1.

0 INTRODUCTION

1.1 Inflammation
Inflammation is everyday and important defensive response to the dangerous stimuli along
with infectious marker. It is as a result of a ramification of stimuli, such as bodily harm, UV
irradiation, microbial assault, and immune reactions [1]. Inflammation is the primary
response to infection or injury that functions to clear the injurious material or agent and
promote tissue repair. It is characterized by the sequential release of mediators including;
bioactive amines, eicosanoids, cytokines, chemokines and growth factors that regulate
increased vascular permeability and recruitment of blood borne leukocytes. Increased
vascular permeability also results in extravasation of plasma proteins that further amplify
the inflammatory reaction. Inflammation is essentially a salutary response that normally
resolves with the restoration of normal tissue structure and function, however when
inflammation persists it can cause tissue damage and loss of function [2].

Inflammation is an important physiological reaction which occurs in response to a

wide variety of injurious agents (eg. bacterial infection, physical trauma, chemicals or any
other phenomenon) ultimately aiming to perform the dual function of limiting damage and
promoting tissue repair [3]. Inflammatory processes are required for immune surveillance,
optimal repair, and regeneration after injury [4]. The inflammatory process protects our
body from diseases by releasing cells and mediators that combat foreign substances and
prevent infection [5]. However, sustained, excessive or inappropriate inflammation is the
cause of numerous diseases including rheumatoid arthritis, psoriasis and inflammatory
bowel disease [6]. Inflammation is a major component of the damage caused by
autoimmune diseases, and is a fundamental contributor of various infectious and non-
infectious diseases such as cancer, diabetes, cardiovascular disease, rheumatoid arthritis,
Alzheimer’s and arteriosclerosis. Depending on the intensity of this process, mediators
generated in the inflammatory site can reach the circulation and cause fever [7-9].

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1.2 Inflammatory Process

1.2.1 Acute inflammatory

Acute inflammatory response is characterized by an increase in vascular permeability and
cellular infiltration leading to oedema formation as a result of extravasation of fluid and
proteins and accumulation of leukocytes at the inflammatory site for short time [10].

1.2.2 Chronic inflammation

Chronic inflammation is the reaction arising when the acute response is insufficient to
eliminate the pro-inflammatory agents [11].

1.2.3Standard drugs for inflammation[12-16]

 Non-steroidal anti-inflammatory drugs (NSAIDs)

 Diclofenac
 Indomethacin
 Aspirin
 Ibuprofen
 Coxib:Rofecoxib, Lumiracoxib, Celecoxib and Etoricoxib
European Medicines Agency (EMEA) as well as by a large number of national drug agencies
all over the world. Thus, a careful evaluation of the risk profiles for adverse events before
prescribing non-selective NSAIDs and coxibs is strongly recommended [17-19].

1.3 Muscle spasm

1.3.1 Clinical presentation of muscle spasm

Muscle spasm presents with a number of features. The patient complains oftightness and
stiffness and the location of this is in a non-dermatomal pattern. Generally heat has a
relaxant effect on musclesin spasm, although the duration of relief is very variable. In some
situationswhat is apparent is a mild increase in muscle tone while at the other end ofthe
spectrum there can be complete spasm of the muscle so that it is tightlycontracted.
Complete spasm of a skeletal muscle can be intermittent andunexpected with sudden onset
of incapacitating spasticity.Along with increases in muscle tone there will often be enthetic
pain,that is pain arising from where the muscle joins onto bone[20, 21].

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1.3.2 Muscle relaxants active on central andperipheral nervous system

a. Baclofen

Baclofen is the p-chlorophenyl derivative of gamma amino butyric acid(GABA). It achieves

its effect by being a GABA B agonist and includesamong its side effects sedation and
confusion.

b. Benzodiazepines

Benzodiazepines bind to benzodiazepine receptors located in the terminals ofprimary

fibers leading to increased chloride flux across the terminal membrane withresultant
increase in membrane potential [22].

c. Carisoprodol

d. Cyclobenzaprine

1.3.3 Local anesthetic nerve blocks

Some nerves have a sensory function, others a motor function, and somesubserve both.
Where a nerve supplies a motor function alone, blockadeof that nerve can cause muscle
relaxation in the absence of skin anesthesia.Despite the fact that the deposition of a local
anesthetic beside a nervecauses only a short-term blockade of that nerve, prolonged relief
of musclespasm may occur.

a. Magnesium

Oral administration of magnesium is ineffective because of the poor absorption of the

substance from the gut. Therefore it is usually given by theparenteral route.

b. Methocarbamol

Methocarbamol is a carbamate derivative of guaifenesin and is structurally relatedto

mephenerin. It is available in both oral and parenteralformulations.

c. Orphenadrine

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Orphenadrine is a monomethylated derivative of diphenhydramine.Centrally acting
antihistamines such as this exhibits muscle relaxant andanalgesic properties.

d. Tizanadine

Tizanadine is an imidazoline derivative that is structurally related to clonidine. Like

clonidine, it is an `2-adrenoreceptor agonist.

1.4 UV- Visible spectroscopy:

The technique of UV- Visible spectrophotometry is one of the most frequently employed in
pharmaceutical analysis. It involves the measurement of the amount of ultraviolet (190-
380nm) or visible (380-800nm) radiation absorbed by a substance in solution. Instruments
which measure the ratio, or a function of the ratio, of the intensity of two beams of light in
UV-Visible region, are called UV-Visible spectrophotometer [23].

The amount of absorption depends on the wavelengths of radiation and the

structure of the compound. In case of organic molecules, the electronic transitions could be
ascribed to a σ, π or η electrons transition from the ground state to an excited state (*, π*
or η* ).There are four types of absorption bands that occur due to the electronic transition
of a molecules:

 R-Bands : Compound with C = O or NO2 groups, λmax< 100.

 K–Bands : Compound in conjugated systems, λmax> 10,000.
 B–Bands : due to aromatic and hetro aromatic (Benzenoid bands) systems, λ max<
2000.
 E–Bands : Compound in aromatic systems, (Ethylene bands) λmax 2000 to 14,000.

1.4.1 Chromophores and Auxochromes :

A chromophore is a covalently unsaturated group responsible for electronic absorption, eg.

C = C, C = O and NO2. An auxochrome represents a saturated group, which when attached to
a chromophore changes both the intensity as well as the wavelength of the absorption
maximum, e.g. OH, NH2, Cl.

1.4.1.1 Transmittance and Absorbance :

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There is no direct method for measuring absorbed light. This information is obtained
indirectly by measuring the light that is not absorbed by the sample, i.e. the transmitted
light. The Beer Lambert law relates the absorption intensity to the incident and transmitted
intensities as well as the concentration of the absorbing material, such that :

Io 1
A =log =ε Cl=log −
It T ----------------- (1)

Where, A = Absorbance.

T = Transmittance.

 = Molar absorption coefficient (molar absorptivity).

C = Concentration in mol L-1.

l = Pathlength in cm.

Io= Intensity of incident beam of light.

It = Intensity of transmitted beam of light.

The molar absorption coefficient  depends upon the wavelength of the incident
light, the temperature and the solvent employed.

The wavelength range of UV radiation starts at the blue end of the visible light and ends at
2000A. The ultraviolet regions is subdivided in to two spectral regions- (i) The region
between 2000 A-4000 A is known as near as UV region. (ii) The region below 2000 A is
called far or vaccum UV region. Wavelengths in the ultraviolet region are usually expressed
in nanometers (1nm=10-7 cm) orangstroms (A) (1A=10-8 cm). Occasionally, absorption is
reported in wave numbers (v= cm-1) [24].

1.4.2 PRINCIPLE

Any molecule has either (n) or combination of these electrons. These bonding and
nonbonding (n) electrons absorb the characteristics radiation and undergo transition from
ground state to exited state. By the characteristic absorption peaks the nature of electrons
present and hence the molecular structure can be elucidated. Ultraviolet absorption
spectra arise from transition of electron or electrons with in a

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molecule or an ion from a lower to higher electronic energy level and the ultraviolet
emission spectra arise from the reverse type of transition. When a molecule absorbs
ultraviolet radiation of frequency v sec-1, the electron in the molecule undergoes transition
from a lower to a higher energy level or molecular orbital, energy difference is given by-
E=hv erg.

The actual amount of energy required depends on the difference in energy between
the ground state E0 and excited state E1 of the electrons. E1-E0=hv. We known that total
energy of a molecule is equal to the sum of electronic vibrational and rotational energy. The
magnitude of these energies decrease in following order:- E etec, E vib and E rot.

The most important characteristic of spectrophotometery are there wide

applicability, high sensitivity, moderate to high sensitivity, good accuracy and convenience.
The assay of an absorbing substance may be quickly carried out by preparing a solvent in a
transparent solvents and measuring this absorbance at a suitable wavelength.

This is governed by Beer-Lambert's law which states A = a.b.c

Where, A = is the absorbance.

a = is the absorbtivity.

b = is the path length.

c = is the concentration.

1.4.3 INSTRUMENTATION

a. Radiation Source

The best source of light is the one which is more stable, more intense and which gives
range of spectrum from 180-360 nm (up to 400 nm). The different sources available are:-

 Tungsten lamp.

 Hydrogen discharge lamp.

 Deuterium lamp.

 Xenon discharge lamp

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 Mercury arc.

 Monochromators

The monochromator is used to disperse the radiation according to the wavelength. The
essential elements of a monochromator are an entrance slip, a dispersing element and an
exit slit. The entrance slit sharply defines the incoming beam of heterochromatic radiation.
Quartz and fused silica prisms which are transparent throughout the entire UV range are
widely used in UV spectrophotometers [25].

b. Sample cells:The cells that are to contain samples for analysis should fulfill three main
conditions:

(i) They must be uniform in construction; the thickness must be constant.

(ii) the material of construction should be inert to solvents.

(iii) They must transmit light of the wavelength used.

The most commonly used cells are made of quartz or fused silica. The path Length of the
cell are 10nm to 1 cm.

c. Solvents

Solvent plays an important role in UV spectra, since compound peak could be obscured by
solvent peak. Hence the solvent for a sample is selected in such a way that the solvent
neither absorbs in the region of measurement nor affects the absorption of the sample.

d. Detectors

Although any one of the detectors used in calorimetric can be used, photomultiplier tubes
are mainly used, since the cost of such UV spectrophotometers are high and more accurate
measurements are to be made.

Example:-

(1) Barrier layer cell photo voltaic cell.

(2) Photo tubes or Photo emissive cell.

(3) Photomultiplier tubes (PMT)

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e. Sample cells

The signal from the Photomultiplier tubes is finally received by the recording system. The
recording is done by recorder pen. The type of arrangement is only done in recording UV
spectrophotometers.

The power supply serves a triple function.

(i) It decreases the line voltage is to the instruments operating level with a transformer.(ii)
It converts A.C. to D.C. with a rectifier it direct current is required by the instrument.

(iii) It smoothes out any ripple which may occur in the line voltage in order to deliver a
constant voltage to the source lamp and instruments.

f. Application

 Detection of functional groups.

 Determination of dissociation constant of acids and bases.

 Chemical kinetic.

 Identification of an unknown compounds.

 Extent of conjugation.

 Detection in conjugated and non-conjugated compounds.

 Detection of polynuclear hydrocarbon.

1.4.4 Spectrophotometric methods for estimation of drugsin combined dosage forms

The spectrophotometric assay of drugs rarely involves the measurement of absorbance of

samples containing only one absorbing component. The pharmaceutical analyst frequently
encounters the situation where the concentration of one or more substances is required in
samples known to contain other absorbing substances which potentially interfere in the
assay. If the recipe of the sample formulation is available to the analyst, the identity and
concentration of the interferents are known and the extent of interference in the assay may
be determined. Alternatively, interference which is difficult to quantify may arise in the
analysis of formulations from manufacturing impurities, decomposition products and

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formulation excipients. Unwanted absorption from these sources is termed irrelevant
absorption and, if not removed, imparts a systematic error to the assay of the drug in the
sample.

The basis of all spectrophotometric methods for multicomponent sample analysis is

the property that at all wavelengths :

(a) The absorbance of a solution is the sum of absorbances of individual components; or

(b) The measured absorbance is the difference between total absorbance of the solution in
the sample cell and that of the solution in the reference (blank) cell [26].

1.4.5 Simultaneous Equation Method or Vierodt’sMethod :

If a sample contains two absorbing drugs (X and Y) each of which absorbs at the max of the
other, it may be possible to determine both drugs by the technique of simultaneous
equations (Vierodt’s method) provided that certain criteria apply.

The information required is :

a. The absorptivities of X at 1 and 2, a X1 and a X2 respectively.

b. The absorptivities of Y at 1 and 2, a Y1 and a Y2 respectively.

c. The absorbances of the diluted sample at 1 and 2, A1 and A2 respectively.

Let CX and CY be the concentrations of X and Y respectively in the diluted sample.

Two equations are constructed based upon the fact that at 1 and2 the absorbance
of the mixture is the sum of the individual absorbances of X and Y.

At 1 A1 = a X1 bCX + a Y1 bCY ……………(1)

At 2 A2 = a X2 bCX + a Y2 bCY ……………(2)

For measurements in 1 cm cells, b = 1

Rearrange eq. (2).

A 2 −a X C X
2
CY =
aY
2

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 Substituting for CY in eq (1) and rearranging

A 2 a Y1 − A 1 a Y2
CX=
aX2 a Y1−aX1 a Y2 ……………(3)

A 1 a X2 − A 2 a X1
CY =
and a X2 a Y1−aX1 a Y2 ……………(4)

As an exercise you should derive modified equations containing a symbol (b) for
pathlength, for application in situations where A 1 and A2 are measured in cells other than 1
cm pathlength.

Criteria for obtaining maximum precision, based upon absorbance ratios, have been
suggested (Glenn, 1960) that place limits on the relative concentrations of the components
of the mixture. The criteria are that the ratios

A 2 /A1 a Y2 /a Y1
and
a X /a X A 2 /A1
2 1

Should lie outside the range 0.1-2.0 for the precise determination of Y and X respectively.
These criteria are satisfied only when the max of the two components are reasornably dissimilar.
An additional criterion is that the two components do not interact chemically, thereby negating
the initial assumption that the total absorbance is the sum of individual absorbances. The
additivity of the absorbances should always be confirmed in the development of a new
application of this technique.

1.4.6 Advantages of UV-Visible Spectrophotometry:

Analytical absorption spectroscopy in ultraviolet and visible region of electromagnetic

spectrum is widely used for quantitative analysis in the field of pharmaceutical and
biomedical analysis. Although, there is steady increase in the use of other advance physical
method of analysis such as GLC, HPLC, HPTLC, and AAS is active, highest specificity,
precision and accuracy and one may be tempted to suggest the use of simple colorimetric
methods based on color reaction has diminished in importance and application. However,
it is relevant to note that even the latest edition of various compendia ( IP/BP/USP/EP )
contain identification tests for different drug substances based on the color reactions of

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one or more specific functional groups present in drug molecule. It is an accepted fact that
real sample analysis is often too complex for direct analysis, no matter what new
technology/instrumentation are available. Selective chemical reactions provide the method
to manipulate a sample to reveal the information desired. They also provide the means to
increase the response of an analyte to a particular functional group or increase the
selectivity of analysis by targeting certain components of the sample to response to the
selected reagents. As a matter of fact, most of the time, analytical chemist attempt to
optimize compendial identification tests based on color reactions for quantitative analysis.
Further speed, simplicity and easy maintenance of such instruments (Colorimeter, UV-
Visible spectrophotometer) are added advantages. So UV-Visible spectrophotometric
methods are widely used for quantitative determination in dosage form [27].

1.5 Validation of Analytical Procedures

This document presents a discussion of the characteristics for consideration during the
validation of the analytical procedures included as part of registration applications
submitted within the EC , Japan and USA. This document does not necessarily seek to cover
the testing that may be required for registration in , or export to , other areas of the world.
Furthermore, this text presentation serves as a collection of terms, and their definitions
and is not intended to provide direction on how to accomplish validation [28].
1.5.1 Types of Analytical Procedures:
Types of analytical procedures to be validated . The discussion of the validation of
analytical procedures is directed to the four most common types of analytical procedures.
 Identification tests
 Quantitative tests for impurities
 Limit tests for the control impurities
 Quantitative tests: of the active moiety in samples of drug substance or drug product
or other selected components (s) in the drug product.
Typical validation Characteristics which should be considered are listed below:

1. Accuracy
2. Precision

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 3. Repeatability
4. Intermediate Precision
5. Specificity
6. Detection Limit
7. Quantification Limit
8. Linearity
9. Range
1. Accuracy: The accuracy of an analytical procedure expresses the closeness of agreement
between the value which is accepted either as a conventional ture value or an accepted
reference value found . This is sometimes terms trueness.
2. Precision: The precision of an analytical procedure expresses the closeness of
agreement ( degree of scatter) between a series of measurements obtained from multiple
sampling of the same homogeneous sample under the prescribed conditions. Precision may
be considered at three levels ; repeatability, intermediate precision and reproducibility.
Precision should be investigated using homogeneous, authentic samples. However , If it is
not possible to obtain a homogeneous sample it may be investigated using artificially
prepared samples or a sample solution [29].
3. Repeatability: Repeatability expresses the precision under the same operating
conditions over a short interval of time , Repeatability is also termed intra-assay precision.
4. Intermediate precision: Intermediate precision expresses within -laboratories
variations: different days, different analysts, different equipment etc.
5. Specificity: Specificity is the ability to assess unequivocally the analyze in the presence
of components which may be expected to be present .Typically these might include
impurities, degarants matrix etc. Lack of specificity of an individual analytical procedure
may be compensated by other supporting analytical procedure(s). This definition has the
following implications to ensure that all the analytical procedures performed allow an
accurate statement of the content of impurities of an analyze Ice. related substances test,
heavy metals, residual solvents content, etc [30].
6. Detection Limit: The Quantization limit of an individual analytical procedure is the lows
amount of analyze in a sample which can be quantitatively determined with suitable
precision and accuracy. The Quantization limit is a parameter of quantitative assays for low

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levels of compounds in samplearticularly for the determination of impurities and/or
degradation products.
7. Quantization Limit: The Quantization limit of an individual analytical procedure is the
lowest amount of analyze in a sample which can be quantitatively determined with suitable
precision and accuracy. The Quantization limit is a parameter of quantitative assays for low
levels of compounds in sample matrices, and is use particularly for the determination of
impurities and/or degradation products.
8. Linearity: The linearity of an analytical procedure is its ability ( Within a give range ) to
obtain test results which are proportional to the concentration ( amount) of analyte in the
sample.
9. Range: The range of an analytical procedure is the interval between the upper and lower
concentration (amounts) of analyze in the sample (including these concentrations) for
which it has been demonstrated that the analytical procedure has a suitable level of
precision, accuracy and literary [31].

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2.0 REVIEW OF LITERATURE

1. Hoang et al., (2020) 32 reported RP-HPLC and UV spectrophotometric analysis of

Paracetamol, Ibuprofen, and Caffeine in solid pharmaceutical dosage forms.
2. Prabha et al., (2018) 33Developed the degradation studies of Methocarbamol from
Methocarbamol injection.
3. Alkhafaji et al., (2018) 34 reported the UV-spectrophotometric methods for simultaneous
determination of Paracetamol, Ibuprofen and Caffeine in bulk and pharmaceutical
formulations
4. Ramalakshmi et al., (2017)35Developed the RP-HPLC method for the simultaneous
estimation of methocarbamol and ibuprofen
5. Patel et al (2016)36Developed and validated RP-HPLC method for simultaneous
determination of Methocarbamol and Diclofenac sodium in injection dosage form.
6. Muftri et al., (2015) 37 reported the zero-crossing derivative spectrophotometric method
for determination of ternary mixture of Paracetamol, Ibuprofen and Caffeine in tablet
dosage form.
7. Priyanka M (2014)38Developed and validated stability indicating HPLC assay method
for Methocarbamol in bulk formulation.
8. Sharaf el Din et al., (2014) 39Developed the derivative and ratio derivatives
spectrophotometry method for simultaneous determination of Methocarbamol and
Aspirin binary mixture in their combined tablets
9. Abdelaleem et al., (2013)40 reported the Validated stability indicating RP-HPLC method
for determination of Paracetamol, Methocarbamol and their related substances
methods.
10. Shah et al (2012)41 developed and validated simultaneous equation
spectrophotometric method for simultaneous estimation of Tolperisonehydrochloride
andDiclofenac sodium in their combined tablet dosage form.
11. Patel et al (2012)42 Development and validation of RP-HPLC method for simultaneous
determination of Omeprazole and Diclofenac sodium in capsule dosage form.
12. Ali et al., (2012) 43Developed the ratio spectra method for the simultaneous
determination of Methocarbamol and Ibuprofen.

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13. Nouruddin et al (2012) 44Developed and validated simultaneous determination of
Methocarbamol and Ibuprofen or Diclofenac potassium using mean centering of the
ratio spectra method.
14. Walash et al., (2011)45 reported the Spectrofluorimetric determination of
Methocarbamol in pharmaceutical preparations and human plasma.
15. Elkady et al., (2011)46Developed the simultaneous spectrophotometric method for the
determination of Diclofenac potassium and Methocarbamol in binary mixture using
chemometric techniques.
16. Issa et al., (2011)47 reported the Simultaneous determination of Ibuprofen and
Paracetamol using derivatives of the ratio spectra method.
17. Elkady et al., (2011) 48 reported the simultaneous spectrophotometric determination of
Diclofenac potassium and Methocarbamol in binary mixture using chemometric
techniques.
18. Manmode et al., (2011) 49 reported the Stability indicating HPLC method for
simultaneous determination of Methocarbamol and Nimesulidefrom tablet formulation.
19. Mavar et al., (2011)50 developed the Stability indicating HPLC method for the
determination of Methocarbamol and Nimesulidefrom tablet matrix
20. Elkady et al., (2010)51 reported the Simultaneous determination of Diclofenac
potassium and Methocarbamol in ternary mixture with guaifenesin by RP-HPLC.
21. Rosasco et al., (2009)52 reported the stability-indicating HPLC method for the
determination of Methocarbamol in veterinary preparations
22. Thomas et al., (2009)53 reported Simultaneous determination of Tramadol and
Ibuprofen in pharmaceutical preparation by 1st order derivative method and LC
method.
23. Subramanian et al., (2005)54 developed the RP-LC method for simultaneous
determination of Paracetamol, Methocarbamol and Diclofenac potassium in tablets
dosage form.
24. Satheeshmanikandan et al., (2004)55 reported the Simultaneous spectrophotometric
estimation of Ibuprofen and Methocarbamol in tablet dosage form.

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25. Erk et al., (2001)56 reported the Simultaneous determination of Paracetamol and
Methocarbamol in tablets by ratio spectra derivative spectrophotometry and LC
method.
26. Vasudevan et al., (2000) 57Developed the RP-HPLC method for the Simultaneous
estimation of Paracetamol, Methocarbamol and Ibuprofen.
27. Zarapkar et al., (1999) 58Developed the RP-HPLC method for the determination of
Ibuprofen, Paracetamol and Methocatbamolin tablets dosages form.
28. Koupai-Abyazani et al., (1994) 59 reported the HPLC method for the determination of
Methocarbamol in equine serum and urine.
29. Weng N et al (1994) 60Developed and validated a HPLC method for the determination of
Methocarbamol in human plasma.
30. Erram et al., (1993)61 reported the RP-HPLC method for Simultaneous determination of
Methocarbamol and Paracetamol from single and combined tablets.
31. Alessi-Severini et al., (1992) 62 reported the High performance liquid chromatographic
analysis of Methocarbamol enantiomers in biological fluids.
32. Kir et al., (1991) 63 reported the 2nd derivative spectrophotometry for the determination
of Paracetamol and Methocarbamol in a pharmaceutical preparation.
33. Atay et al., (1990) 64 reported the Quantitative analysis of Methocarbamol and
Paracetamol containing tablets by spectrophotometric methods.
34. Rao, et al., (1990) 65 reported the Gas liquid chromatographic determination of
Paracetamol and Methocarbamol in single and combined dosage forms.

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2.1RESEARCH ENVASIAGED

In pharmaceutical industry, the quality of the manufactured drug in various dosages forms
must be carefully controlled. Slight changes in the composition or in the purity of the drug
itself can affect the therapeutic value. Therefore, to detect these, there is a greater need for
development of newer and better methods of pharmaceutical analysis. Many methods are
available for the quantitative analysis of pharmaceutical dosage forms like HPLC, HPTLC,
UV, Colorimetric method.
Among numerous estimation methods spectrophotometric is a simple, relatively
sensitive and cost-effective method. It can generate meaningful data for interpretation of
results. UV-Visible spectrophotometer can be made into more accurate and freer from
interference from other components by employing advance version of it i.e. derivative
spectroscopy, area calculation, simultaneous equation methods and multi component
analysis.
Methocarbamol (MET) is chemically known as 2-hydroxy- 3-(2-methoxyphenoxy)
propyl carbamate [66]. It is a centrally acting skeletal muscle relaxant whose action may be
due to its general depressant effect on the CNS. Ibuprofen (IBU) is 2-(4-isobutylphenyl)
propionic acid which is a non-steroidal drug with anti-inflammatory, antipyretic and
analgesic properties. They are co-formulated together for treatment of pain associated with
muscle spasm. After exhaustive literature review, there are many reports have been
published for determination of MET and IBU mixture either in their binary mixture or in
mixtures with other drugs.
The combination of Ibuprofen and Methocarbamol is already approved by FDA [67].
The combination is very crucial for the patient suffering from muscle spam, pain and
inflammation. Therefore, the simultaneous analysis of these two drugs ibuprofen and
methocarbomol is highly desirable as this will allow more efficient clinical data generation
in the patients who are co-infected with inflammation [68].
Literature survey revealed that there is no reported simultaneous equation method
is available for the estimation of ibuprofen and methocarbomolby UV spectrophotometric
using the Vierodt’s method. The non-availability of suitable modern analytical method for

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the concurrent analysis of this combination of drug prompted us to pursue the present
research work. It is also planned to validate the developed methods as per ICH guideline.

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2.2 PLAN OF WORK

Analysis of drugs will be carried out on following lines.

1. Review of Literature.
2. Procurement and Identification of the standard drugs.
3. Preliminary screening of the individual drugs.
4. Selection of common solvent for both drugs.
5. Analytical method development by UV spectrophotometer.
6. Validation of developed methods.
a. Specificity
b. Linearity
c. Accuracy
d. Precision
e. Robustness
f. Ruggedness
g. Limit of Detection
h. Limit of Quantization
7. Application of developed method.
8. Statistical evaluation and compilation of data.

3.0 Drug Profile

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3.1 IBUPROFEN [69]
a. Molecular Structure

5
H3C
6
4 O

CH3 3 2
1 1
OH
2

CH3
3

b. Chemical Name: (2RS)-2-[4-(2-Methylpropyl)phenyl]propanoic acid.

c. Molecular Formula: C13H18O2
d. Molecular Weight: 206.3g/mol
e. Category: Cyclo-oxygenase inhibitor; analgesic; anti-inflammatory.
f. Description: White or almost white; crystalline powder or colourless crystals.

g. Description: Ibuprofen is a non-steroidal anti-inflammatory drug (NSAID) derived from

propionic acid and it isconsidered the first of the propionics. The formula of ibuprofen is 2-
(4-isobutylphenyl) propionicacid and its initial development was in 1960 while
researching for a safer alternative for aspirin.Ibuprofen was finally patented in 1961 and
this drug was first launched against rheumatoidarthritis in the UK in 1969 and USA in
1974. It was the first available over-the-counter NSAID.On the available products,
ibuprofen is administered as a racemic mixture. Once administered, theR-enantiomer
undergoes extensive interconversion to the S-enantiomer in vivo by the activity ofthe
alpha-methylacyl-CoA racemase. In particular, it is generally proposed that the S-nantiomer
iscapable of eliciting stronger pharmacological activity than the R-enantiomer

h. Solubility: Practically insoluble in water, freely soluble in acetone, in methanol and in

methylenechloride.
i. Identification: Melting Point: 75-78C

j. Storage: Store in a tightly closed container in a dry place.

k. Indication: Ibuprofen is the most commonly used and prescribed NSAID. It is very
common over the countermedication widely used as an analgesic, anti-inflammatory and

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antipyretic.The use of ibuprofen and its enantiomer Dexibuprofen in a racemic mix is
common for themanagement of mild to moderate pain related to dysmenorrhea, headache,
migraine,postoperative dental pain, spondylitis, osteoarthritis, rheumatoid arthritis, and so̭
tissuedisorder.Due to its activity against prostaglandin and thromboxane synthesis,
ibuprofen has beenattributed to alteration of platelet function and prolongation of
gestation and labor.

l. Mechanism of action:Ibuprofen is considered anNSAID and thus it is a non-selective

inhibitor of cyclooxygenase, which is an enzyme involved inprostaglandin (mediators of
pain and fever) and thromboxane (stimulators of blood clotting)synthesis via the
arachidonic acid pathway.Ibuprofen is a non-selective COX inhibitor and hence, it inhibits
the activity of both COX-1 andCOX-2. The inhibition of COX-2 activity decreases the
synthesis of prostaglandins involved inmediating inflammation, pain, fever, and swelling
while the inhibition of COX-1 is thought to causesome of the side effects of ibuprofen
including GI ulceration.

m.International brands: Act-3 (Healthcare Logistics) / Actiprofen (Sterling Health) /

Adex (Dexxon) / Aktren (Bayer) / Alges-X (Axapharm) / Algoflex(Chinoin) / Algofren
(Intermed) / Alivium (Bristol-Myers Squibb) / Artofen (Teva) / Betagesic (Adcock Ingram
Pharmaceuticals) /Betaprofen (Be-Tabs Pharmaceuticals) / Bonifen (Krka) / Brufen
(Abbott) / Brupro (Sandoz).

3.2 Methocarbamol [70]

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a. IUPAC Name:2-hydroxy-3-(2-methoxyphenoxy)propyl carbamate

b. Chemical structure

3
O
4
O H3C 2

1 3 1
2 5
H2N O O
6

OH

c. Molecular weight: 241.24

d. Molecular Formula: C11H15NO5

e. Synonyms: Methocarbamolum

f. Description: Methocarbamol was developed in the early 1950s as a treatment for

musclespasticity and theassociated pain. It is a guaiacol glyceryl ether.Methocarbamol
tablets and intramuscular injections are prescription medicines indicated in theUnited
States as an adjunct to rest, physical therapy, and other measures for the relief
ofdiscomforts associated with acute, painful musculoskeletal conditions. In
Canada,methocarbamol can be sold as an over the counter oral medicine at a lower dose
that may becombined with acetaminophen or ibuprofen. A combination product with
acetylsalicylic acidand codeine is available in Canada by prescription.Methocarbamol was
FDA approved on 16 July 1957.

g. Indication: Methocarbamol tablets and intramuscular injections are indicated in the

United States as anadjunct to rest, physical therapy, and other measures for the relief of
discomforts associated withacute, painful musculoskeletal conditions. Oral methocarbamol
in America may be given upto 1500mg 4 times daily for 2-3 days.

h. Mechanism of action: The mechanism of action of methocarbamol is thought to be

dependant on its central nervoussystem depressant activity. This action may be mediated
through blocking spinal polysynapticreflexes, decreasing nerve transmission in spinal and
supraspinal polysynaptic pathways, andprolonging the refractory period of muscle cells.

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Methocarbamol has been found to have noeffect on contraction of muscle fibres, motor end
plates, or nerve fibres.

i. Protein binding: Methocarbamol is 46-50% protein bound in healthy patients and 47.3-
48.9% protein bound inhemodialysis patients.

j. Half-life: The elimination half-life is 1.14 hours in healthy subjects and 1.24 hours in
subjects with renalinsufficiency.

k. Clearance: 0.2-0.8L/h/kg

l. International/Other Brands:Bolaxin (Ying Yuan) / Carbaflex (Paill) / Delaxin

/DoloVisan (Dr.KadePharmaceutische Fabrik) / Fubaxin (Grape King) /
Lumirelax(Juvise) / Metocarbamol (AZ Pharma) / Mioflex (Labinco) / Miorel (Cheminter) /
Musxan (Pharmasant) / Ortoton (Recordati) /Rebamol (Winston) / Robaxin-750 (Pfizer) /
Robinax (Khandelwal) / Sinaxar (Armofar) / Taspan (Honten)

m. Manufacturers:Marsam pharmaceuticals, Watson laboratories, Baxter healthcare,

Ferndale laboratories, Able laboratories, Impax laboratories, Mylan pharmaceuticals,
Roxane laboratories, Sandoz, Solvay pharmaceuticals, Schwarz pharma

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