Association of Antioxidant Vitamins A, C, E and Carotenoids with Cognitive Performance over Time: A Cohort Study of Middle-Aged Adults

Carotenoids may strengthen the association of antioxidant vitamins A, C, and E with favorable cognitive outcomes over time, though a few prospective studies have examined this hypothesis. We evaluated the longitudinal data from 1251 participants in the Healthy Aging in Neighborhoods of Diversity across the Life Span (HANDLS) study (Age at visit 1 in 2004–2009 (v1): 30–65 years). Vitamins A, C, and E dietary intakes and total and individual dietary carotenoids were computed using two 24-h recalls at v1. Cognitive tests, covering global mental status and domains of memory/learning, attention, psychomotor speed, visuo-spatial, language/verbal, and executive function were conducted at v1 and/or v2 (2009–2013); mean ± SD follow-up: 4.66 ± 0.93 years. Mixed-effects linear regression models detected an interaction between vitamin E and total (and individual) carotenoids for three of 11 cognitive tests at v1, with only one meeting the statistical significance upon multiple testing correction whereby vitamin E was linked with greater verbal memory performance in the uppermost total carotenoid tertile (γ0a = +0.26 ± 0.08, p = 0.002), a synergism largely driven by carotenoid lycopene. Vitamins A and C showed no consistent interactions with carotenoids. In conclusion, we provide partial evidence for synergism between vitamin E and carotenoids in relation to better baseline cognitive performance, pending further studies with time-dependent exposures and randomized trials directly examining this synergism.


Digit Span Forward and Backward (DS-F and DS-B)
The Wechsler Adult Intelligence Scale, Revised [4] Digit Span Forward and Backward primarily capture attention and working memory, a component of executive function. The tests were administered according to the manual's instructions. The outcome variable was the total score, which was the total number of correct answers for each test.

Category Fluency
Category fluency [5,6] is a measure of semantic verbal fluency, where participants are asked to generate as many animals as possible within a 60 second duration. Higher scores indicate better category fluency. The outcome variable was the total number of correctly generated words (i.e., words that were not intrusions and perseverations).

Brief Test of Attention (BTA)
For the BTA [7], a test of divided auditory attention, the examiner administered up to 10 trials of letters and numbers (4-18 items) that increased in length with each trial. Only the numbers portion of the test was administered. For each trial, participants were asked to disregard the number of letters read, while tracking how many numbers were recited. They were also told to keep their hands in fists to avoid finger counting. The outcome variable was the total number of correct trials.

Trail Making Tests A and B (TRAILS A and B)
The Trail Making Tests A and B [8] primarily capture attention and executive functioning, respectively. The main executive function subdomain that TRAILS B captures is set-shifting and cognitive control. Both trials also measure visuo-motor scanning and processing speed.
Participants were asked to draw a line between consecutive numbers (TRAILS A) and alternate between numbers and letters (TRAILS B) as quickly as they could. They were informed that they were being timed. The examiner pointed out errors that were then corrected by the participant.
Errors were captured via increased time. Scores for TRAILS A and B reflected seconds to completion, where higher scores indicate poorer performance.

Clock Drawing Test -Clock to Command (CDT)
The Clock Drawing Test [9] is a measure of visuo-spatial abilities, that also captures elements of memory and executive function. Participants are instructed to draw a clock, put in all of the numbers, and set the hands to 10 minutes past 11. Performance is based off correct drawings of the clock face (0-2), numbers (0-4) and hands (0-4). Scores ranged from 0 to 10, with higher scores indicating better performance. Participants who did not score a perfect score on the command portion of the test were also asked to copy a clock with the hands set to 10 minutes after 11.

Wide Range Achievement Test -3 rd Edition: Word and Letter Reading Subtest (WRAT)
The WRAT Word and Letter Reading Subtest [10] is a test of reading ability that is often used as a proxy for literacy and quality of education. Participants were instructed to correctly read a list of 50 words that increased in difficulty. If the first five words were not correctly pronounced, letter reading was also administered. Standard instructions were used with the tan form. The outcome variable used was the total number of correctly pronounced words.

Center for Epidemiological Studies Depression Scale (CES-D)
The CES-D [11] is a 20-item measure of depressive symptomatology. Participants are asked to consider the frequency and severity of their symptoms over the last week. Scores ranged from 0 to 60. Scores of >16 indicated significant depressive symptoms and scores of >20 indicated a clinically significant amount of depressive symptoms.

Method S2: Mixed-effects regression models
The main multiple mixed-effects regression models can be summarized as follows: Multi-level models vs. Composite models Eq.

1.1-1.4
Where Yij is the outcome (Each cognitive test score measured at v1 and/or v2) for each individual "i" and visit "j"; is the level-  (γ0a) and their interaction with TIME (γ1a), as described in a previous methodolgical paper. [12] i 0  b Mixed-effects linear regression models were conducted with each cognitive test score as the outcome and each of dietary individual carotenoids and vitamins A, C and E as exposures entered simultaneously. Random effects were added to the intercept and TIME (See Method S2 for notations). Exposures and covariates were interacted with TIME. Minimally adjusted models (Model 1) are presented and included age at v1 (in years) centered at 48, sex (Male vs. Female), race (African-American vs. White), poverty status (Below vs. Above Poverty), education (<HS, HS, >HS), WRAT-3 total score (centered at 42) and employment status (Unemployed vs. employed, Missing vs. employed), total energy intake (kcal/d, centered at 2006) and the inverse mills ratio. Findings are summarized in terms of significant interactions for a specific cognitive test score, at type I error of 0.05 and the direction of the parameter point estimate: +: positive; -: negative.
All cognitive test scores are in the direction of higher score → better performance with the exception of BVRT (# of errors) and TRAILS A and B (# of sec. to complete). See Table 1 for tertile distribution of total carotenoids.