Journal of Analytical Sciences, Methods and Instrumentation, 2012, 2, 103-107 Published Online June 2012 ( 103
New Approach for the Determination of Tricyclic
Antidepressant Amitriptyline Using β-Cyclodextrin-PEG
System via Spectrophotomerty
Pankaj Soni1, Santosh K. Sar2, Rajmani Patel1
1Department of Applied Chemistry, Faculty of Engineering & Technology, Shri Shankaracharya Group of Institutions, Junwani,
Bhilai, India; 2Department of Engineering Chemistry, Bhilai Institute of Technology, Bhilai House, Durg, India.
Received December 22nd, 2011; revised January 16th, 2012; accepted January 31st, 2012
A new and simple procedure for the spectrophotometric determination of the tricyclic antidepressant drug amitriptyline
is proposed. The method is based on enhancement o f sensitivity of the [AMIYTP]+ β-cyclodextrin and PEG molecules
involved in formation of molecules inclusion complex, in presence of polyethylene glycol (PEG) medium. The mole
ratio of [AMIYTP]+ β-cyclodextrin and PEG molecules in inclusion complex were determined by the curve fitting
method. The value of molar absorptivity of {[AMIYTP: (β CD)] PEG} complex in term of the drug lies in rag e of (2.20
- 2.23) × 104 L·mole–1·cm–1 at absorption maximum 242 nm. The slope, intercept and correlation co-efficient were
found to be 14.21, 0.0046, and 0.998, respectively. The effect of analytical variables on the determination of the drug
and composition of the ion associated complex are discussed in the paper. This method is applicable in the determina-
tion of amitriptyline in the pharmaceutical preparations.
Keywords: Amitriptyline; PEG; β-Cyclodextrin; Spectrophotometric Determination; AMIYTP-β-Cyclodextrin
Complex; Pharmaceuticals
1. Introduction
Amitriptyline [3-(10,11-dihydro-5H-dibenzol[a,d]cyclo-
hept-5-ylidene) propyldimethylamine] constitute an im-
portant class of neurotherapeutics [1] belonging to the
first generation of antidepressant drug [2,3] The former
has a carbocyclic structure with an exocyclic double
bond at C-5 which is substituted with an N, N-dimethyl-
1-propanamino side chain (Figure 1). The value of angle
of these molecules is important, the more nearly planar
the greater the neuroleptic activity. It is believed that
amitriptyline acts by blocking the receptors of neuro-
transmitters, naradrenaline in the synopsis in the central
nervous system, which results an increase of concentra-
tion of both molecules with a subsequent—enhancement
of their antidepressant patency [3]. However, this drug
suffers from several drawbacks, such as antiarrhytmic,
anticholinergic, cardiovascular and/or hyperthermia side
effect [2,3], which may be reduced if the drugs are suita-
bly vectored to the organism. In this field cyclodextrins
(CDs) are considered as one of the most suitable artificial
receptors for the vectorization of guest hydrophobic
molecules (drug, dyes, detergents, pesticides, etc.) in
aqueous media [4-6]. In fact, the use of CDs as a new
family of pharmaceutical excipient and drug carriers has
become an increasingly accepted method for many
therapeutic molecules [7].
Various analytical methods have been reported for de-
termination of amitriptyline including spectrofluoromet-
ric [8], fluorospectrophotometric [9-14] flow injection
method [15] atomic absorption spectroscopic [16], con-
ductometric [17,18], high performance liquid chroma-
tographic [19,20], voltammetric [21] and chroma-
tographic [22] methods. Some of these methods are not
Figure 1. [AMIYTP] Cl Chemical Name: [3-(10,ll-dihydro-
5H-dib enzol [a ,d] cyclohep t-5 -ylid ene) prop yldime thylam ine] .
Copyright © 2012 SciRes. JASMI
New Approach for the Determination of Tricyclic Antidepressant Amitriptyline Using
β-Cyclodextrin-PEG System via Spectrophotomerty
simple for the routine analysis and they need sophisti-
cated instruments which are not commonly available in
the routine laboratories and conventional batch process
solvent extraction is a tedious and time consuming pro-
cedure. Therefore it seems necessary to develop a sensi-
tive, simple and fast identification and determination of
Amitriptyline. The proposed method is based on the en-
hancement of sensitivity of [AMIYTP: βCD] co mplex in
presence of polythene glycol (PEG) medium. The opti-
mization of analytical variables is discussed in this paper.
The method is simple, sensitive and reproducible. This
method has been applied for determination of amitrip-
tyline in pharmaceutical preparations.
2. Experimental
2.1. Reagents
All chemicals used were of analytical grade reagent
(Merck). The standard solution of amitriptyline (l000
ppm) was prepared by dissolving its 1.000 g in 1 liter
deionized double distilled water. The working solutions
were prepared by the appropriate dilution of the stock
solution. Solutions used were prior filtered. The
β-cyclodextrin 1000 ppm (8.81 × 10–4 M) solution was
prepared by dissolving its 0.1gm in 100 ml deionized
double distilled water, and further diluted to 800 ppm
(7.04 × 10–4 M) with double distilled water. The PEG
solution 1000 ppm (2.5 × 10–3 M) was prepared by dis-
solving its 0.1 gm in 100 ml deionized double distilled
water, and it was further diluted to 20 ppm (5 × 10–5 M)
with deionized distilled water. All working solutions
employed were prior filtered and degassed, by the de-
gassing unit.
2.2. Equipment
Systronics UV-VIS double beam spectrophotometer
–2201 matched with 1 cm quartz cell were employed to
determine the absorbance. The absorbance of the solu-
tions of different concentrations of amitriptyline are
shown in Figure 2.
2.3. Procedure for determination of
2 ml aliquots of the standard solution of amitriptyline
Figure 2. Calibration Graph for Determination of Amitrip-
tyline 0.1 to 1.0 ppm (mg·L–1).
having varying concentrations from 0.1 to 1.0 ppm were
taken in 10 ml volumetric flasks. In each volumetric
flask 2 ml of β-CDs solution and 2 ml of PEG solution
were added. Then made up the solution up to the mark on
volumetric flask with double distilled water. Then meas-
ured their absorbance at 242 nm against the reagent blank,
and prepared the calibration graph by plotting absorbance
versus concentration of amitriptyline, Figure 2. The
similar procedure was repeated with sample solution,
which was prepared from pharmaceutical product. The
concentration of amitriptylin e in the sample solution was
computed by using the calibration curve prepared under
similar condition.
3. Results and Discussion
3.1. Reaction Mechanism and Composition of
Amitriptyline gives ion-associate species with β-CD and
PEG system and the absorbance increases with increase
in the concentration of amitriptyline. Amitriptyline cation
reacts with β-CDs to give an inclusion complex. Their
stoichiommetry are 1:1 indicating that the complex is
formed by the association of a molecule of β-CD per
each molecule of [AMIYTP]+, as usually found for most
cyclodextrin/drug complexes. Considering this 1:1 sto-
ichiommetry the molecular encapsulation process can be
represented as:
 
mn m
where n = 1 to 2.
This reaction has been used for the determination of
cationic antidepressant drug Amitriptyline in presence of
PEG molecule. The solubilization as well as the viscosity
of an aqueous solution of polymer bound system is
higher than the solution of pure polymer. The expected
reaction in the PEG medium can be expressed as:
 
mn m
 
Copyright © 2012 SciRes. JASMI
New Approach for the Determination of Tricyclic Antidepressant Amitriptyline Using
β-Cyclodextrin-PEG System via Spectrophotomerty 105
where, value of “χ” may vary from 3-4 and [AMIYTP]+ =
cation of the drug Amitriptyline.The mole ratio of [AMI-
YTP]+ to β-CD and PEG molecules involved in formula-
tion of molecules inclusion complex, were determined by
the curve fitting method by plotting log(Aeq./Amax. – Aeq.)
vs. Concentration. Where, Aeq. = Absorbance of the
complex when reagent was in equilibrium, and Amax. =
Absorbance when reagent was in constant excess. The
value of slope for β-CD and PEG were found to be in
ratio 0.140 and 0.436, respectively Figures 3 and 4.
Curve-fitting method suggested the molar ratio of
β-CD and PEG and drug cation in the complex to be in
1:3 ratios. As per the curve-fitting method, the composi-
tion of the inclusion complex is expected to be as:
 
3.2. Absorption Spectra
The {[AMIYTP](βCD)(PEG)3}+ Cl complex exhibit the
absorption maximum at 242 nm. The position of
max is
changed when the PEG is added and absorbance was
increased. The absorption maxima of only {[AMIYTP]
(βCD)} complex were found to be at 250 nm. The posi-
tion of λmax is change when the PEG is added and the
absorbance was increased. The {[AMIYTP](βCD)
(PEG)3}+Cl complex exhibits the absorbance maximum
at 242 nm Figure 5. In presence of the drug/PEG, hy-
perchromic and hypsochromic shift was observed due to
formation of molecular inclusion complex.
3.3. Optimum Concentration Range, Detection
Limit and Statistics
The absorbances for the different concentration of amitrip-
tyline are sho wn in Figure 2. The method followed line-
arity in the range 0.1 - 1.0 ppm of amitriptyline with the
slope, intercept and correlation coefficient of 14.21,
0.00458, and 0.998 respectively. The value of molar ab-
sorptivity [calculated by taking the molar concentration
of amitriptyline and path length of the cell to be 1 cm]
with amitriptyline drug was found in the range (2.20 -
2.23) × 10–4 L·mole–1·cm–1 at the absorption maximum
242 nm. The detection limit (absorbance > 3 × SD) of the
method was 0.034 ppm. The relative standard deviation
for the analysis of seven different solutions containing
0.5 ppm amitriptyline was found to be ±0.023%.
3.4. Application of the Method
The proposed method was applied for the determination
of amitriptyline in commercial pharmaceuticals tablets
Amil-25 (From Mano Pharma India) and Amitriptyline
Hydrochloride tablet USP/IP—25 mg (From (Iron) drugs
and pharmaceuticals Pvt. Ltd. India). Sample solutions
were prepared from the tablets. Five tablets of each pro-
duce were weighed and powdered. The powder equiva-
lent to their single tablet was filtered and evaporated to
dryness. Then the residue was dissolved in distilled water
and made up the volume up to 100 ml in a volumetric
flask. These stock solutions were further diluted to con-
tain the requisite concentration. Then followed the pro-
cedure as described earlier for the proposed method, and
it was compared with the results obtained from the offi-
cial method, Tables 1 and 2.
Figure 3. Curve fitting method for determination of molar
ratio of β-cyclodextrin in ion-associate complex.
Figure 4. Curve fitting method for determination of mole
atio of the PEG in the ion-associate complex. r
Copyright © 2012 SciRes. JASMI
New Approach for the Determination of Tricyclic Antidepressant Amitriptyline Using
β-Cyclodextrin-PEG System via Spectrophotomerty
Figure 5. Absorption spectra of {[AMIYTP](β-CD)}+Cl and {[(AMIYTP](β-CD)] (PEG)3}+Cl Ions—associate complexes.
Table 1. The result of analysis of Amitriptyline tablets by proposed and official method.
Recovery, mg
S.No. Pharmace u t i c al Product Official Method Proposed Method Error, %
1. Amit 25, 25 mg 25.3 ± 0.03 24.87 ± 0.16 1.6%
2. Amitriptyline Hydrochloride tablet 25mg 25.2 ± 0.25 24.72 ± 0.32 1.9%
Table 2. Comparison with other established spec trophotometric method.
S.No. Method λmax, (nm)Working Range, (ppm)R.S.D* (%) Correlation Co-ef f i cie nt
1. Extraction sp ectrophotometer ammonium reinkate method 523 0.1 - 6.0 0.8 0.994
2. Proposed spectrophotometric {AMIYTP(βCD)(PEG)3}+Cl
ion associate complex method 242 0.1 - l.0 2.0 0.998
4. Conclusion
The method was successfully applied for the determina-
tion of amitriptyline in the pharmaceutical preparations.
The method is very simple as there is no requirement of
prior separation or extraction of the complex and the re-
agents are cheap and commonly available in routine
laboratories. The results obtained from the proposed
method were comparable with the established methods.
The method has good potential in simplicity and sensitiv-
5. Acknowledgements
Authors are thankful to the Head, Department of Applied
Chemistry and the Director, Shri Shankaracharya College
of Engineering & Technology, Bhilai, for providing
laboratory facilities for this work.
[1] G. A. Gilman, W. T. Rall, S. A. Nies and P. Taylor, “The
Pharmacological Basis of Therapeutics,” Ninth Edition,
Mc Graw Hill, New York, 1996.
[2] C. M. Hall and R. A. Nugent, “Encyclopedia of Chemical
Technology,” John Wiley & Sons, New York, 1992.
[3] W. C. Bowman and M. L. Rand, “Textbook of Pharma-
cology,” Blackwell Science Publication, London, 1992.
[4] J. L. Atwood, J. E. D. Daviess, D. D. Mac Nicol and F.
Vogtle, “Comprehensive Supramolecular Chemistry Per-
gamon,” Oxford, 1996.
[5] K. A. Connors, “The Stability of Cyclodextrine Complex
in Solution,” Chemical Reviews, Vol. 97, No. 5, 1997, pp.
1325-1357. doi:10.1021/cr960371r
[6] V. T. D’Souza and K. B. Lipkowitz, “Cyclodextrines,”
Chemical Reviews, Vol. 98, 1998, p. 1741.
[7] D. O. Thompson, “Cyclodextrine—Enabling Excipients.
Copyright © 2012 SciRes. JASMI
New Approach for the Determination of Tricyclic Antidepressant Amitriptyline Using
β-Cyclodextrin-PEG System via Spectrophotomerty 107
Their Present and Future Use in Pharmaceuticals,” Criti-
cal Reviews in Therapeutic Drug Carrier, Vol. 14, No. 1,
1997, pp. 1-104.
[8] F. A. Mohamed, H. A. Mohamed, S. A. Hussein and S. A.
Ahmed, “A Validated Spectrofluorimeter Method for De-
termination of Some Psychoactive Drug,” Journal of
Pharmaceutical and Biomedical Analysis, Vol. 39, No.
1-2, 2005, pp. 139-146. doi:10.1016/j.jpba.2005.03.024
[9] J. E. F. Reynolds (Ed.), “The Extra Pharmacopoeia,” 30th
Edition, The Pharmaceutical Press, London, 1993, p.
[10] J. Karpinska, J. Suszymska and J. Trace, “The Spectro-
phometric Simultaneous Determination of Amitriptyline
and Chloropromazine Hydrochloride in Their Binary
Mixture,” Microprobe Technology, Vol. 19, 2001, pp.
355- 364. doi:10.1081/TMA-100105051
[11] A. F. Faten, N. El-Die n, G. Mohamed Gehad and A. Mo-
hamed, “Nehad Spectrophoto Metric Determination of
Aminopline, Trazodone, Amitriptyline through Using
Methyl Orange and Bromocresol Green Reagent,” Spec-
trochemi Acta Part A, Vol. 65, 2006, pp. 26-30.
[12] A. Tehssen, A. A Kazi, I. H. Mohamed and A. Tal,
“Spectrophotometric Determination of Amitriptyline Hy-
drochloride in Pure and Pharmaceutical Preparation,”
Analytical letters, Vol. 33, No. 12, 2000, pp. 2477-2490.
[13] J. O. Onah, “Spectrophotometric Determination of
Amitriptyline by Method of Charge Transfer Complexa-
tion with Chloranic,” Acid Global Journal of Puro and
Applied Science, Vol. 11, No. 2, 2005.
[14] W. Micink and M. Tarasiewicz, “Application of Thiocy-
nate Complex of Titanium(IV) to the Extractive Spectro-
photometric Determination of Amitriptyline Hydrochlo-
ride,” Microchemical Journal, Vol. 78, No. 107, 1993, pp.
[15] R. M. El-Nashar, N. T. Ghani Abdel and A. A. Bioumy,
“Flow Injection Potentiometric Determination of Amitrip-
tyline Hydrochloride,” Microchemical Journal, Vol. 78,
No. 2, 2004, pp. 107-113.
[16] E. M. Elnemma, F. M. ElZawawy and S. M. Hassan,
“Saad Determination of Amitriptyline Impiramine and
Orphenadrine in Antidepressant Drug by Potentiometry,
Spectrometry, and Atomic Absorption Spectrophotome-
try,” Microchimica Acta, Vol. 110, No. 13, 1993, pp. 79-
88. doi:10.1007/BF01243988
[17] T. Ghani Nour, A. El. Nashar, R. M. Bioumy and A.
Abeer, “Conductimetric Determination of the Antide-
pressant Amitriptyline and Dothiepin Hydrochloride and
Tranylcypromine Hemisulphate in their pharmaceutical
formulation,” FABAD Journal of Pharmaceutical Sci-
ences, Vol. 29, 2004, pp. 195-201.
[18] C. Jessica, R. Alberto, A. Emillio and J. Elena, “Tem-
perature Effect on the Complex Formation between Tri-
cyclic Antidepressant Drug (Amitriptyline or Imipramine
and Hydroxypropyl-β-cyclo-dextrine in Water,” Journal
of Inclusion Phenomena and Macrocyclic Chemistry, Vol.
59, No. 3-4, 2007, pp. 279-285.
[19] G. V. Deshmane, J. R. Vakil, S. R. Dhaheshwar, K. R.
Mahadik and S. S. Kadam, “Simultaneous Spectropho-
tometric Estimation of Amitriptyline Hydrochloride and
Chlordiazepoxide,” Indian Drug, Vol. 34, 1997, pp. 443-
[20] R. Gandhi, P. Sharma, H. P. S. Chaw la, C. L. Kaul and R.
Panchagnula, “HPLC Analysis of Tricyclic Antidepres-
sant in Skin Diffusion Studies,” S.S.P. Pharmaceutical
Science, Vol. 10, 2000, pp. 234-238.
[21] W. Lu, S. A. Shamsi, T. D. McCarley and I. M. Warner,
“Online Capillary Electrophorosis Electrospray Ionization
Mass Spectroscopy Using Polymerized Anionic Surfac-
tant,” Electrophoresis, Vol. 19, No. 12, 1998, pp. 2193-
2199. doi:10.1002/elps.1150191225
[22] L. Simonson and K. Nelson, “Adsorption of Amitrip-
tyline Hydrochloride in HPLC Injector Loops,” LC-GC,
Vol. 10, No. 7, 1992, pp. 533-535.
[23] E. Janquera, S. Romero and C. Aicart, “Behavavior of
Tricyclic Antidepressanls in Aqueous Solution: Seif Ag-
gregation and Association with β-Cyclodextrin,” Lang-
muir, Vol. 17, No. 6, 2001, pp. 1826-1832.
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