To Evaluate and Compare the effects of First Generation Anti-Histamine

(Chlorpheniramine Maleate) and Second Generation Anti-Histamine (Loratadine)

on Isolated Trachea of Rabbit
G. A. Jalbani1, K. Aamir2, A.M. Shaikh3, M.A. Unar4 , R. Ashraf5, F. M. Soomro6
Liaquat University Hospital, Hyderabad/Jamshoro1, Department of Pharmacology and Therapeutics, BMSI, Jinnah Postgraduate Medical Centre2, Karachi,
Chandka Medical College3,4, Larkana, Fatima Jinnah Dental College5, Karachi and PNS Rahat,, Karsaz6, Karachi.

Abstract
Objective: The incidence of respiratory allergy has increased gradually over the past several years and current
estimates suggest that allergic rhinitis affects approximately 20% of the population. Large scales population sur-
veys indicate that upto 38% of patients with rhinitis have asthma. The allergic response in the airways is an
important pathogensis to cause bronchoconstriction owing to increased responsiveness of tracheo bronchial tree
to various stimuli and also causes the release of histamine and other chemical mediators from mast cells.
Histamine has been shown to be an important mediator of an allergic reaction in both the upper and lower res-
piratory airways. Chlorpheniramine maleate is a stable, most potent, sedative first generation anti-histamine and
is effective in the treatment of allergic disorders. Loratadine is a highly potent, non-sedating, long acting tricyclic,
second generation anti-histamine. It is indicated in allergic rhinits, chronic idiopathic urticaria and allergic
bronchial asthma. The purpose of study was to evaluate the antagonistic effects of chlorpheniramine maleate
and loratadine on histamine induced contractions in isolated trachea of rabbit and also to compare the effects of
first generation anti-histamine (chlorpheniramine maleate and second generation anti-histamine Loratadine).
Methods: In this study twenty-four experiments were performed on isolated trachea of rabbit, in the presence of
selected standard concentration of histamine dihydrochloride, antagonistic effects of various concentrations of
chlorpheniramine maleate (10-18 to 10-3 gm/ml) and loratadine from concentrations 10-18 to 10-3 gm/ml were
recorded by Polygraph Model 7B in terms of rate and amplitude.
Results: Chlorpheniramine maleate showed non-significant antagonistic effect from concentrations 10-18 to
10-3gm/ml in case of rate and 10-18 to 10-8 gm/ml in case of amplitude. Significant response showed from con-
centrations 10-8 to 10-3 gm/ml in case of rate (P<0.001) and 10-7 to 10-3 gm/ml in case of amplitude
(P<0.001)while, loratadine showed non-significant response from concentrations10-18 to 10-12 gm/ml in case of
rate and from concentration 10-18 to 10-14gm/ml in case of amplitude. Significant response observed from con-
centrations 10-11 to 10-3 gm/ml in case of rate and 10-13 to 10-3 gm/ml in case of amplitude.
Conclusion: It was concluded that chlorpheniramine maleate antagonized the histamine induced contractions
80.65% at concentration 10-3 gm/ml in case of amplitude and 11.35% at concentration 10-3 gm/ml in case of rate
and loratadine 76.82% in case of amplitude and 10.59% in case of rate (JPMA 54:556;2004).

Introduction postrema of brain.

Histamine is generally considered as the principal It occurs in plants as well as in animal tissues. Also
mediator of acute inflammatory process and allergic and occurs as a component of venoms and secretions from insect
anaphylactic reaction1, in both the upper and lower respira- sting.7 It is formed by decarboxylation of the amino acid L-
tory airways.2 It is the natural candidate as mast cell media- histadine. In mammalian tissues this reaction is catalyzed by
tor3, also has an important role in gastric acid secretion and the enzyme histadine decarboxylase. After formation it is
function as neurotransmitter and neuromodulator.4 It is either stored or rapidly inactivated contractions of smooth
found in all tissues, but high amount is found in lung, skin muscles by histamine is usually associated with depolariza-
and gastrointestinal tract, high concentration in mast cells or tion and increased adenosine phosphate discharge.
basophils.5 At least three storage sites of histamine are Histamine also contracts isolated fragments of smooth mus-
found. cles of cell membrane. In tracheal muscle an increase in
cAMP was detected.
i) In granules of mast cells and basophils, where it is
bound with heparin and cannot exert its effects or may be The anti-histamines are the classic H1 receptors
metabolized.6 mediated response blockers and competitively block the
receptor mediated response of a target tissue.8 They are
ii) In the mucosal layer of the gastrointestinal tract. divided into first and second generation anti-histamines.9
iii) A third fraction is held in the hypothalamus and area The main distinguishing points between first and second
 generation anti-histamines are that first generation gm, sodium dihydrophosphate 0.60 gm, potassium chloride
drugs are widely distributed throughout the body and are 1.85 gm, magnesium chloride 0.23 gm, distilled water 5000
more likely to block autonomic receptors and enter the cen- ml.
tral nervous system readily while the second generation
Preparation and Isolation of Tracheal Smooth Muscle
drugs are less lipid soluble and enter the central nervous
system with difficulty or not at al, so they show less seda- Twenty-four healthy adult rabbits male and female
tive and antichlorinergic effects.10 (non-pregnant), approximately 2 kg weight were selected
and used for the present study.18 The animals were sacri-
Chlorpheniramine Maleate
ficed, trachea was removed and transferred to petri dish
It is a stable, most potent, first generation, sedative anti- containing aerated (oxygenated) Kreb's bicarbonate solu-
histamine belongs to alkylamine group and acts by reversible tion, where it was cleaned of extraneous tissues.19 A chain
competitive antagonism at H1 receptors.11 It is widely distrib- of tracheal section was made by cutting several rings of car-
uted in the body, including passage into the central nerve sys-
tilages and tying them together loosely in such a way that
tem. It is used in the treatment of urticaria, allergic rhinitis,
muscles of two rings were at 180° to each other.20 Chain
angioedema, conjunctivitis, and in pruritic skin disorders. It is
was suspended vertically in an inner organ bath containing
a common ingredient of cough and cold preparations.
20 ml Kreb's bicarbonate solution with the help of tissue
Loratadine holder and connected to the Polygraph with the help of force
It is highly potent2, non-sedative and long acting tri- transducer.21 The nutritional solution was continuously aer-
cyclic11 second generation anti-histamine12 with selective ated 10-12 bubbles per minute and temperature was main-
competitive peripheral histamine H1 receptor antagonistic tained at 37°C. The preparations were allowed to equilibrate
activity13, belonging to the piperidine group and structural- in Kreb's bicarbonate solution for 90 minutes under an ini-
ly related to azatadine.14 It is less lipophilic, has no central tial lead of 2 gm.
nervous system activity and is essentially free of sedation.15
Bath solutions were changed after every 15 minutes.
It is proven to be effective in the treatment of seasonal aller-
The drugs were added in small quantities (1 ml) at each
gic rhinitis, chronic idopathic urticaria and allergic
bronchial asthma.16 interval to inner organ bath from lower concentration 10-18
gm/ml to higher concentration 10-3 gm/ml according to
Study Design experimental protocol and responses from each dilution
The purpose of study was to evaluate the antagonis- were recorded on Grass Polygraph under resting tension of
tic effects of chlorpheniramine maleate and loratadine on 1 gm.22
histamine induced contractions of isolated trachea of rabbit
and also to compare the effects of first generation anti-his- Methodology
tamine (chlorpheniramine maleate) and second generation First of all spontaneous contractions of tracheal
anti-histamine (loratadine). smooth muscles were recorded as a baseline. In group I, tis-
All experimental works were carried out in the sues were challenged with serial dilutions 10-18 to 10-3
Department of Pharmacology and Therapeutics, Basic gm/ml of histamine dihydrochloride and responses were
Medical Sciences Institute, Jinnah Postgraduate Medical recorded. From those responses, standard concentration of
Centre, Karachi. histamine dihydrochloride was selected, which had pro-
duced maximum response.
Material and Methods
In group II, tissues were challenged with serial dilu-
Preparation of Serial Dilution of Drugs
tions 10-18 to 10-3 gm/ml of chlorapheniramine maleate in
Serial dilutions were made by taking 1 ml of drug (1 the presence of selected standard concentration of histamine
ml/ml) and adding 9 ml of distilled water to make the ratio dihydrochloride and responses were recorded for each dilu-
1:9. In this way serial dilutions of different drugs were pre-
tion, respectively.
pared from concentrations 10-3 to 10-18 gm/ml.
In group III, tissues were challenged with serial dilu-
Nutrition Solution tions 10-18 to 10-3 gm/ml of loratadine in the presence of
In this vitro project Kreb's bicarbonate solution was selected standard concentration of histamine dihydrochlo-
used for the perfusion of isolated tracheal tissues.17 For the ride and responses were recorded for each dilution, respec-
preparation of 5 liters of Kreb's bicarbonate solution, fol- tively. After taking response of each concentration the tis-
lowing quantities of ingredients were used: sodium chloride sues were washed and giving rest for 3 minutes before
34.50 gm, sodium bicarbonate 10.50 gm, D-glucose 10.00 applying the next concentration.
 difference between the mean values of these two 11. Therapeutic drugs (Cetirizine). 2nd ed. 1999;1:C152-C156.

data showed that loratadine dose dependently antagonized 12. Druce HM, Thoden WR, Mure P, et al. Brompheniramine, loratadine and
placebo in allergic rhinitis: a placebo-controlled comparative clinical trial. J
the rate and amplitude 10.59% and 76.82%, respectively as Clin Pharmacol 1998;38:382-9.
shown in Figure 2(a) and 2(b). Statistically: The antagonism 13. Carpio JD, Kabbash L, Turenne Y, et al. Efficacy and safety of loratadine (10
in rate at the concentrations of 10-18 to 10-12 gm/ml was mg once daily), terfenadine (10 mg twice daily) and placebo in the treatment
non-significant, while the concentrations 10-11 to 10-3 of seasonal allergic rhinitis.J Allergy Clin Immunol 1989;84:741-6.
gm/ml showed the significant response (P<0.001). 14. Gutkowski A, Bedard P, Carpio JD, et al. Comparison of the efficacy and safe-
ty of loratadine, terfenadine and placebo in the treatment of seasonal allergic
The antagonism in amplitude at the concentrations rhinitis. J Allergy Clin Immunol 1988;81:902-7.
10-18 to 10-14 gm/ml was non-significant, while at the con- 15. Reicin Alise, White R, Weinstein SF, et al. Montelukast, a leukotriene recep-
centrations 10-13 to 10-3 gm/ml showed significant tor antagonist in combination with loratadine, a histamine receptor antagonist,
response (P<0.001). in the treatment of chronic asthma. Arch Intern Med 2000;160:2481-8.
16. Day JH, Briscoe M, Widlitz MD. Cetirizine, loratadine, or placebo in subjects
Discussion with seasonal allergic rhinits: Effects after controlled ragweed pollen chal-
lenge in an environmental exposure unit. J Allergy Clin Immunol
In the present in vitro study, we have observed the 1998;101:638-45.
effects of first generation anti-histamine (chlorpheniramine 17. Compton MR, Hillery SJ, Kerr SJ, et al. Different immunological sensitizing
maleate) and second generation anti-histamine (loratadine), regimens and airway reponsiveness in vitro to contractile agonists in guinea
on histamine induced contractions of tracheal smooth mus- pigs. Eur J Pharmacol 1988;158:129-34.

cles. In this study histamine produced a prominent action on 18. Bristow M, Sherrod TR, Green RD. Analysis of Beta receptor drug interac-
tions in isolated rabbit atrium, aorta, stomach and trachea. J Pharmacol Exp
the smooth muscles of the tracheal tissues of the rabbit char- Therap 1970;171:52-9.
acterized by contractions and both the anti-histamines 19. Drazen JM, Schneider MW. Comparative responses of tracheal spirals and
showed significant antagonistic effects, but the effects of parenchymal strips to histamine and carbachol in vitro. J Clin Invest 1977;
61:1441-7.
chlorpheniramine maleate were more than the loratadine.
20. Blattner R, Lassen HG, Dehhert H, et al. Experiments on isolated smooth
This study is according to in vivo study done by Druce et muscle preparations. HSE Biological Techniques 1978. Hugosachs Electroni
al12 in allergic rhinits who has compared the symptomatic KKG. Germany.
effects of brompheniramine (belongs to same group as 21. Diamantis CW, Sofia RD. Modulation of in vitro anaphylaxis of guinea-pig
isolated tracheal segments by azelastine, inhibitors of arachidonic acid metab-
chlorpheniramine maleate) and loratadine. He found that olism and selected anti-allergic drugs. Pharac 1986;87:443-8.
brompheniramine was superior than loratadine in clinical 22. Brink C, Duncan PG, Douglas JS. The response and sensitivity to histamine
of respiratory tissues from normal and ovalbumin-sensitized guinea pigs:
efficacy. Still there is no in vitro study available on lorata- Effects of cyclo-oxygenase and lipoxygenase inhibition. J Pharmacol Exp
dine to observe its effects on the respiratory tract. More Therap 1981;217:592-601.
work on this drug is necessary. feature characteristic of embryonal RMS is its lack of gene
References amplification. The cellular DNA content of embryonal
1. Gozsy B, Kato L. Investigations into the mechanism of action of antihista- RMS is hyperdiploid.15-18
mines. Ind J Med Res 1961;49:595-601.
RMS clinically presents with ptosis (droopy eyelid),
2. Corren J, Harris AG, Aaronson D, et al. Efficacy and safety of loratadine plus
pseudoephedrine in patients with seasonal allergic rhinitis and mild asthma. J
and/or unilateral proptosis (eye prominence), as a sub-con-
Allergy Clin Immunol 1997;100:781-8. junctival tumor, dislocation of the lens, or impairment of
3. Roquet A, Dahlen B, Kumlin M, et al. Combined antagonism of leukotrienes ocular mobility. It is usually found in the superonasal orbit
of allergen-induced early and late phase airway obstruction in asthmatics. Am (that is under the upper lid near the nose). Due to its vari-
J Resp Crit Care Med 1997;155:1856-63.
able manifestations it may imitate orbitocellulitis, chalazae,
4. Morrow JD, Roberts LJ, Brow N, et al. Autacoids; drug therapy of inflamma-
tion In: Goodman and Gilman's Pharmacological basis of therapeutics. 10th epibulbar papilloma or as naso lacrimal duct obstruc-
ed. New York: McGraw Hill 2001, pp 211-12. tion.11,19,20 Rapid evolution, tumour burden (tumour size
5. Katzung BG. Histamine serotonin and the ergolalkaloids: basic and clinical >5.cms.) and regional lymph node involvement are indica-
pharmacology 8th ed. New York: McGraw-Hill 2001, pp 272-91.
tors of poor prognosis. Computed axial tomography (CT-
6. Estelle F, Simons R. A double-blind, single-dose, crossover comparison of
cetirizine, terfenadine, loratadine, astemizol and chlorpheniramine versus scan) and magnetic resonance imaging (MRI) typically
placebo: Suppressive effects on histamine-induced wheals and flares during show a mass adjacent to or attached to one of the ocular or
24 hours in normal subject. J Allergy Clin Immunol 1990;540-7. orbital muscles. CT is particularly helpful because it shows
7. Lippincott's, autocoids and Autocoid antagonists. In: Harvey RA, Champe PC orbital bone tumor invasion and characteristic focal calcifi-
(eds.). Lippincott's illustrated reviews 2nd edition. Philadelphia 2000; 419-28.
8. Kay GG, Berman B, Mockoviak SH, et al. Initial and steady-state effects of
cation. Magnetic resonance (MR) imaging allows this tumor
diphenhydramine and loratadine on sedation, cognition, mood and psychomo- to be differentiated from pseudogliomas, such as Coats dis-
tor performance. Arch Intern Med 1997;157:2350-6. ease and retrolental fibroplasia. CT and MR imaging help in
9. Nicholson AN. Therapeutics: antihistamines and sedation. Lancet the differentiation from dermoid, cavernous hemangioma,
1983;2:211-12.
and lymphangioma.21
10. Sheffer AL, Samuels LL. Cetirizine: antiallergic therapy beyond traditional
H1 antihistamines. J Allergy Clin Immunol 1990;86:1040-6. Early diagnosis, complete surgical resection fol-
 Table 1 (b). Effects of histamine on isolated trachea of rabbit in group 1 (rate).

Drug Baseline Histamine Baseline to P value

concentration Mean SEM Mean SEM histamine (%)

10-18 28.50 0.42 29.38 0.46 0.03 NS

10-17 28.50 0.42 29.50 0.42 3.50 NS
10-16 28.75 0.45 29.75 0.53 3.47 NS
10-15 29.25 0.53 30.13 0.52 0.88 NS
10-14 29.50 0.42 30.50 0.42 3.38 NS
10-13 30.25 0.45 31.00 0.42 2.47 NS
10-12 30.50 0.42 31.50 0.42 3.27 NS
10-11 31.00 0.46 32.13 0.40 3.64 NS
10-10 31.38 0.46 32.38 0.46 3.18 NS
10-9 31.88 0.52 32.88 0.52 3.13 NS
10-8 32.25 0.53 33.75 0.59 4.65 NS
10-7 32.88 0.55 36.63 0.46 11.40 0.001
10-6 33.63 0.53 39.13 0.67 16.35 0.001
10-5 34.50 0.60 41.25 0.90 19.56 0.001
10-4 34.75 0.49 42.75 0.77 23.02 0.001
10-3 35.63 0.56 45.25 0.84 26.99 0.001

SEM = Standard error of mean

NS = Non-significant
gm/ml = Gram per milliliter

Figure 1 (a). Antagonistic effects ofchlorpheniramine maleate on amplitude of histamine induced Figure 1 (b). Antagonistic effects of chlorpheniramine maleate on rate of histamine induced con-
contractions in isolated trachea of rabbit (group II). tractions in isolated trachea of rabbit (group II).
difference between the mean values of these two the rate and amplitude 10.59% and 76.82%, respectively as
data showed that loratadine dose dependently antagonized shown in Figure 2(a) and 2(b). Statistically: The antagonism
 in rate at the concentrations of 10-18 to 10-12 gm/ml the smooth muscles of the tracheal tissues of the rabbit char-
was non-significant, while the concentrations 10-11 to 10-3 acterized by contractions and both the anti-histamines
gm/ml showed the significant response (P<0.001). showed significant antagonistic effects, but the effects of
The antagonism in amplitude at the concentrations chlorpheniramine maleate were more than the loratadine.
10-18 to 10-14 gm/ml was non-significant, while at the con- This study is according to in vivo study done by Druce et
centrations 10-13 to 10-3 gm/ml showed significant al12 in allergic rhinits who has compared the symptomatic
response (P<0.001). effects of brompheniramine (belongs to same group as
chlorpheniramine maleate) and loratadine. He found that
brompheniramine was superior than loratadine in clinical
efficacy. Still there is no in vitro study available on lorata-
dine to observe its effects on the respiratory tract. More
work on this drug is necessary.
References
1. Gozsy B, Kato L. Investigations into the mechanism of action of antihista-
mines. Ind J Med Res 1961;49:595-601.
2. Corren J, Harris AG, Aaronson D, et al. Efficacy and safety of loratadine plus
pseudoephedrine in patients with seasonal allergic rhinitis and mild asthma. J
Allergy Clin Immunol 1997;100:781-8.
3. Roquet A, Dahlen B, Kumlin M, et al. Combined antagonism of leukotrienes
of allergen-induced early and late phase airway obstruction in asthmatics. Am
J Resp Crit Care Med 1997;155:1856-63.
4. Morrow JD, Roberts LJ, Brow N, et al. Autacoids; drug therapy of inflamma-
tion In: Goodman and Gilman's Pharmacological basis of therapeutics. 10th
ed. New York: McGraw Hill 2001, pp 211-12.
5. Katzung BG. Histamine serotonin and the ergolalkaloids: basic and clinical
pharmacology 8th ed. New York: McGraw-Hill 2001, pp 272-91.
6. Estelle F, Simons R. A double-blind, single-dose, crossover comparison of
Figure 2 (a). Antagonistic effects of loratadine on amplitude of histamine induced contractions in cetirizine, terfenadine, loratadine, astemizol and chlorpheniramine versus
isolated trachea of rabbit (group III). placebo: Suppressive effects on histamine-induced wheals and flares during
24 hours in normal subject. J Allergy Clin Immunol 1990;540-7.
7. Lippincott's, autocoids and Autocoid antagonists. In: Harvey RA, Champe PC
(eds.). Lippincott's illustrated reviews 2nd edition. Philadelphia 2000; 419-28.
8. Kay GG, Berman B, Mockoviak SH, et al. Initial and steady-state effects of
diphenhydramine and loratadine on sedation, cognition, mood and psychomo-
tor performance. Arch Intern Med 1997;157:2350-6.
9. Nicholson AN. Therapeutics: antihistamines and sedation. Lancet
1983;2:211-12.
10. Sheffer AL, Samuels LL. Cetirizine: antiallergic therapy beyond traditional
H1 antihistamines. J Allergy Clin Immunol 1990;86:1040-6.
11. Therapeutic drugs (Cetirizine). 2nd ed. 1999;1:C152-C156.
12. Druce HM, Thoden WR, Mure P, et al. Brompheniramine, loratadine and
placebo in allergic rhinitis: a placebo-controlled comparative clinical trial. J
Clin Pharmacol 1998;38:382-9.
13. Carpio JD, Kabbash L, Turenne Y, et al. Efficacy and safety of loratadine (10
mg once daily), terfenadine (10 mg twice daily) and placebo in the treatment
of seasonal allergic rhinitis.J Allergy Clin Immunol 1989;84:741-6.
14. Gutkowski A, Bedard P, Carpio JD, et al. Comparison of the efficacy and safe-
ty of loratadine, terfenadine and placebo in the treatment of seasonal allergic
rhinitis. J Allergy Clin Immunol 1988;81:902-7.
15. Reicin Alise, White R, Weinstein SF, et al. Montelukast, a leukotriene recep-
tor antagonist in combination with loratadine, a histamine receptor antagonist,
Figure 2 (b). Antagonistic effects of loratadine on rate of histamine induced contractions in isolat-
in the treatment of chronic asthma. Arch Intern Med 2000;160:2481-8.
ed trachea of rabbit (group III).
16. Day JH, Briscoe M, Widlitz MD. Cetirizine, loratadine, or placebo in subjects
Discussion with seasonal allergic rhinits: Effects after controlled ragweed pollen chal-
In the present in vitro study, we have observed the lenge in an environmental exposure unit. J Allergy Clin Immunol
1998;101:638-45.
effects of first generation anti-histamine (chlorpheniramine
17. Compton MR, Hillery SJ, Kerr SJ, et al. Different immunological sensitizing
maleate) and second generation anti-histamine (loratadine), regimens and airway reponsiveness in vitro to contractile agonists in guinea
on histamine induced contractions of tracheal smooth mus- pigs. Eur J Pharmacol 1988;158:129-34.
cles. In this study histamine produced a prominent action on 18. Bristow M, Sherrod TR, Green RD. Analysis of Beta receptor drug interac-
18. Bristow M, Sherrod TR, Green RD. Analysis of Beta receptor drug interac- KKG. Germany.
tions in isolated rabbit atrium, aorta, stomach and trachea. J Pharmacol Exp 21. Diamantis CW, Sofia RD. Modulation of in vitro anaphylaxis of guinea-pig
Therap 1970;171:52-9. isolated tracheal segments by azelastine, inhibitors of arachidonic acid metab-
19. Drazen JM, Schneider MW. Comparative responses of tracheal spirals and olism and selected anti-allergic drugs. Pharac 1986;87:443-8.
parenchymal strips to histamine and carbachol in vitro. J Clin Invest 1977; 22. Brink C, Duncan PG, Douglas JS. The response and sensitivity to histamine
61:1441-7. of respiratory tissues from normal and ovalbumin-sensitized guinea pigs:
20. Blattner R, Lassen HG, Dehhert H, et al. Experiments on isolated smooth Effects of cyclo-oxygenase and lipoxygenase inhibition. J Pharmacol Exp
muscle preparations. HSE Biological Techniques 1978. Hugosachs Electroni Therap 1981;217:592-601.