary syndrome 51 . Recent data indicated that curcumin has a preventive stroke effect by reducing the oxidative stress levels related to signaling the uncoupling of protein 2 to strengthen endothelial vascular function 52 . Curcumin also has a profound effect on the microglial response, facilitate microglial M2 polarization, and prevents the pro-in ammatory response of microglia. Additionally, curcumin post-treatment among ischemic stroke patients diminishes brain damage and strengthens vascular endothelial function 53 . These data showed the potential bene ts of curcumin in diabetes and CVDs. Consequently, curcumin is not only a promising therapeutic alternative for type 2 diabetes because of its anti-in ammatory property but also a preventive strategy for CVDs.
To our knowledge, this large-scale study is the rst to report the dual effect of vitamin, curry, heavy metal consumption, and MetS on a national level in Korea. However, this study has several limitations. First, the cross-sectional method could not evaluate causality between MetS, and levels of thiamine and curry, heavy metal consumption. Second, as no physiological markers of antioxidant status in KNHANES were measured, oxidation status and vitamin levels in plasma and tissues were not evaluated. Third, vitamin consumption is measured from 24-h recall data which is based on recall; therefore, participants could be under-or overestimated their food consumption. However, the 24-h recall may be an effective technique to assess food intake, especially, before evaluating the food consumption, all participants were instructed to uphold their usual dietary habits. Finally, there were few individuals who consumed curry with high frequency, which could be underestimated the association between curry consumption and MetS.
The prevalence of MetS and exposure to heavy metal in Korea tends to be increasing [54][55][56] , it gets worse during the COVID-19 pandemic 57 . MetS are signi cant factors of risk for severe COVID-19, but the mechanisms have been somewhat uncertain. Furthermore, the dramatic global rise of urbanization and industrialization has increased the risk of exposure to heavy metals. It is critical to developing a prevention strategy targeting the high-risk population to slow down this progression to postpone risk factors related to heavy metals and reduce prevalence. Vitamins and curry supplements could reverse MetS, these results indicate that the potential health bene ts resulting from thiamine and curry supplementation could guard the public against the dual burden of communicable and noncommunicable diseases in Korea. However, further works are required to determine the mechanistic dual effects of vitamins and curry in MetS.

Declarations
The author(s) declare no competing interests.   The levels of vitamin intake (A), serum heavy metals and serum CRP (B) in subjects with or without metabolic syndrome (MetS). Two-sample t test with unequal variances. ***, P<0.001.

Figure 3
Crude odds ratio and adjusted odds ratio (95% con dence interval) for the risks of Metabolic syndrome.
Adjusted for monthly household income, residential areas, energy intake, age group, occupation, sex, family history of CVDs, family history of diabetes mellites, family history of hyperlipidemia, BMI group, smoking status, high risk drinking, physical activity, education level, hypertension, dyslipidemia, type 2 diabetes, stroke, myocardial infarction or angina, myocardial infarction, angina, arthritis, osteoarthritis, rheumatoid arthritis, kidney failure, depression, thyroid disease, and asthma. (shown in the supplementary material).