methods in nutrition

A study of caffeine in tea

I. A new spectrophotometric micro-method

II. Concentration of caffeine in

various strengths, brands, blends,

Downloaded from ajcn.nutrition.org by guest on April 7, 2014

and types of teas1-3
Daniel S. Groisser

ABSTRACT A new spectrophotometric micro-method for the determination of caffeine in tea

is described. This method is then used to evaluate the caffeine content of a variety of brands and
blends of bagged and loose hot tea prepared in different strengths and by different brewing
methods. In addition, the caffeine content of instant tea, ice tea, and Mr. Coffee automatic tea is
evaluated. Am. J. Clin. Nut,. 31: 1727-1731, 1978.

Tea, next to water, is the most frequently The purpose of this study is to evaluate the
consumed beverage in the world. Conse- caffeine content of various brands and
quently, the ingestion of caffeine, an impor- strengths of teas, prepared by the dipping
tant ingredient of tea as well as coffee, has method or by brewing for consumption by
become a daily habit for many. Because med- tea drinkers.
ical literature concerning the physiological In addition, a new improved spectropho-
effects of caffeine has shown the drug to be tometric micro-method has been developed
an important contributing factor in some pa- to analyze the caffeine concentration in tea.
tients with heartburn, diarrhea, and peptic This method and the evaluation of the caf-
ulcer disease, and because of its well-known feine content of tea is described.
effect as a central nervous system stimulant,
a study of the caffeine content of a cup of An improved method for caffeine
various brands and strengths of tea has been determination In tea
performed (1-5). Several spectrophotometric methods for analyzing
The question, “how much caffeine is in a caffeine concentrations in liquids are available (6-9).
cup of tea?” is complex; the number of vari- Although accurate, these methods are time-consuming
ables is great. There are a wide variety of and/or use large volumes of material. The new proce-

brands and blends of bagged and loose tea,

different brewing methods, and personal pre- From the Department of Pathology and Laborato-
ferred strengths. As yet the literature does not ries, Mountainside Hospital, Montclair, New Jersey.
2Supported in part by a grant from the New Jersey
contain information concerning the quantity
Academy of Science.
of caffeine ingested in tea that attempts to 3This work was presented in part at the Annual
consider all or many of these diverse ele- Meeting of the New Jersey Academy of Science, April 2,
ments. 1977.

The American Journal of Clinical Nutrition 31: OCTOBER 1978, pp. 1727-173 1. Printed in U.S.A. 1727
1728 GROISSER

dure, partially an outgrowth of the Axehrod-Reichenthal TABLE 2

method (6), is inexpensive, uses less reagents, is easily Reproducibility of color matching of tea to
performed, is less time-consuming (20 min per aliquot standards-as represented by caffeine content mg/180
compared to 60 or more min) and is highly accurate. mr
In this procedure caffeine is isolated from tea at pH
Trial no. Brand Weak Medium Strong
8 to 9 (20 C) by extraction into the relatively nonpolar
organic solvent benzene. The extraction is augmented by 1 Tetley 18 48 70
the additioi of NaCl, which helps to break the emulsion 2 Tetley 19 48 71
formed by the oils present in the beverage. The caffeine 3 Tetley 18 48 70
is further isolated in H2SO4, separating it from other 4 Tetley 18 48 70
constituents of tea such as theobromine and theophylline. a Each trial was carried out by using different tea bags
The optical density of the H2S04 layer is read on a on each of 4 different days. Three aliquots per sample
spectrophotometer and the amount of caffeine present
in the beverage is calculated. were analyzed and the mean value is reported.

cuvet more readily without damaging the spectropho-

Method
tometer, and the procedure is less time-consuming.
Materials. All chemicals and equipment used are of
analytical grades and are used without further purifica-
tion. Determination of caffeine concentration In
Caffeine standards of 2.5, 5.0, 7.5, and 10.0 cg/m1 are tea
prepared with 5 N H2S04.

Downloaded from ajcn.nutrition.org by guest on April 7, 2014

A model DB spectrophotometer is used. (Beckman
Methods
Instruments, Inc., Fullerton, Calif. 92634).
Using the previously described method, various
strengths and brands of tea were studied for caffeine
Procedure
content. The comparison was made of teas prepared in
Make 180 ml (6 ounces) of tea to desired strength. two ways: 1) on the basis of strength (color); and, 2) on
Adjust pH of the beverage to 8 to 9 using 1 N NaOH at the basis of package directions for brewing.
20 C. Pipet 0.1 ml of tea into a 20 ml teflon capped tube.
Pipet 10 ml benzene into the tube and add about 0.5 g Criteria for “weak,” “medium,” and “strong” cups of tea
NaCI. Shake tube for I min using a vortex super mixer.
The choice of a weak, medium, and strong cup of tea
Centrifuge for 10 min at 3500 rpm. Carefully pipet S ml
is highly variable and individualistic. The popular con-
from the benzene (top) layer into a teflon capped test
cept is that most people choose their strength of tea on
tube containing 5 ml of 5 N H2SO4. Shake for 30 sec.
the basis of color development. To verify this assump-
Centrifuge for 5 min. Carefully pipet some of the H2S04
tion, 700 people were polled as to the criteria they use to
(bottom) layer into a quartz cuvet. Read the optical
choose their strength of tea. Of those polled 95% choose
density of the sample versus blank H2S04 cuvet at 273
their strength of tea by the color of the beverage. Utiliz-
nm. Calculate the caffeine concentration as mg/h80 ml
ing this information, 20 different color gradations of a
of beverage.
single brand of tea were made. (Black tea, which pro-
duces the copperish color most Americans are accus-
Reproducibility
tomed to, was the blend used.) A survey in which 205
Three recovery experiments in which 10, 25, and 50 hospital personnel were asked to visually select one weak,
mg of caffeine were added to tea were performed. Re- one medium, and one strong cup often from the 20 cups
covery was better than 95% (see Table I). often was conducted. Of the 205 people 90% chose color
concentrations that were almost identical in each of the
Advantages weak, medium, and strong range. The color of the weak,
medium, and strong tea that received the most frequent
The described method has the following advantages:
positive responses in each of the categories was used as
Sample and reagent quantities have been reduced, the
the standard for the caffeine experiment.
procedure utilizes the relatively non-volatile H2S04
which allows samples to be pipeted into and out of the Tea prepared by color comparison

Tea bags or porous metal tea bags containing 2.35 g

TABLE 1 of loose tea leaves (the amount of tea found in most
Recovery of caffeine from tea (mg/h80 ml)a bagged brands) were dunked at a uniform rate into
Percent recov
uniform glass jars containing boiling water. Against a
Tea plus caffeine added Ideal recovery Actual recovery uniform background, weak, medium, and strong cups of
cry
tea were made by color matching with these standards.
ing
The standards were in the same type of jar and were at
Tea+ 0 48
the same temperature as the samples. Black and Oolong
Tea+l0 58 57 98
teas were made in this fashion. The reproducibility of
Tea+25 73 72 99
this color matching was excellent and is shown in Table
Tea+50 98 95 97
2. Green tea, which yields a pale green color, cannot be
a Three aliquots per sample were analyzed and the
compared with the color of Black and Oolong. Instant,
mean value reported. ice, and Mr. Coffee Automatic teas were not made in
 CAFFEINE IN TEA 1729

this fashion. These latter teas were analyzed for caffeine of all the teas tested by package directions.
content only by following package directions for prepa-
Tables 3 and 4 include the type of tea (hot,
ration.
ice, instant, or Mr. Coffee Automatic tea), the
Tea prepared by package instruction type of packaging and the blend of tea (Black,
In addition, all teas tested were made by standard Green, or Oolong).
package directions. Bagged tea, loose tea in porous metal The most commonly consumed domestic
bags, and unencased loose tea were left to steep for 4 brands-Lipton, Tetley, Salada, and Red
min with the jar covered. (Package directions on most
Rose-display a wide range of caffeine levels
brands call for a 3 to S min steep.) Instant, ice, and
automatic tea were made by individual package instruc-
per strength and package directions (Table
tions. 3); weak tea 18 to 45 mg; medium tea 48 to
84 mg; strong 70 to 107 mg and by package
Results directions 24 to 45 mg. For each strength,
Red Rose contains the most caffeine followed
The caffeine content of weak, medium, and by Salada, Lipton, and Tetley.
strong tea for each brand tested is listed in For each of the three strengths and package
Table 3. Table 4 includes the caffeine content directions, the domestic teas contain less caf-

Downloaded from ajcn.nutrition.org by guest on April 7, 2014

TABLE 3
Tea made by strengths-comparison of caffeine content mg/180 ml of domestic an d imported tea. Stren gths of tea
established by color comparison”

Brand Type Packed Blend Weak Medium Strong

Domestic teas
Red Rose Hot Bag Black 45 62 90
Salada Hot Bag Black 25 60 78
Lipton Hot Bag Black 25 53 70
Tetley Hot Bag Black 18 48 70

Imported teas
English Breakfast (Twinings) Hot Bag Black 26 78 107
English Breakfast (Twinings) Hot Loose in metal bag Black 39 84 90
Darjeeling (Twinings) Hot Bag Black 39 74 91
Formosa (Jacksons) Hot Loose in metal bag Oolong 42 65 78
a Three aliquots per sample were analyzed and the mean value is reported.

TABLE 4
Tea made by package directions-comparison of caffeine content (mg/180 ml) of domestic and imported teas”

Brand Type Packed Blend Four-mm steep

Red Rose Hot Bag Black 46

Salada Hot Bag Black 40
Lipton Hot Bag Black 38
Tetley Hot Bag Black 25
English Breakfast (Twinings) Hot Bag Black 52
English Breakfast (Twinings) Hot Loose and metal bag Black 58
English Breakfast (Twinings) Hot Loose Black 77
Daijeeling (Twinings) Hot Bag Black 65
Formosa (Jackson) Hot Loose and metal bag Oolong 42
Chinese (Hong Kong) Hot Bag Green 36
Pan fried Hot Loose and metal bag Green 81
Lipton Instant Loose 62
Nestea Instant Loose 48
Lipton Ice water sugar and Loose 76
lemon
Nestea Ice water sugar and Loose 67
lemon
Lipton Mr. Coffee automatic Loose 71
Tetley Mr. Coffee automatic Loose 68
a Three aliquots per sample were analyzed and the mean value is reported.
1730 GROISSER

feine than the more expensive imported black third the quantity of caffeine found in a cup
teas (Tables 3 and 4). Mean concentration of coffee or a strong cup of tea. The advice of
reveals 28 versus 37 mg for weak, 56 versus parents and occasionally of physicians to
75 mg for medium, 77 versus 92 mg for strong “take a weak cup of tea” to “settle your
and 37 versus 63 mg for package directions. stomach,” therefore, does not result in the
When package directions were followed for introduction of a physiologically significant
all hot teas in Table 3, the caffeine content amount of caffeine into the body (if indeed
was approximately between that of a weak the patient restricts himself to one or even
and medium cup (Table 4). For all hot teas two cups of weak tea). Likewise, those who
tested, mean concentration of caffeine was: prepare tea according to package instructions
weak 32 mg, medium 66 mg, and 46 mg as obtain a level of caffeine that is not signifi-
prepared by package directions. cantly greater than a weak cup of tea.
When the leaves of loose English Breakfast It is of interest that the tea bag or porous
Tea are encased in the metal tea bag, the metal container tends to retard some of the
caffeine yield a similar to that of bagged caffeine leakage into the cup-thus tea pre-
English Breakfast Tea (Tables 3 and 4). How- pared with loose tea leaves delivers more
ever, made by package directions, unencased caffeine for the same degree of color devel-
and time exposure

Downloaded from ajcn.nutrition.org by guest on April 7, 2014

loose English Breakfast leaves yielded a con- opment to the water.
siderably higher caffeine level than the Eng- Although decaffeinated products are avail-
lish Breakfast in paper or metal tea bags able for coffee, it has not been deemed fea-
made under the same condition. sible yet to decaffeinate tea. In addition, be-
Instant tea, (mean 55 mg), has a similar cause caffeine contributes significantly to the
caffeine content to that of a medium cup of flavor of the tea, its removal would render
hot domestic tea (mean 56 mg). Ice tea, (mean the tea unpalatable to the tea drinker (14).
72 mg), and Mr. Coffee Automatic tea, (mean In any experiment regarding caffeine in
70 mg), yield similar caffeine concentrations tea, the size of the cup is important. Many
that compare with that of a strong cup of the experiments have been performed with 5 to
hot domestic teas (mean 77 mg). 5Y2 ounce cups. Other experiments have failed
to mention cup size. One author has sug-
Discussion gested that perhaps a 150 ml cup be used as
In addition to tea, caffeine may be ingested a standard (13). We have chosen a 6 ounce
in many forms as in the over-the-counter cup as a standard throughout the entire in-
medications such as APC (32 mg per tablet), vestigation.
prescription medication such as Cafergot (100 Caffeine is not the only xanthine present
mg per tablet), in cocoa (50 mg/6 ounces), in in tea. However, only small quantities of
milk chocolate (3 mg/ounce), coca cola (40 theobramine (2 mg per cup) and theophylline
to 72 mg/12 ounces), NoDoze tablets (100
(1 mg per cup) have been detected (13, 15).
mg per tablet), as well as in coffee (100 to 150
mg/6 ounce cup) (10-12). The amount of
Conclusion
ingested caffeine that produces increased A new method for measuring the quantity
awareness and acuity, possibly to the point of of caffeine in tea and other beverages has
restlessness or sleeplessness in some people,
been described.
varies with the individual. Approximately
Utilizing this new technique, the caffeine
100 to 150 mg is considered a physiological
content of different brands and strengths of
dose and may produce side effects (5). tea has been evaluated. The relationship of
There has, in general, been scanty infor-
the caffeine content of tea as contrasted to
mation concerning the amount of caffeine in
the caffeine content of other beverages has
various strengths of tea (13), and certainly no
been discussed.
definitive information exists in the literature
The author thanks R.
Sandhu, Ph.D., Biochemist at
concerning the quantity of caffeine in weak
Mountainside Hospital,Montclair, New Jersey, for his
tea. The present experiment establishes that kind guidance and for his review of this manuscript. The
the so-called weak tea, as judged by its color author is also indebted to Ms. Sally Bailey for her
development, contains approximately one- constant encouragement and advice. The author should
 CAFFEINE IN TEA 1731

also like to thank F. Watson, M.D., Director of Moun- biological material. J. Pharmacol. Exptl. Ther. 107:
tainside Hospital Laboratories for providing the labora- 519, 1953.
tory facilities for this research. 7. Association of Official Analytical Chemists. Official
Methods of Analysis. 11: 239, 1970.
References 8. Association of Official Agricultural Chemists. Offi-
1. GOODMAN, L. S., AND A. GILMAN. The Pharmaco- cial methods of analysis. 10: 215, 1965.
logical Basis of Therapeutics. New York. 1975, p. 9. NEWTON, J. M. J. Assoc. Off. Anal. Chem. 52: 1133,
367. 1969.
2. COHEN, S., AND G. H. BOOTH, JR. Gastric acid 10. Nutrition and the M.D. 9: 3, 1976.
secretion and lower esophageal sphincter pressure in 11. NAGY, M. Caffeine content of beverages and choc-
response to coffee and caffeine. New EngI. J. Med. olate. J. Am. Med. Assoc. 229: 337, 1974.
293: 897, 1975. 12. LEE, S. Better procedures for analyzing caffeine
3. ROTH, J. A., AND A. C. Ivy. The effect of caffeine could help better standards. Tea Coffee Trade J.
upon gastric secretion in the dog, cat and man. Am. 144: 26, 1973.
J. Physiol. 141: 454, 1944. 13. BURG, A. N. How much caffeine in a cup often and
4. BABKA, J. D., AND D. CASTELL. The effects of specific coffee. Tea Coffee Trade J. 1: 40, 1975.
foods on the lower esophageal sphincter. Digest. 14. MILLAN, D. J., D. J. CRISPIN AND D. SNAINE. Non-
Diseases 18: 391, 1973. volatile components of black tea and their contri-
5. GOLDSTEIN, A., G. KAISER AND R. WARREN. Psy- bution to the character of the beverage. J. Agr. Food
chotropic effects of caffeine in man. J. Pharmacol. Chem. 17: 717, 1969.

Downloaded from ajcn.nutrition.org by guest on April 7, 2014

Exp. Ther. 150: 146, 1965. 15. MARTINEK, R. G. Xanthines, tannins, and sodium in
6. AXELROD, J., AND J. REICHENTHAL. The fate of coffee, tea, and cocoa. J. Am. Med. Assoc. 158: 1030,
caffeine in man and a method for its estimation in 1955.