AuthorsInsects and Mites: W. J. Bentley, UC IPM Program, Kearney Agricultural Center, Parlier;R. H. Beede, UCCE Kings Co.;K. M. Daane, Biological Control, UC Berkeley/Kearney AgriculturalCenter, Parlier T. J. Michailides, Kearney AgriculturalCenter, Parlier; B. L. Teviotdale,Kearney Agricultural Center, Parlier August 2004 B. B. Westerdahl, Nematology, UC Davis
Cellmarkbiopharma.comThe combined effects of L-theanine andcaffeine on cognitive performance and mood Gail N. Owen1, Holly Parnell1, Eveline A. De Bruin2, Jane A. Rycroft1
1Unilever Research and Development, Colworth House, Sharnbrook, Bedford, UK 2Unilever Food and Health Research Institute, Unilever R&D Vlaardingen, The Netherlands The aim of this study was to compare 50 mg caffeine, with and without 100 mg L-theanine, oncognition and mood in healthy volunteers. The effects of these treatments on word recognition,rapid visual information processing, critical flicker fusion threshold, attention switching and moodwere compared to placebo in 27 participants. Performance was measured at baseline and again 60min and 90 min after each treatment (separated by a 7-day washout). Caffeine improved subjectivealertness at 60 min and accuracy on the attention-switching task at 90 min. The L-theanine andcaffeine combination improved both speed and accuracy of performance of the attention-switchingtask at 60 min, and reduced susceptibility to distracting information in the memory task at both 60min and 90 min. These results replicate previous evidence which suggests that L-theanine andcaffeine in combination are beneficial for improving performance on cognitively demanding tasks.
Keywords: L-theanine, caf eine, cognitive performance, mood
associated with changes in subjective state, includingincreased feelings of well-being, energy, motivation, Black tea is traditional y associated with relaxation and self confidence, alertness and concentration.5 Direct refreshment. The active components in tea associated with comparisons of coffee and tea containing the same this revitalising effect are caffeine and L-theanine. Caffeine dose of caffeine indicate that both produce similar is a wel -known stimulant that makes up 2–5% of the dry weight of tea leaves; a typical serving of black tea contains consistent levels of arousal during the day and is less Published by Maney Publishing (c) W.S Maney & Son Limited around 40 mg caffeine depending on the blend and brewing likely to subsequently disrupt sleep.7 procedure.1 Peak plasma concentrations are at ained (γ-N-ethylglutamine) is one of 30–120 min after consumption.2 Caffeine crosses easily predominant amino acids found in species of the tea through the blood–brain barrier and, once in the brain, it plant Camellia and constitutes 1–2% of the dry weight indirectly increases neurotransmit er activity by blocking of tea,8–10 which corresponds to up to 60 mg per 200-ml the inhibitory actions of the neuromodulator adenosine.1 serving. Within tea, the predominant form of theanine is the L isomer (around 98% of total theanine).
attention specifically, have been shown at doses equal L-theanine crosses the blood–brain barrier within 30 to and lower than the amount of caffeine typically min and increases activity in the alpha frequency band found in a single serving of tea.3,4 Caffeine is also of the electroencephalogram (EEG) not only at a highdose (200 mg)11,12 but also at a lower dose (50 mg),13suggesting that L-theanine even in low amounts (such Correspondence to: Eveline A. de Bruin, Unilever Food and Health Research Institute, PO Box 114, 3130 AC Vlaardingen, The Netherlands.
as present in 2–3 cups of tea) contributes to a relaxed, Tel: +31 10 460 5831; Fax: +31 10 460 5794; yet alert, feeling at rest. Furthermore, a recent study Received 14 November 2007, revised manuscript accepted 20 May 2008 has shown that L-theanine improves attention-related 2008 W. S. Maney and Son Ltd
Owen et al. Combined effects of L-theanine and caffeine on cognitive performance and mood anticipatory alpha during an attention task, indicating week). Additionally, they were free from over-the- improvement of the ability to focus attention when counter or prescribed medication (with the exception mentally active.14 of the contraceptive pill).
Despite the common co-consumption of caffeine L-theanine in tea, there is virtually no research that explicitly examines the interaction of the two L-theanine and caffeine solutions were prepared by substances. In a recent study, a combined dose of 150 adding 100 mg L-theanine and/or 35 mg caffeine to 250 mg caffeine and 250 mg L-theanine significantly ml Lipton Iced Tea (LIT). LIT contains 15 mg caffeine improved simple reaction time but also speed of from tea powder (thus total caffeine was 50 mg/serving).
numeric working memory and word recognition, and L-theanine was administered in a synthetic form of pure accuracy of sentence verification in the absence of any L-theanine known as Suntheanine™ (Taiyo Kagaku Co.
effect of caffeine or L-theanine alone.15 Furthermore, Ltd, Yokkaichi, Japan), an odourless, white, crystal alertness and tiredness were also improved by L- powder with a slightly sweet taste. L-theanine and theanine and caffeine in combination. In another caffeine were added immediately before serving. The study investigating EEG during the execution of an placebo solution was 250 ml LIT with tea powder attention task, 50 mg caffeine and 100 mg L-theanine (including 15 mg caffeine) removed.
in combination desynchronised tonic alpha and improved accuracy at the behavioural level, whereas A cross-over, double-blind, randomised, placebo- caffeine or L-theanine alone did not.16 Additionally, controlled design was used. Participants completed a series of three test sessions and consumed one of the reduction in the alpha attention effect seen after three drinks during each session – placebo, caffeine, L- ingestion of caffeine alone.
theanine+caffeine. Treatments were allocated using a The aim of the current study was to explore further Latin square design such that the order of treatments the combined effects of L-theanine and caffeine on was counterbalanced across participants. Sessions cognitive function. As caffeine and L-theanine are were separated by at least 7 days. Each session was usually consumed together in tea, the vehicle used for standardised according to the procedures below and administration was a tea-based soft drink (iced tea).
lasted approximately 2 h.
The combined effects of L-theanine+caffeine, andcaffeine alone were compared to placebo using tests of attention, arousal, memory and mood.
Prior to the start of the study, participants were given theopportunity to familiarise themselves with the tests of Subjects and methods
cognition and mood to be used during the study. On theday of the experiment, they arrived at the laboratory having consumed their normal breakfast and having The ethics committee at Unilever Research and abstained, from 9 p.m. the previous night, from caffeinated Published by Maney Publishing (c) W.S Maney & Son Limited approved the study and participants gave written medications, and mushrooms (the edible Bay bolete informed consent. A power calculation based on the contains L-theanine). On arrival, they completed baseline results of a simple reaction-time task in a pilot study tests of mood and cognitive performance in the order (unpublished data) suggested that a study of 27 shown below. Following completion of the baseline test participants should have sufficient power to detect a battery, they were given 5 min in which to consume the test difference in change in mean reaction time from drink. Further test batteries were complete 60 min and 90 baseline of 20 ms between treatments with 80% min from the end of the 5-min ingestion period. When the certainty at a significance level of 5%. Therefore, a participants were not being tested, they were allowed to total of 27 participants (13 females, 14 males) were read quietly.
recruited from employees at Unilever. They had amean age of 28.3 ± 5.34 years and a mean body mass Test battery index (BMI) of 25.8 ± 4.65 kg/m2. All reported being The test battery consisted of a number of tests of free of neurological disorders, were in general good mood and cognition. The choice of tests was based on health, and had an average State-Trait Anxiety Inventory the study by Haskell et al.15 and a pilot study (STAI) score of 35.4 ± 5.74. All participants were regular (unpublished results). The tests took about 14 min to caffeine consumers (9.6 ± 5.51 servings of caffeine per complete in total and were always performed in the Owen et al. Combined effects of L-theanine and caffeine on cognitive performance and mood same order. They are described below in the order in task-switching tests, the participant was required to which they appeared in the test battery.
characteristics of the stimulus pair for each trial.
Word recognition When the letter and number appeared in red, the At the start of the test battery, 15 words matched for participant was required to attend to the letter and frequency and concreteness were presented in sequence press a designated key only if the letter was a vowel.
for 1 s each with a 1-s ISI. At the end of the test battery, When the two items appeared in purple, participants these original words (old words) plus 15 distracter words responded only if the number was even. Despite the (new words) were presented, one at time, in randomised predictable pattern of task switching, performance of order. For each word, the participant had to indicate such tasks is typically poorer on trials when a switch is whether it was in the original list by pressing a designated required compared to performance when no switch is key as quickly as possible. Mean reaction times and accuracy were recorded. The total duration of word In the current study, the test duration was 5 min and presentation was 30 s and the duration of the recognition average response time, number of correct responses, test was 1 min.
number of missed targets and numbers of false alarmswere recorded. Performance for the trials requiring a Rapid visual information processing (RVIP) switch (i.e. first stimulus pair to appear in either red or A single digit (between 1 and 9) was presented in the purple following three successive stimuli in the centre of a computer screen in a continuous stream.
alternative colour) were also analysed separately.
Each digit was displayed for 600 ms with no intervalbetween stimuli. The participant was required to press Bond–Lader mood questionnaire a designated key as soon as possible following a Mood state was measured using the Bond–Lader sequence of either three consecutive odd or even analogue mood rating scale.19 The scale comprises 16 numbers. The task continued for 4 min and a target visual analogue scales (VAS) anchored by adjective occurred on 12.5% presentations. Mean reaction pairs (e.g. tense/relaxed). Participants place a vertical times, the number of correct responses, missed targets mark on the VAS to represent how they are feeling in and false alarms were recorded.
relation to the dimension defined by the adjectives. Bycombining scores for each adjective pair, it is possible Critical flicker fusion test (CFFT) to measure three distinct mood factors – alertness, Participants were required to discriminate flicker from contentedness, and calmness.
fusion, and vice versa, in a set of light-emitting diodes.
The diodes were held at a constant fixation distance.
Data analysis Scores for cognitive performance and mood at 60 min ascending (flicker to fusion) and three descending and 90 min post-ingestion were calculated as change (fusion to flicker) scales. The mean of these three from baseline. The data were subjected to a mixed ascending and descending presentations gives the model analysis of variance with ‘participant' as a threshold frequency in Hertz. The duration of the task random effect and ‘treatment', ‘gender', ‘BMI' and Published by Maney Publishing (c) W.S Maney & Son Limited ‘STAI anxiety score' as fixed effects. Omnibus tests ofthe main effects of treatment were followed by planned comparisons, using Tukey–Kramer tests to Participants were presented simultaneously with a assess the efficacy of the individual treatments in letter (A, E, G, I, K, M, R or U) and number (2, 3, 4, comparison with the placebo.
5, 6, 7, 8 or 9) one either side of, and at a set distancefrom, a central fixation. The position of the letter andnumber varied randomly between trials but with equal probability that either could occur to the left or rightof the fixation point. The number and letter were Word recognition chosen at random with the only constraint being that There was no evidence of a treatment effect for response neither had appeared in the preceding trial.
time for either new or old words. Average response time The stimuli appeared together for 1 s in either a red did decrease significantly between 60 min and 90 min, or purple font. The colour of the font changed in a but this occurred across all treatments.
fixed sequence every fourth presentation (i.e. three red There were no treatment effects for the number of followed by three purple and so forth). As in most correctly identified original (old) words, but there was Owen et al. Combined effects of L-theanine and caffeine on cognitive performance and mood Figure 1 Change in number of correctly identified ‘new'
Figure 2 Change in response time in the attention-
words in the word recognition task
a main effect of treatment for the number of correctly times were faster in both the L-theanine+caffeine and identified new words (F(2,108) = 6.14; P < 0.01). In caffeine conditions at 60 min but not 90 min.
the L-theanine+caffeine condition only, the number of There was a treatment by time interaction for the correctly identified words was greater at 60 min and 90 number of correct responses (F(2,115) = 49.34; P < min than at baseline (Fig. 1).
0.001). A direct comparison of L-theanine+caffeine andcaffeine showed that the number correct increased Rapid visual information processing (RVIP) significantly in the L-theanine+caffeine condition after There were no significant treatment effects for this 60 min (P < 0.001), and at 90 min for the caffeine only task. The number of false alarms decreased at 60 min condition (P < 0.01; Fig. 3).
and 90 min but there were no significant differences The response time for trials requiring a switch between treatments.
followed the same pattern as that seen for overallperformance of this task and the response time Critical flicker fusion test (CFFT) decreased relative to baseline in all conditions.
There was a significant decrease in arousal between 60 Although there were no significant treatment effects, min and 90 min across all treatments. Although there response times were fastest in the L-theanine+caffeine were no significant treatment effects, the decrease in arousal at 60 min appeared to be slightly reduced inboth Bond–Lader mood questionnaire There was a main effect of treatment for the composite alertness (F(2, 120) = 4.08; P < 0.05). Alertness There was a decrease in response time in all conditions increased in the caffeine condition relative to placebo (P that may be indicative of a practice effect (Fig. 2).
< 0.01). L-theanine+caffeine also appeared to increase Published by Maney Publishing (c) W.S Maney & Son Limited There were no treatment effects although response alertness compared to placebo, but this was not Figure 3 Change in the number of correct responses in the attention-switching task
Owen et al. Combined effects of L-theanine and caffeine on cognitive performance and mood statistically significant. There was a decrease in calmness combined.15 However, in this study, the doses used between 60 min and 90 min post-ingestion, but this was were significantly higher (150 mg caffeine, 250 mg L- common to all treatments. There were no treatment or theanine) than those in the current study. The current co-variate effects for the contentedness scale.
absence of effects on RVIP in combination with thetendency for both caffeine and L-theanine+caffeine toprotect against the decline in arousal (as measured by CFFT) seen in the placebo condition suggest that thecurrent study may have been underpowered for Both the caffeine and L-theanine+caffeine treatments demonstrating these effects.
produced significant changes in performance relative Both caffeine and L-theanine+caffeine increased to placebo. After caffeine ingestion, response speed subjective alertness relative to placebo, although the and accuracy improved in the attention-switching latter was non-significant. Improvements in objectively task, at 60 min and 90 min, respectively. The L- measured attention without concurrent changes in theanine and caffeine combination also produced subjective alertness are common and have been faster responses and improved accuracy during this reported for other studies as well (see, for example, task but this occurred simultaneously after 60 min.
Hindmarch et al.7 and Lieberman et al.20). In the The combined treatment also led to an increase in the current study, this may be explained by the ‘dampening' number of correctly identified new words in the word effect of L-theanine on caffeine as previously suggested.
recognition task. This effect was significant relative to For example, one study showed an inhibitory effect of placebo at both 60 min and 90 min post-ingestion.
L-theanine on caffeine stimulation as evaluated by EEG RVIP performance was not significantly altered by in the rat when the dose of L-theanine was five times either treatment and, although caffeine increased that of caffeine, but not when the level of L-theanine subjective ratings of alertness at 60 min, there was no was less than that of caffeine.21 A human study recently change in arousal as measured by CFFT.
found an interactive effect of a combination of caffeine The task most significantly affected by both treatments and L-theanine indicating that L-theanine antagonised was the attention switching task. Given that this was the the caffeine-induced rise in blood pressure but had no most cognitively demanding task of the test battery, this significant effects on blood pressure when ingested is not entirely unexpected. What was interesting is that although both caffeine and the L-theanine+caffeinecombination improved response speed and accuracy, itwas only the latter that improved them simultaneously. L- theanine is known to improve attention-related antici-patory alpha14 and caffeine, as demonstrated in the current This study represents a systematic assessment of L- study, can increase alertness. Thus, the combination was theanine and caffeine interactions and replicates associated with an improved ability to focus attention, previous work which suggests a synergy between these whilst caffeine alone lead to an improvement in basic two tea components. In particular, L-theanine and response speed or accuracy but not both.
caffeine in combination appear to improve, signifi- Published by Maney Publishing (c) W.S Maney & Son Limited In the word recognition task, response speed for cantly, aspects of memory and attention to a greater correctly recognised words (old words) tended to extent than caffeine alone. The levels of L-theanine improve after 60 min but returned to baseline by 90 and caffeine used here are higher than those found in min. This effect was not significant and did not vary real tea beverages and also in the reverse ratio as in tea across treatments. There was also no treatment effect the caffeine level is typically higher than that of L- with regard to the number of correctly identified old theanine (approximately 2:1). Further studies should words. However, whilst there was a decrease in the therefore be aimed at establishing a full dose-range number of correctly rejected distracter (new) words effect for L-theanine and caffeine in combination.
post-treatment in the placebo and caffeine conditions,there was no impairment in the L-theanine+caffeinetreatment. Participants were, therefore, less susceptible to distraction following ingestion of L-theanine and Fredholm BB, Battig K, Holmen J, Nehlig A, Zvartau EE. Actions caffeine in combination.
of caffeine in the brain with special reference to factors that There was no effect of either treatment on RVIP contribute to its widespread use. although improved RVIP accuracy has been found Passmore AP, Kondowe GB, Johnstone GD. Renal andcardiovascular effects of caffeine: a dose response study. previously for both caffeine and L-theanine+caffeine Owen et al. Combined effects of L-theanine and caffeine on cognitive performance and mood Durlach PJ. The effects of a low dose of caffeine on cognitive tea, and its effect on mental state. Smit HJ, Rogers PJ. Effects of low doses of caffeine on cognitive 14. Gomez-Ramirez A, Higgins BA, Rycroft JA et al. The deployment performance, mood and thirst in low and high caffeine consumers.
of intersensory selective attention: a high-density electrical mapping study of the effects of theanine. Mumford GK, Evans SM, Kaminski BJ, Preston KL, Sannerud CASK, Griffiths RR. Discriminative stimulus and subjective 15. Haskell CF, Kennedy DO, Milne AL, Wesnes KA, Scholey AB.
effects of theobromine and caffeine in humans.
The effects of L-theanine, caffeine and their combination on cognition and mood. Hindmarch I, Quinlan PT, Moore KL, Parkin C. The effects of 16. Kelly SP, Gomez-Ramirez M, Montesi JL, Foxe J. Synergistic black tea and other beverages on aspects of cognition and effects of L-theanine and caffeine in combination affect human psychomotor performance. recognition as evidenced by oscillatory alpha-band activity and Hindmarch I, Rigney U, Stanley N, Quinlan P, Rycroft J, Lane J. A attention task performance. J Nutr 2008; In press.
naturalistic investigation of the effects of day-long consumption of 17. Wylie GR, Javitt DC, Foxe JJ. Cognitive control processes during tea, coffee and water on alertness, sleep onset and sleep quality.
an anticipated switch of task. 18. Wylie GR, Javitt DC, Foxe JJ. The role of response requirements in task Cartwright RA, Roberts EAH, Wood DJ. Theanine an amino acid switching: dissolving the residue. Neuroreport 2004; 15: 1079–1087.
of N-ethyl amide present in tea. 19. Bond A, Lader M. The use of analogue scales in rating subjective Finger A, Kuhr S, Engelhardt UH. Chromatography of tea feelings. Br J Psychol 1974; 47: 211–218.
20. Lieberman HR, Wurtman RJ, Emde GG, Roberts C, Coviella IL.
10. Wickremasinghe RL. Tea (in food research). The effects of low doses of caffeine on human performance and 11. Juneja LR, Chu D, Okubo T, Nagato Y, Yokogoshi H. L-theanine 21. Kakuda T, Nozawa A, Unno T, Okamura N, Okai O. Inhibiting – a unique amino acid of green tea and its relaxation effects in effects of theanine on caffeine stimulation evaluated by EEG in the 12. Song CH, Jung JH, Oh JS, Kim KS. Effects of theanine on the 22. Rogers PJ, Smith JE, Heatherley SV, Pleydell-Pearce CW. Time for release of alpha brain waves in adult males. Korean J Nutr 2003; tea: mood, blood pressure and cognitive performance effects of caffeine and theanine administered alone and together.
13. Nobre AC, Rao A, Owen G. L-theanine, a natural constituent of Published by Maney Publishing (c) W.S Maney & Son Limited
This Application Note contains important information about this product AFFINILUTE MIP – β-agonists Part Number AFFINILUTE MIP β-agonists 25 mg/10 mL AFFINILUTE MIP β-agonists 25 mg/3 mL Molecularly imprinted polymers (MIPs) are a class of highly cross-linked polymers- engineered to bind one target compound or a class of structurally related target compounds with high selectivity. Selectivity is introduced during MIP synthesis inwhich a template molecule, designed to mimic the analyte, guides the formation of specific cavities or imprints that aresterically and chemically complementary to the target analyte(s). It is therefore critical for analysts to use the methodologydescribed below when using this phase. Conventional generic methodologies employed with conventional SPE chemistries(e.g., reversed-phase C18) will yield sub-optimal results when employed with this phase.