the thiamine content of herbs and medicinal plants

4
SCIENTIFIC EDITION The Thiamine Content of Herbs and Medicinal Plants* By ALEXANDER S. CHAIKELISt By means of a photofluorometric assay tech- nique, eliminating the elution phase, devised to investigate the thiamine content of plant material ranging from rhizomes and roots to barks, leaves, and flower tops, the thiamine concentration in herbs and medicinal plants was determined. A spot check analysis of 41 genera belonging to 20 distinct and separate botanical families showed a thiamine range from 125 to 2880 micrograms per 100 Gm. of dried substance. This vitamin B1 content was postulated as a possible modus operandi of the many infusions and de- coctions of medicinal herbs prescribed in cases of anorexia, dyspepsia, general debility, and nervousness. OR MANY CENTURIES the.use of plants in the alleviation and cure of bodily ills received wide attention but little official sanction until in the reign of King Henry VIII Parliament enacted what is now popularly known in England, “The Herba- lists’ Charter” (1). Healers, like Nicholas Culpepper (1616- 1654) in England and Samuel Thompson (1769-1843) in America, did much to ad- vance the study and practice of herbalism. Apart from the claims made by the herba- lists, the use of herbs as sources of drugs, like those described in articles and books on pharmacognosy and/or materia medica, has done much to justify their usefulness in medicine (2-7). With our present knowledge of the various sources of vitamins, can it be that the cumu- lative beneficial results produced by the use of herbs, exclusive of their medicinal proper- ties, might be the effects of increased vita- min intake? To obtain information along this line of thought, it was decided to make a spot check on some herbs and other plants, * Received July 9, 1946, from the Biology Department, The City College, and the Department of Industrial Hygiene, School of Public Health, Columbia University, New York City. t The author expresses his gratitude to Dr. Frederick B. Flinn, Director of the Department of Industrial Hygiene, School of Public Health, Columbia University, for his en- couragement, and to Hoffman-LaRoche, Inc., Nutley, N. J., for donations of thiamine. 343 emphasis to be on those which were and are, a t present, described as possessing tonic, stomachic, and stimulant actions in cases of dyspepsia, constipation, general debility, and nervousness. Since all higher plants are able to not only synthesize thiamine but to excrete it through the roots, it seemed logical to investigate the existence and quantity of thiamine in these and other parts of the dried plants that had recognized medicinal value. Samples of the herbs and medicinal plants were purchased in the dried state from a reliable distributor of such material. The plants so bought represented 41 genera belonging to 20 separafe botanical families. This number of specimens was considered to be an adequate sampling of the entire group of medicinals. EXPERIMENTAL Based on the idea of procedure for thiochrome determination described by Hennessy (8) and the digestion method of plant material to release the thiamine in the free state as well as the phosphory- lated form (cocarboxylase) suggested by Platt and Block (9), the thiamine content of the plant material was analyzed by the following method devised to fit the demands of the material under investigation and the requirement of a quick, reliable technique. A. Reagents Required.-The many reagents needed to effectively carry out the thiochrome determinations included those listed in the U. S. P., those enumerated by Platt and Block (9), and those, such as the needed standard solutions, described in (10). The digesting enzymes papain and clarase before use were treated in the following manner: A 5-Gm. batch of each enzyme is suspended in 50 cc. distilled water and shaken thoroughly. To this suspension is added 50 cc. isobutanol, and the mixture again shaken in a separatory funnel. Discard the super- natant isobutanol fraction, repeating the process until the isobutanol portion when oxidized with NaOH and KIFe(CN)B shows no evidence of thio- chrome when examined in a photofluorometer. The aqueous fraction is then treated with ethanol to precipitate the enzymes, filtered, and the residue dried in vucuo. The dry amorphous powder is then ready for use in the digestion phase of the treatment section of the procedure.

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Page 1: The thiamine content of herbs and medicinal plants

SCIENTIFIC EDITION

The Thiamine Content of Herbs and Medicinal Plants*

By ALEXANDER S. CHAIKELISt

By means of a photofluorometric assay tech- nique, eliminating the elution phase, devised to investigate the thiamine content of plant material ranging from rhizomes and roots to barks, leaves, and flower tops, the thiamine concentration in herbs and medicinal plants was determined. A spot check analysis of 41 genera belonging to 20 distinct and separate botanical families showed a thiamine range from 125 to 2880 micrograms per 100 Gm. of dried substance. This vitamin B1 content was postulated as a possible modus operandi of the many infusions and de- coctions of medicinal herbs prescribed i n cases of anorexia, dyspepsia, general debility,

and nervousness.

OR MANY CENTURIES the.use of plants in the alleviation and cure of bodily

ills received wide attention but little official sanction until in the reign of King Henry VIII Parliament enacted what is now popularly known in England, “The Herba- lists’ Charter” (1).

Healers, like Nicholas Culpepper (1616- 1654) in England and Samuel Thompson (1769-1843) in America, did much to ad- vance the study and practice of herbalism. Apart from the claims made by the herba- lists, the use of herbs as sources of drugs, like those described in articles and books on pharmacognosy and/or materia medica, has done much to justify their usefulness in medicine (2-7).

With our present knowledge of the various sources of vitamins, can i t be that the cumu- lative beneficial results produced by the use of herbs, exclusive of their medicinal proper- ties, might be the effects of increased vita- min intake? To obtain information along this line of thought, it was decided to make a spot check on some herbs and other plants,

* Received July 9 , 1946, from the Biology Department, The City College, and the Department of Industrial Hygiene, School of Public Health, Columbia University, New York City. t The author expresses his gratitude to Dr. Frederick B. Flinn, Director of the Department of Industrial Hygiene, School of Public Health, Columbia University, for his en- couragement, and to Hoffman-LaRoche, Inc., Nutley, N. J., for donations of thiamine.

343

emphasis to be on those which were and are, a t present, described as possessing tonic, stomachic, and stimulant actions in cases of dyspepsia, constipation, general debility, and nervousness. Since all higher plants are able to not only synthesize thiamine but to excrete i t through the roots, it seemed logical to investigate the existence and quantity of thiamine in these and other parts of the dried plants that had recognized medicinal value.

Samples of the herbs and medicinal plants were purchased in the dried state from a reliable distributor of such material. The plants so bought represented 41 genera belonging to 20 separafe botanical families. This number of specimens was considered to be an adequate sampling of the entire group of medicinals.

EXPERIMENTAL

Based on the idea of procedure for thiochrome determination described by Hennessy (8) and the digestion method of plant material to release the thiamine in the free state as well as the phosphory- lated form (cocarboxylase) suggested by Platt and Block (9), the thiamine content of the plant material was analyzed by the following method devised to fit the demands of the material under investigation and the requirement of a quick, reliable technique. A. Reagents Required.-The many reagents

needed to effectively carry out the thiochrome determinations included those listed in the U. S. P., those enumerated by Platt and Block (9), and those, such as the needed standard solutions, described in (10).

The digesting enzymes papain and clarase before use were treated in the following manner: A 5-Gm. batch of each enzyme is suspended in 50 cc. distilled water and shaken thoroughly. To this suspension is added 50 cc. isobutanol, and the mixture again shaken in a separatory funnel. Discard the super- natant isobutanol fraction, repeating the process until the isobutanol portion when oxidized with NaOH and KIFe(CN)B shows no evidence of thio- chrome when examined in a photofluorometer.

The aqueous fraction is then treated with ethanol to precipitate the enzymes, filtered, and the residue dried in vucuo. The dry amorphous powder is then ready for use in the digestion phase of the treatment section of the procedure.

Page 2: The thiamine content of herbs and medicinal plants

TA

BL

E C

OM

POSI

TE

SUM

MARY

OF T

HIA

MIN

E

CO

NTE

NT IN

V

ARI

OU

S DR

IED

HE

RB

S AN

D OT

HER M

ED

ICIN

AL

PLANTS

Av.

T

hiam

ine

Thi

amin

e T

hiam

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Com

mon

Nam

e A

grim

ony

Ang

ostu

ra

Bal

mon

y B

arbe

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Bay

Lau

rel

Bla

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lder

B

less

ed T

hist

le

Bon

eset

B

uchu

B

ugle

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alam

us

Car

away

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cari

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Cat

nip

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erv

Ciy

mom

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Tre

e C

ondu

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ande

lion

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ogw

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aica

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verf

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den

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enti

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Gro

und

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H

orse

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Page 3: The thiamine content of herbs and medicinal plants

SCIENTIFIC EDITION 345 B. Treatment of the Substance to Be Ana-

lyzed.-l. Place 10 Gm. of the substance, dried and finely ground, in a 250-cc. flask.

2. Add 90 cc. N/lO HzSO~, mix thoroughly, and then add 0.2 Gm.-batches each of the purified papain and clarase.

Now add 10 cc. of the M/10 sodium acetate- acetic acid buffer solution. Mix well (check PH with bromcresol green so that i t is 4.0; correct with either 10% HCl or 10% NaOH).

4. Stopper flask and heat the mixture a t 40-45' for from sixteen to eighteen hours.

5. Remove flask, add 0.2 cc. glacial acetic acid, and heat mixture to 80" to coagulate the proteins.

6. Cool flask to room temperature; centrifuge a 20-cc. portion, and save the clear supernatant aqueous layer for the analysis.

Oxidation (by means of alkaline ferricyanide) of thiamine t o thiochrome, which is extracted by isobutanol, is followed by measurement of the fluorescence in ultraviolet light of the thiochrome formed. The thiamine pyrophosphate also forms a fluorescent thiochrome, which is not, however, soluble in isobutanol-hence one reason for the enzyme treatment to hydrolyze the thiamine pyro- phosphate to thiamine.

With a Coleman Universal Spectrophotometer #11 adapted with an ultraviolet illuminator and the requisite filters for thiamine study, determine the degree of fluorescence using the substitute working standard of quinine sulfate (3 gamma per cc.). See details in the direction booklet (11).

3.

7.

8.

RESULTS

The values for the vitamin B1 content of the 41 genera of herbaceous medicinal plants are repre- sented in Table I. The thiamine, in micrograms per gram of dried substance, as listed in the table represents the mean value of three distinct and separate samples of, each plant studied.

Of the total number of plants studied in this spot check, it can be seen by reference to Table I that a group of seven medicinal plants contain thiamine in excess of 1500 ?/lo0 Gm. of substance. These seven plants arranged in order of the highest thia- mine content are: Prickly Ash (Xanthoxylum americanum Miller), Black Alder (Ilex verticilluta Linn.), White Ash (Fraxinus americana Linn.). Buchu (Barosm betulina Bartling & Wendland), Cherry (Prunus serotina Linn.), Quassia (Picrasma excelsa Panchon), and Dandelion (Taraxacum oficinale Weber) .

On the other hand, five genera show a thiamine content of less than 300 ~/100 Gm. of dried sub- stance. Arranged in order from the lowest to the highest concentration these five herbs are: Parsley (root) (Petroselinum sativum Hoffman), Southern- wood (Artemisia abrotanum Linn.), Agrimony (Agrimony eupatorium Linn.) , Strawberry (wild) (Fragaria virginiana Linn .) , and Bugleweed (Lycopus uni$orus Linn.). The remaining herbs

in the series range from 365-1154 y/100 e m . of substance.

DISCUSSION

Many factors influence the effectiveness with which a plant synthesizes vitamins. Therefore the values reported in this paper cannot be considered as constant but merely as indicative of the concentration of thia- mine contrary to the claim made that leaves, for instance, belonging to higher plants regardless of the botanical family source contain a constant thiamine concentration equal to 75 micrograms per 100 Gm.

It is an accepted fact that animals (in- cluding man) require a daily intake of thiamine because this vitamin is not stored in the organism although, as Westerbrink (12) reported, relatively higher concentra- tions are found in the liver, kidneys, heart, muscle, and brain than in the blood stream. The animal organism absorbs only as much thiamine as is needed for the time being, all excess is excreted and to a small extent destroyed in the body.

The physiological action of thiamine in the animal organism in the form of its pyrophosphoric acid ester is intimately concerned with carbohydrate, fat, and water metabolism. Thus, glycogen, the specific animal carbohydrate, requires the presence of this vitamin to be properly metabolized. The fat metabolism is in- fluenced by vitamin B1 only in so far as the synthesis of fat from carbohydrate is con- cerned. The water metabolism in the animal organism seems connected with the action of this vitamin, probably through the carbohydrate metabolism. Edema and water imbibitions in the heart and in other organs are symptoms of thiamine inade- quacy. Vitamin B1 deficiency affects, first of all, the emotions and the tonus of the nervous system, for this substance takes part in the conduction of nerve impulses as shown by Minz (13) that acetylcholine and vitamin B1 are liberated during impulse conduction.

Further symptoms of thiamine deficiency are loss of appetite (anorexia), lowered physical endurance associated with unusual susceptibility to fatigue, gastrointestinal

Page 4: The thiamine content of herbs and medicinal plants

346 JOURNAL OF THE AMERICAN PHARMACEUTICAL ASSOCIATION

disturbances (dyspepsia, constipation), mus- cular weakness, and a decrease of the blood pressure.

Attention is called to the fact that it is precisely for such discomforts that many herb infusions and decoctions, known as “tonics,” have been prescribed. The indica- tions are that these prescriptions materially benefited the ailing individual, a t least in a small measure, by providing him with readily absorbable thiamine and its phos- phorylated form a t a time when his system was in great need of these items.

SUMMARY

1. The thiamine content of a spot check analysis of 41 genera of herbs and medicinal plants was determined by means of the photofluorometric technique.

2. A photofluorometric assay technique was devised employing the suggestions of Hennessy (8) and those of Platt and Block

These 41 genera of herbs and medici- (9) -

3.

nal plants belonging to 20 separate botan- ical families showed a thiamine content ranging from 125-2880 micrograms per 100 Gm. of dried substance.

The rational basis for the use of herb infusions and decoctions in the form of “tonics” is suggested on the basis of the physiological action of thiamine in the animal organism.

4.

REFERENCES

(1) Warcl. 11.. “Herbal Manpl,” London. 1936. L. I < . . “A Modern Herbal.” ork, 1931. Wirth. E. H., “Pharmacog-

iiaaeiphia, 1936. , “Textbook of Pbarmacognosy.” adelphia, 1543 rhe Spice Handbook.” Chemical w York. 1945. A Textbook of Pharmacognosy.” ,ondon, 1945. ull. Torrey Bolan. Club. 73, 60-72

(8 ) Hennessy, D. J . , Ind. Eng . Chem., Anal . Ed . , 13,

(9) Platt. B. S.. and Block, C. F., Biochem. J . , 37, 436

\ - - I ” , .

2 16-17 ( 1941 ) .

f 19431. ~ ~ ~ ~ . , . (10) Hoffman-LaRoche. Inc., Nut!ey. N. J.. “Determina-

tion of Vitamin BI in Food Products (11 ) “Zirections for Model 11 ’Universal Spectropho-

tometer, D-142, Coleman Electric Co., Maywood. 111..

1944.

1945, p. 11.

560(1932). (12) Westerbrink, H. C. K . , Arch. nlcrland. physiol . , 17,

(13) Minz, B., C o m p f . r e n d . sot. b i d . , 127, 1251(1938).

Persistence of Penicillin in Saliva After Local Oral Application*

By MILTON GJELHAUG LEVINE and ROBERT E. HOYT

Penicillin for local application to the mouth area was administered in a chicle base chew- ing gum, in flavored parafKn, and in a mouth- wash. The retention time of penicillin in the saliva for each of these dosage forms with varying concentrations of the drug is recorded

in this report.

HE USE of penicillin in the buccal cavity Thas been suggested in hemolytic strepto- coccus infections, tonsillitis, Vincent’s disease (acute ulcerative gingivostomatitis), Ludwig’s angina, for lesions of the oral cavity involving loss of surface, and for prophylactic use in operative procedures of the mouth. Of all these, the most con- vincing therapeutic evidence has been ad-

*Received Aug. 12, 1946, from the Institute of Experi- mental Medicine. College of Medical Evangelists, 1.0s Angeles, Calif.

duced for the use of local preparations in Vincent’s angina.

Our own experiences and those reported by other investigators (1, 2, 9, 10) indicate that in most cases Vincent’s disease may be cured in from twenty-four to forty-eight hours by the local application of penicillin. The results for other infections so far have been inconclusive, although preliminary reports (2-5) indicate that Group A strepto- cocci may be reduced in number and even eliminated by local applications of peni- cillin. However, more satisfactory results may be obtained by parenteral administra- tion of the drug. Parenteral administration of penicillin has also proved to be of value in Ludwig’s angina (6, 7).

The treatment of acute ulcerative stoma-