The recruited participants completed a cognitive questionnaire together with the first questionnaire from the 2011–2012 and 2013–2014 NHANES surveys. As shown in Fig. 1, this study included 2,854 elderly American participants.
The mean age of study participants was 69.4 years. Study participants had a mean score of 19.0 for the CERAD word list learning test, 5.9 for the CERAD recall test, 16.7 for the animal fluency test, and 46.2 for the digital symbol test. The mean CERAD word list learning test score was 1.7 points higher in the 2013–2014 cycle than in the 2011–2012 cycle, while the mean CERAD recall test was 0.7 points higher. The animal fluency test and digital symbol test were not significantly different in the 2011–2014 two cycles. Mean theobromine intake was 81.8 mg/d. Theobromine intake did not significantly vary between the 2011–2012 and 2013–2014 cycles. The characteristics of other covariates are presented in Table 1.
Table 1
Study population characteristics by survey cycle
|
Total1
|
2011–2012 Cycle1
|
2013–2014 Cycle1
|
P value
|
Age, yr, mean ± SE
|
69.4 ± 0.2
|
69.3 ± 0.3
|
69.5 ± 0.2
|
0.90
|
Male,
|
9859 (46.0)
|
4856 (45.5)
|
5003 (46.4)
|
0.62
|
Race
|
|
|
|
0.09
|
Mexican American
|
3085 (3.7)
|
1355 (2.9)
|
1730 (4.4)
|
|
Other Hispanic
|
2036 (3.9)
|
1076 (4.5)
|
960 (3.4)
|
|
Non-Hispanic White
|
6647 (78.0)
|
2973 (78.2)
|
3674 (77.9)
|
Non-Hispanic Black
|
4950 (8.9)
|
2683 (8.5)
|
2267 (9.2)
|
|
Other races including multi-racial
|
3213 (5.5)
|
1669 (5.9)
|
1544 (5.2)
|
|
Education
|
|
|
|
0.07
|
Less than 9th grade
|
1005 (6.2)
|
550 (7.5)
|
455 (5.0)
|
|
9-11th grade (includes 12th grade with no diploma)
|
1573 (10.4)
|
782 (10.8)
|
791 (10.0)
|
|
High school graduate
|
2472 (22.4)
|
1169 (22.4)
|
1303 (22.5)
|
Some college
|
3427 (31.1)
|
1657 (29.9)
|
1770 (32.2)
|
College graduate or above
|
2840 (29.8)
|
1397 (29.4)
|
1443 (30.2)
|
Marriage status
|
|
|
|
0.73
|
Married/living with partner
|
2300 (4.4)
|
1188 (4.6)
|
1112 (4.3)
|
|
Widowed/divorced/separated
|
6505 (64.9)
|
3123 (65.4)
|
3382 (64.4)
|
Never married
|
2514 (30.7)
|
1242 (30.0)
|
1272 (31.3)
|
Home status
|
|
|
|
0.46
|
Owned or being bought
|
10553 (83.5)
|
5003 (84.3)
|
5550 (82.9)
|
Rented
|
8703 (14.5)
|
4434 (14.1)
|
4269 (14.9)
|
Other arrangement
|
482 (2)
|
259 (1.7)
|
223 (2.2)
|
|
Employment
|
|
|
|
0.39
|
Employed
|
6101 (69.6)
|
3085 (68.8)
|
3016 (70.3)
|
Unemployed
|
6530 (30.4)
|
3087 (31.2)
|
3443 (29.7)
|
Smoking Status
|
|
|
|
0.11
|
Never
|
6716 (49.7)
|
3184 (49.7)
|
3532 49.7)
|
Previous
|
2606 (39.6)
|
1259 (38.4)
|
1347 (40.7)
|
Current
|
2340 (10.7)
|
1108 (11.9)
|
1232 (9.7)
|
|
BMI
|
|
|
|
0.24
|
Underweight
|
3640 (1.4)
|
1833 (1.1)
|
1807 (1.6)
|
|
Normal or healthy weight
|
5471 (25.2)
|
2669 (26.6)
|
2802 (34.0)
|
Overweight
|
6603 (58.7)
|
3214 (58.5)
|
3389 (58.9)
|
Obese
|
1943 (14.8)
|
886 (13.8)
|
1057 (15.6)
|
History of disease
|
6438 (68.5)
|
3114 (64.0)
|
3324 (72.5)
|
< 0.01
|
Heart disease
|
900 (17.9)
|
428 (17.6)
|
472 (18.3)
|
0.63
|
History of stroke
|
431 (6.6)
|
229 (6.0)
|
202 (7.2)
|
0.21
|
Hypertension
|
4205 (58.8)
|
2031 (56.2)
|
2174 (61.1)
|
0.01
|
Diabetes
|
|
|
|
0.00
|
No
|
17365 (76.1)
|
8524 (78.0)
|
8841 (74.4)
|
Borderline
|
310 (4.3)
|
125 (3.0)
|
185 (5.5)
|
|
Yes
|
1445 (19.6)
|
708 (19.0)
|
737 (20.1)
|
|
Sleep disorder
|
1064 (12.5)
|
485 (11.8)
|
579 (13.1)
|
0.30
|
Depression
|
2534 (21.1)
|
1178 (16.6)
|
1356 (25.0)
|
< 0.01
|
Dietary intake
|
|
|
|
Energy, kcal
|
1839.5 ± 16.9
|
1833.7 ± 21.3
|
1844.6 ± 25.7
|
0.46
|
Protein, g
|
72.1 ± 0.7
|
71.2 ± 1.1
|
72.9 ± 0.8
|
0.39
|
Lutein + Zeaxanthin, mg
|
1750.1 ± 96.8
|
1957.0 ± 186.7
|
1567.3 ± 78.5
|
0.07
|
Folic acid, mcg
|
373.3 ± 5.7
|
383.9 ± 9.4
|
364.0 ± 6.8
|
< 0.01
|
Vitamin B12, mcg
|
4.7 ± 0.1
|
5.0 ± 0.3
|
4.3 ± 0.1
|
< 0.01
|
Added vitamin B12, mcg
|
0.9 ± 0.0
|
1.0 ± 0.1
|
0.8 ± 0.1
|
0.16
|
Vitamin D (D2 + D3), mcg
|
4.7 ± 0.1
|
4.7 ± 0.2
|
4.7 ± 0.2
|
0.80
|
Magnesium, mg
|
281.6 ± 3.1
|
282.5 ± 5.1
|
280.8 ± 3.9
|
0.27
|
Iron, mg
|
13.8 ± 0.2
|
14.1 ± 0.3
|
13.5 ± 0.3
|
< 0.01
|
Zinc, mg
|
9.9 ± 0.1
|
9.9 ± 0.2
|
9.9 ± 0.1
|
0.55
|
Copper, mg
|
1.2 ± 0.0
|
1.3 ± 0.0
|
1.2 ± 0.0
|
< 0.01
|
Selenium, mg
|
102.0 ± 1.1
|
99.3 ± 1.4
|
104.5 ± 1.6
|
0.05
|
Caffeine, mg
|
146.7 ± 4.8
|
150.9 ± 8.5
|
142.9 ± 5.1
|
< 0.01
|
Theobromine, mg
|
31.8 ± 1.3
|
31.0 ± 2.1
|
33.1 ± 1.4
|
0.29
|
CERAD: immediate recall score
|
19.0 ± 0.2
|
18.1 ± 0.3
|
19.7 ± 0.2
|
< 0.01
|
CERAD: delayed recall score
|
5.9 ± 0.1
|
5.6 ± 0.1
|
6.3 ± 0.1
|
< 0.01
|
Animal fluency score
|
16.7 ± 0.1
|
16.5 ± 0.2
|
16.8 ± 0.2
|
0.59
|
Digital symbol score
|
46.2 ± 0.6
|
45.4 ± 1.0
|
46.9 ± 0.8
|
0.43
|
1: Categorial variables are presented as unweighted sample size (weighted percentage); Continuous variables as mean ± SE. |
Abbreviations: Body Mass Index, BMI; GED; AA; Consortium to Establish a Registry for Alzheimer's disease, CERAD |
Table 2 Weighted regression models evaluating association between dietary theobromine intake and cognitive performance.
CERAD: Immediate Recall Score
|
Theobromine intake, mg/d
|
Non-adjusted1
|
Adjust I2
|
Adjust II3
|
Adjust III4
|
β
|
95% CI
|
P value
|
β
|
95% CI
|
P value
|
β
|
95% CI
|
P value
|
β
|
95% CI
|
P value
|
Continuous Form
|
0.002
|
(0.000, 0.004)
|
0.01
|
0.002
|
(-0.000, 0.003)
|
0.07
|
0.002
|
(-0.000, 0.003)
|
0.07
|
0.001
|
(-0.001, 0.003)
|
0.16
|
Log form
|
0.588
|
(0.151, 1.025)
|
0.01
|
0.627
|
(0.228, 1.027)
|
<0.01
|
0.672
|
(0.262, 1.082)
|
<0.01
|
0.661
|
(0.222, 1.101)
|
<0.01
|
CERAD: Delayed Recall Score
|
Theobromine intake, mg/d
|
Non-adjusted1
|
Adjust I2
|
Adjust II3
|
Adjust III4
|
β
|
95% CI
|
P value
|
β
|
95% CI
|
P value
|
β
|
95% CI
|
P value
|
β
|
95% CI
|
P value
|
Continuous Form
|
0.001
|
(-0.000, 0.002)
|
0.24
|
0.000
|
(-0.001, 0.001)
|
0.48
|
0.000
|
(-0.001, 0.001)
|
0.45
|
0.000
|
(-0.001, 0.001)
|
0.56
|
Log form
|
0.172
|
(-0.041, 0.386)
|
0.11
|
0.214
|
(0.014, 0.413)
|
0.04
|
0.207
|
(0.005, 0.409)
|
0.05
|
0.232
|
(0.016, 0.449)
|
0.04
|
Animal Fluency Score
|
Theobromine intake, mg/d
|
Non-adjusted1
|
Adjust I2
|
Adjust II3
|
Adjust III4
|
β
|
95% CI
|
P value
|
β
|
95% CI
|
P value
|
β
|
95% CI
|
P value
|
β
|
95% CI
|
P value
|
Continuous Form
|
0.004
|
(0.002, 0.006)
|
<0.01
|
0.001
|
(-0.001, 0.004)
|
0.15
|
0.002
|
(-0.001, 0.004)
|
0.14
|
0.001
|
(-0.001, 0.003)
|
0.3
|
Log form
|
0.729
|
(0.194, 1.264)
|
<0.01
|
0.528
|
(0.063, 0.994)
|
0.03
|
0.492
|
(0.009, 0.976)
|
0.05
|
0.393
|
(-0.122, 0.907)
|
0.13
|
Digital Symbol Score
|
Theobromine intake, mg/d
|
Non-adjusted1
|
Adjust I2
|
Adjust II3
|
Adjust III4
|
β
|
95% CI
|
P value
|
β
|
95% CI
|
P value
|
β
|
95% CI
|
P value
|
β
|
95% CI
|
P value
|
Continuous Form
|
0.015
|
(0.008, 0.022)
|
<0.01
|
0.009
|
(0.004, 0.014)
|
<0.01
|
0.009
|
(0.004, 0.014)
|
<0.01
|
0.009
|
(0.004, 0.015)
|
<0.01
|
Log form
|
1.525
|
(-0.002, 3.053)
|
0.05
|
1.5
|
(0.320, 2.680)
|
0.01
|
1.631
|
(0.456, 2.806)
|
0.01
|
1.395
|
(0.140, 2.649)
|
0.03
|
1Adjusted non-variables: Age, sex, race, education, marital status, home status, employment, smoking status, BMI, history of disease, heart disease, history of stroke, hypertension, diabetes, depression, sleep disorder, and sex were adjusted.
2Adjusted model I for sex, age, race, education, marriage, home status, and employment.
3Adjusted for sex, age, race, education, marriage, home status, employment, smoking status, BMI, history of disease, heart disease, history of stroke, hypertension, diabetes, depression, and sleep disorder.
4Adjusted model III for sex, age, race, education, marriage, home status, employment, smoking status, BMI, history of disease, heart disease, history of stroke, hypertension, diabetes, depression, sleep disorder, and dietary nutrients (including alcohol, energy, protein, lutein, zeaxanthin, folic acid, vitamin B12, vitamin B12, vitamin D, magnesium, iron, zinc, copper, selenium, and caffeine).
Abbreviations: Consortium to Establish a Registry for Alzheimer's disease, CERAD.
Figure 2 presents a weighted distribution of daily theobromine intake in log form with respect to quartiles of immediate recall score, delayed recall score, animal fluency score, and DSST score. Daily theobromine intake was not significantly different among these quartile subgroups. As shown in Supplementary Fig. 1, daily theobromine intake did not follow a normal distribution. However, the log transformation distribution did. Thus, we used a continuous form and a log transform of daily theobromine intake to analyze the association between dietary theobromine and cognitive performance, as shown in Table 3 In the initial model, log transformation of daily theobromine intake was significantly associated with immediate recall test score (β, 95%CI, P value: 0.588, 0.151–1.025, 0.01), animal fluency test score (β, 95%CI, P value: 0.729, 0.194–1.264, < 0.01), and DSST score (β, 95%CI, P value: 1.525, -0.002-3.053, 0.05), but not with the delayed recall test (β, 95%CI, P value: 0.172, -0.041-0.386, 0.11). In the fully adjusted model, dietary theobromine intake was positively associated with immediate recall score (β, 95% CI, P: 0.661, 0.222–1.101, < 0.01), delayed recall score (β, 95% CI, P = 0.232, 0.016–0.449, 0.04), and DSST score (β, 95%CI, P value: 0.009, 0.004–0.015, < 0.01). Daily theobromine intake was not associated with the poor cognitive function measured by the four indexes, independent of whether confounding variables were adjusted or not.
To study the association between theobromine intake and cognitive performance we performed subgroup analyses. Theobromine was significantly positively associated with immediate recall score in those who never-smoked (β, 95%CI, P value: 0.006, 0.003–0.010, < 0.01), had hypertension (β, 95%CI, P value: 0.004, 0.001–0.008, 0.02), but not depression (β, 95%CI, P value: 0.005, 0.002–0.008, < 0.01), the 3rd quartile of daily energy (β, 95%CI, P value: 0.002, 0.002–0.011, < 0.01), and protein (β, 95%CI, P value: 0.006, 0.001–0.010, 0.01) intake.
Daily theobromine intake was negatively associated with delayed recall test scores in males (β, 95%CI, P value: -0.002, -0.004-0.000, 0.02), age > 80 (β, 95%CI, P value: -0.004, -0.008-0.000, 0.03), and the 3rd quartile of Lutein + Zeaxanthin (β, 95%CI, P value: -0.004, -0.006- -0.001, < 0.01). However, it was positively associated with delayed recall test scores in the 2nd quartile of Lutein + Zeaxanthin (β, 95%CI, P value: 0.004, 0.001–0.007, 0.01).
Theobromine intake was positively associated with animal fluency score in never-smokers (β, 95%CI, P value: 0.006, 0.002–0.011, < 0.01), normal BMI (β, 95%CI, P value: 0.010, 0.004 − 0.0015, < 0.01), non-depression (β, 95%CI, P value: 0.004, 0.000-0.007, 0.04), the 2nd quartile of Lutein + Zeaxanthin (β, 95%CI, P value: 0.008, 0.002–0.015, 0.01), and the highest quartile of vitamin D2 + D3 (β, 95%CI, P value: 0.008, 0.002–0.015, 0.01) subgroups.
In terms of execution ability, theobromine was associated with DSST score among age > 60 and ≤ 70 years old (β, 95%CI, P value: 0.011, 0.005–0.017, < 0.01), non-Hispanic white race (β, 95%CI, P value: 0.011, 0.004–0.018, 0.01), collage graduate or above (β, 95%CI, P value: 0.011, 0.003–0.019, 0.01), never-smoker (β, 95%CI, P value: 0.020, 0.010–0.031, < 0.01), overweight (β, 95%CI, P value: 0.010, 0.004–0.016, < 0.01), no history of stroke (β, 95%CI, P value: 0.010, 0.004–0.015, < 0.01), hypertension (β, 95%CI, P value: 0.012, 0.005–0.018, < 0.01), non-diabetes (β, 95%CI, P value: 0.012, 0.005–0.018, < 0.01), and depression (β, 95%CI, P value: 0.016, 0.008–0.024, < 0.01) subgroups. With respect to nutrition subgroups, theobromine was also positively associated with DSST among the 3rd (β, 95%CI, P value: 0.016, 0.002–0.031, 0.02) and 4th (β, 95%CI, P value: 0.009, 0.003–0.015, 0.01) quartile of energy intake. In the highest quartile of protein (β, 95%CI, P value: 0.017, 0.005–0.029, 0.01), lutein + zeaxanthin (β, 95%CI, P value: 0.027, 0.015–0.038, < 0.01), vitamin B12 (β, 95%CI, P value: 0.030, 0.017–0.043, < 0.01), and vitamin D2 + D3 (β, 95%CI, P value: 0.030, 0.016–0.044, < 0.01) subgroups, dietary theobromine and DSST were also associated.
To further analyze any non-linear associations, we conducted a curve fitting analysis between theobromine intake and cognitive performance. A non-linear association was found, as presented in Supplementary Fig. 3.