research article development and validation of uv...
TRANSCRIPT
Hindawi Publishing CorporationISRN SpectroscopyVolume 2013 Article ID 534830 4 pageshttpdxdoiorg1011552013534830
Research ArticleDevelopment and Validation of UV SpectrophotometricMethod for Simultaneous Estimation of Hesperidin and Diosminin the Pharmaceutical Dosage Form
Doddi Srilatha Mahesh Nasare Borra Nagasandhya Valluri Prasad and Prakash Diwan
Department of Pharmaceutical Analysis and Quality Assurance Anurag Group of Institutions Venkatapur GhatkesarRanga Reddy 501301 India
Correspondence should be addressed to Doddi Srilatha pinkybcmgmailcom
Received 8 August 2013 Accepted 5 September 2013
Academic Editors P Forgo and I P Gerothanassis
Copyright copy 2013 Doddi Srilatha et al This is an open access article distributed under the Creative Commons Attribution Licensewhich permits unrestricted use distribution and reproduction in any medium provided the original work is properly cited
A simple rapid precise and highly selective spectrophotometric method was developed for simultaneous estimation of Hesperidinand Diosmin in tablet dosage form This method involves the measurement of absorbances of Hesperidin and Diosmin at thewavelengths of 285 nm (120582max of Hesperidin) and 268 nm (120582max of Diosmin)TheUV spectrarsquos of Hesperidin andDiosmin preparedin different solvents water methanol and acetonitrile and 02N sodium hydroxide were recorded These two drugs showed goodabsorbances when dissolved in 02N NaOH Hence 02N NaOH was selected as the solvent for the method Two wavelengths 285and 268 nmwere selectedwhich are120582max of twodrugsHesperidin andDiosmin respectivelyDifferent concentrations ofHesperidin(5ndash50 120583gmL) andDiosmin (2ndash24 120583gmL) and amixture of Hesperidin andDiosmin were prepared scanned and absorbances werenoted at the two wavelengths were fixed for the studyThemethod showed good reproducibility and recovery with RSD less than2 The LOD of Hesperidin and Diosmin was found to be 0139 120583gmL and 0048 120583gml and LOQ of Hesperidin and Diosmin wasfound to be 042120583gmL and 0147 120583gmL respectively Thus the proposed method was found to be rapid specific precise accurateand cost effective quality control tool for the routine analysis of Hesperidin and Diosmin in bulk and combined dosage form
1 Introduction
Hesperidin is 3101584057-trihydroxy-41015840methoxy flavanone 7-o-120573-rutinoside which is greatly found in citrus species and is theactive constituent of tangerine peel (citrus reticulate anti-inflammatory effect by the inhibition of eicosanoids synthesisother biological activities including blood cholesterol lower-ing effect hypotensive effect protective agent against sepsisantioxidant activity and diuretics) It prevents poisoningcaused by lead strontium and heavy metals It is also used fordiabetics and gastroesophageal reflux diseases Hesperidinconverted to hesperitn by intestinal microflora and subse-quently absorbed from intestinal mucosa (see Figure 1)
Diosmin is a synthetic drug (modified Hesperidin) of theflavonoid family It is an oral phlebotropic drug used in thetreatment of venous disease that is chronic venous insuffi-ciently (CVI) and hemorrhoidal diseases Diosmin aglyconeis diosmetin IUPAC name is 3101584057-trihydroxy-4 methoxy
flavone 7-rutinosode It might have potential in the treatmentof neurodegenerative disease such as alzheimerrsquos diseasesand its anti-inflammatory and antiapoptotic activities havebeen demonstrated in neuronal cells It reduces venoushyperpressure present in patients suffering from CVI It alsoimproves lymphatic drainage by increasing the frequency andintensity of lymphatic contraction and by increasing the totalnumber of functional lymphatic capillaries (see Figure 2)
The Literature Review According to the literature survey itwas found that few analytical methods such as UV [1ndash5]Fluorometric method [6] LC-method [7] LC-MS method[8] HPLC method [9 10] capillary electrophoresis [11]simple colorimetric method [12] thin layer chromatography(TLC) [13] and adsorptive stripping voltammetry [14] werereported for hesperidin and Diosmin To our knowledgeno study related to the UV spectrophotometric method forthe simultaneous estimation of hesperidin and diosmin has
2 ISRN Spectroscopy
O O
O
OO
O
O
OH
OH
HO
HO
HO
HO
OH
HO
Figure 1 Chemical structure of Hesperidin
O
O
O
OO
O
O
OH
OH
HO
HO
HO
HO
HO
HO
Figure 2 Chemical structure of Diosmin
been reported in literature Therefore there is a challengeto develop UV spectrophotometric method for the simul-taneous estimation of hesperidin and diosmin The presentstudy was involved in a research effort aimed at developingand validating a simple specific accurate economical andprecise UV spectrophotometric method for the simultaneousestimation of two drugs in pharmaceutical dosage form
2 Materials and Methods
21 Materials and Reagents Hesperidin and Diosmin work-ing standards were procured from Mylan LaboratoriesHyderabad India Commercially available Daflon 500mgwas purchased from local pharmacy Methanol Acetonitrileand Sodium hydroxide were obtained from Merck Chem-icals Water was prepared by using Millipore Milli Q Pluswater purification system
22 Instrument UV-Visible double beam spectrophotometer(Shimadzu Model 1800) was employed with a spectral bandwidth of 1 nm and a wavelength accuracy of 03 nm (with
1
1
2
2
44
5Absorbance
06
04
02
0220 250 300 350 400 450
(nm)
Hesperidin
Diosmin
Figure 3 Overlain normal spectra of Hesperidin and Diosmin in02N NaOH
automatic correction with a pair of 1 cm matched quartzcells)
23 Method The UV spectra of Hesperidin and Diosmin indifferent solvents like water methanol ethanol acetonitrilesodium hydroxide (02N) were recorded The two drugsshowed good absorbances when dissolved in 02N NaOHHence 02N NaOH was selected as the solvent for themethod two wavelengths 285 and 268 nm were selectedwhich are 120582max two drugs Hesperidin and Diosmin respec-tively (see Figure 3)
Different concentrations of Hesperidin (5ndash50120583gmL)and Diosmin (2ndash24120583gmL) and a mixture of Hesperidinand Diosmin were prepared and scanned Two absorbanceswere noted at the two wavelengths fixed for the study Theabsorbance of Hesperidin and Diosmin was measured andabsorptivity values 119864 (1 1 cm) were determined at all thewavelengths The concentration of two drugs in mixture canbe calculated using the following equations
119862Hesperidin =(11986021198861199101 minus 11986011198861199102)
11988611990921198861199101minus 11988611990911198861199102
119862Diosmin =(11986011198861199092 minus 11986021198861199091)
11988611990921198861199101minus 11988611990911198861199102
(1)
where119862Hesperidin and 119862Diosmin are the concentrations of Hes-peridin and Diosmin respectively in mixture and in samplesolution 1198601 and 1198602 are the absorbances of sample at 285 nmand 268 nm respectively 1198861199091 and 1198861199092 are the absorptivityof Hesperidin at 285 nm and 268 nm respectively 1198861199101 and1198861199102 are the absorptivity of Diosmin at 285 nm and 268 nmrespectively (see Figures 4 and 5)
3 Results and Discussion
The analytical method was validated with respect to param-eters such as linearity precision accuracy limit of detection(LOD) limit of quantification (LOQ) and ruggedness
ISRN Spectroscopy 3Ab
sorbance
109080706050403020100 10 20 30 40 50 60
Concentration
y = 0018x minus 0003
R2= 0999
Figure 4 Calibration curve of Hesperidin at 285 nm
Absorbance
Concentration
12
1
08
06
04
02
00 5 10 15 20 25 30
y = 0041x minus 0021
R2= 0999
Figure 5 Calibration curve of Diosmin at 268 nm
Table 1 Linearity values of Hesperidin and Diosmin
Parameter Hesperidin DiosminRegression equation 119884 = 0018119909 minus 0003 119884 = 004119909 + 0021
Linearity 120583gmL 5ndash50 2ndash24Correlation coefficient 0999 0999
Table 2 Precision values of Hesperidin and Diosmin
Drug Concentration(120583gmL)
Intraday( RSD)
Interday( RSD)
Hesperidin 20 021 0055Diosmin 10 014 021
31 Linearity Linearity was established by least squareslinear regression analysis of the calibration curve and thecalibration curves were linear over the concentration rangeof 5ndash50120583gmL for hesperidin and 2ndash24 120583gmL for DiosminAbsorbances were plotted versus respective concentrationsand linear regression analysis was performed on the resultantcurves correlation coefficient was found to be 0999 and0999 for hesperidin and Diosmin respectively (see Table 1)
32 Precision To check the degree of repeatability of themethod suitable statistical evaluation was carried out Theconcentrations of the two drugs were measured 3 times onthe same day at intervals of one hour and on three differentdays for intra- and inter-day study The standard deviation(SD) and relative standard deviation (RSD) were calculatedThe results are given in Table 2
33 Accuracy Recovery studies were carried out by applyingthe method to a drug sample to which known amount of
1
2
4
3
220 250 300 350 400 450
(nm)
0
1
2
3
Absorbance
Figure 6 Spectrum of Hesperidin and Diosmin
standard Hesperidin and Diosmin corresponding to 50 100and 150 of label claim had been added At each level of theamount three determinations were performed The resultsare given in Table 3
34 LOD and LOQ The LOD of hesperidin and diosminwas found to be 0139 120583gmL and 0048120583gmL respectivelyand the LOQ was found to be 0042120583gmL and o147 120583gmLrespectively the results are given in Table 4
35 Analysis of Marketed Formulation Twenty tablets(Daflon 500mg) were powdered and the average weightwas calculated The Quantity equivalent to 10mg of drugwas dissolved in 02N NAOH and to it 8mg of (standardaddition method) such that sample contains 9mg of eachHesperidin and Diosmin Absorbances were noted at 285 nmand 268 nm respectively (see Table 5 and Figure 6)
4 Conclusion
This method is considered simple reliable and selectiveproviding satisfactory accuracy precision with lower limitsof detection more specific quantification and sensitivity Thegood recoveries were obtained in all cases and the reliableagreement with the reported procedure proved that theproposedmethod could be applied efficiently for the determi-nation of Hesperidin and Diosmin in oral dosage form withsatisfactory precisionmoreover the shorter duration of anal-ysis of Hesperidin and Diosmin marks the reported methodsuitable for routine analysis in pharmaceutical dosage forms
Authorsrsquo Contribution
Author Srilatha managed the literature survey and analysisof the study and wrote the first draft of the paper AuthorMahesh Nasare designed the study plan and authors Dr VV L N Prasad and Dr Prakash V Diwan provided chemicalsand instruments
4 ISRN Spectroscopy
Table 3 Recovery values of Hesperidin and Diosmin
Drug Recovery RSD50 100 150 50 100 150
Hesperidin 1006 9866 101 0289 0260 0281Diosmin 102 993 100 0280 0204 0239 RSD of three observations
Table 4 LOD and LOQ of Hesperidin and Diosmin
Drug LOD LOQHesperidin 0139 042Diosmin 0048 0147
Table 5 Analysis of formulation and its RSD values
Drug Amount labeled Amount found label claim RSDHesperidin 50 498 996 024Diosmin 450 452 1004 019
Acknowledgment
The authors are grateful to the School of Pharmacy AnuragGroup of Institutions Hyderabad for giving permission tocarry out their research work
References
[1] Z Moldovan A A Bunaciu M Al-Omara and H Y Aboul-Enein ldquoA spectrophotometric method for diosmin determina-tionrdquoThe Open Chemical and Biomedical Methods Journal vol3 pp 123ndash127 2010
[2] V Kuntic N Pejic and S Micic ldquoDirect spectrophotometricdetermination of hesperidin in pharmaceutical preparationsrdquoActa Chimica Slovenica vol 59 pp 436ndash441 2012
[3] S Wei X Ze and L Xin Detection of Hesperidin in OrangeSkin By the UV-VIS Spectrophotometry Chinese Journal ofSpectroscopy Laboratory 2005
[4] A El Bayoumi ldquoModified H-point standard addition methodand logarithmic function for the spectrophotometric and spec-trodensitometric determination of hesperidin and diosmin inmixturesrdquo Analytical Letters vol 32 no 2 pp 383ndash400 1999
[5] Z Moldovan and H Y Aboul-Enein ldquoA sensitive spectropho-tometric method for determination of diosmin using sodiumnitroprusside as a chromogenic reagentrdquo Instrumentation Sci-ence and Technology vol 39 no 2 pp 135ndash148 2011
[6] L A Pavun J M D Markovic P T Durdevic et al ldquoDevel-opment and validation of a fluorometric method for the deter-mination of hesperidin in human plasma and pharmaceuticalformsrdquo Journal of the Serbian Chemical Society vol 77 no 11pp 1625ndash1640 2012
[7] AM El-Shafae andMM El-Domiaty ldquoImproved LCmethodsfor the determination of diosmin andor hesperidin in plantextracts and pharmaceutical formulationsrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 26 no 4 pp 539ndash5452001
[8] M A Campanero M Escolar G Perez E Garcia-QuetglasB Sadaba and J R Azanza ldquoSimultaneous determination of
diosmin and diosmetin in human plasma by ion trap liq-uid chromatography-atmospheric pressure chemical ionizationtandem mass spectrometry application to a clinical phar-macokinetic studyrdquo Journal of Pharmaceutical and BiomedicalAnalysis vol 51 no 4 pp 875ndash881 2010
[9] I Saeidi M R Hadjmohammadi M Peyrovi et al ldquoHPLCdetermination of hesperidin diosmin and eriocitrin in Iranianlime juice using polyamide as an adsorbent for solid phaseextractionrdquo Journal of Pharmaceutical and Biomedical Analysisvol 56 no 2 pp 419ndash422 2011
[10] L S Mazzaferro and J D Breccia ldquoQuantification of hesperidinin citrus-based foods using a fungal diglycosidaserdquo Food Chem-istry vol 134 no 4 pp 2338ndash2344 2012
[11] G Chen L Zhang J Zhao and J Ye ldquoDetermination ofhesperidin and synephrine in Pericarpium Citri Reticulataeby capillary electrophoresis with electrochemical detectionrdquoAnalytical and Bioanalytical Chemistry vol 373 no 3 pp 169ndash173 2002
[12] S A Mir A A Ahangar and A S Bhat ldquoA new simple col-orimetric method for assaying diosmin and flavonoids indaflon tablets and orange peel extractsrdquo International Journalof PharmTech Research vol 5 no 2 pp 341ndash348
[13] Z Janeczko U Hubicka J Krzek and I Podolak ldquoQualitativeand quantitative analysis of diosmin in tablets by thin-layerchromatography with densitometric UV detectionrdquo Journal ofPlanar Chromatography Modern TLC vol 16 no 5 pp 377ndash380 2003
[14] M S El-Shahawi A S Bashammakh and T El-Mogy ldquoDeter-mination of trace levels of diosmin in a pharmaceutical prepa-ration by adsorptive stripping voltammetry at a glassy carbonelectroderdquo Analytical Sciences vol 22 no 10 pp 1351ndash13542006
Submit your manuscripts athttpwwwhindawicom
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Inorganic ChemistryInternational Journal of
Hindawi Publishing Corporation httpwwwhindawicom Volume 2014
International Journal ofPhotoenergy
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Carbohydrate Chemistry
International Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Journal of
Chemistry
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Advances in
Physical Chemistry
Hindawi Publishing Corporationhttpwwwhindawicom
Analytical Methods in Chemistry
Journal of
Volume 2014
Bioinorganic Chemistry and ApplicationsHindawi Publishing Corporationhttpwwwhindawicom Volume 2014
SpectroscopyInternational Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014
Medicinal ChemistryInternational Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Chromatography Research International
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Applied ChemistryJournal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Theoretical ChemistryJournal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Journal of
Spectroscopy
Analytical ChemistryInternational Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Quantum Chemistry
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Organic Chemistry International
ElectrochemistryInternational Journal of
Hindawi Publishing Corporation httpwwwhindawicom Volume 2014
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
CatalystsJournal of
2 ISRN Spectroscopy
O O
O
OO
O
O
OH
OH
HO
HO
HO
HO
OH
HO
Figure 1 Chemical structure of Hesperidin
O
O
O
OO
O
O
OH
OH
HO
HO
HO
HO
HO
HO
Figure 2 Chemical structure of Diosmin
been reported in literature Therefore there is a challengeto develop UV spectrophotometric method for the simul-taneous estimation of hesperidin and diosmin The presentstudy was involved in a research effort aimed at developingand validating a simple specific accurate economical andprecise UV spectrophotometric method for the simultaneousestimation of two drugs in pharmaceutical dosage form
2 Materials and Methods
21 Materials and Reagents Hesperidin and Diosmin work-ing standards were procured from Mylan LaboratoriesHyderabad India Commercially available Daflon 500mgwas purchased from local pharmacy Methanol Acetonitrileand Sodium hydroxide were obtained from Merck Chem-icals Water was prepared by using Millipore Milli Q Pluswater purification system
22 Instrument UV-Visible double beam spectrophotometer(Shimadzu Model 1800) was employed with a spectral bandwidth of 1 nm and a wavelength accuracy of 03 nm (with
1
1
2
2
44
5Absorbance
06
04
02
0220 250 300 350 400 450
(nm)
Hesperidin
Diosmin
Figure 3 Overlain normal spectra of Hesperidin and Diosmin in02N NaOH
automatic correction with a pair of 1 cm matched quartzcells)
23 Method The UV spectra of Hesperidin and Diosmin indifferent solvents like water methanol ethanol acetonitrilesodium hydroxide (02N) were recorded The two drugsshowed good absorbances when dissolved in 02N NaOHHence 02N NaOH was selected as the solvent for themethod two wavelengths 285 and 268 nm were selectedwhich are 120582max two drugs Hesperidin and Diosmin respec-tively (see Figure 3)
Different concentrations of Hesperidin (5ndash50120583gmL)and Diosmin (2ndash24120583gmL) and a mixture of Hesperidinand Diosmin were prepared and scanned Two absorbanceswere noted at the two wavelengths fixed for the study Theabsorbance of Hesperidin and Diosmin was measured andabsorptivity values 119864 (1 1 cm) were determined at all thewavelengths The concentration of two drugs in mixture canbe calculated using the following equations
119862Hesperidin =(11986021198861199101 minus 11986011198861199102)
11988611990921198861199101minus 11988611990911198861199102
119862Diosmin =(11986011198861199092 minus 11986021198861199091)
11988611990921198861199101minus 11988611990911198861199102
(1)
where119862Hesperidin and 119862Diosmin are the concentrations of Hes-peridin and Diosmin respectively in mixture and in samplesolution 1198601 and 1198602 are the absorbances of sample at 285 nmand 268 nm respectively 1198861199091 and 1198861199092 are the absorptivityof Hesperidin at 285 nm and 268 nm respectively 1198861199101 and1198861199102 are the absorptivity of Diosmin at 285 nm and 268 nmrespectively (see Figures 4 and 5)
3 Results and Discussion
The analytical method was validated with respect to param-eters such as linearity precision accuracy limit of detection(LOD) limit of quantification (LOQ) and ruggedness
ISRN Spectroscopy 3Ab
sorbance
109080706050403020100 10 20 30 40 50 60
Concentration
y = 0018x minus 0003
R2= 0999
Figure 4 Calibration curve of Hesperidin at 285 nm
Absorbance
Concentration
12
1
08
06
04
02
00 5 10 15 20 25 30
y = 0041x minus 0021
R2= 0999
Figure 5 Calibration curve of Diosmin at 268 nm
Table 1 Linearity values of Hesperidin and Diosmin
Parameter Hesperidin DiosminRegression equation 119884 = 0018119909 minus 0003 119884 = 004119909 + 0021
Linearity 120583gmL 5ndash50 2ndash24Correlation coefficient 0999 0999
Table 2 Precision values of Hesperidin and Diosmin
Drug Concentration(120583gmL)
Intraday( RSD)
Interday( RSD)
Hesperidin 20 021 0055Diosmin 10 014 021
31 Linearity Linearity was established by least squareslinear regression analysis of the calibration curve and thecalibration curves were linear over the concentration rangeof 5ndash50120583gmL for hesperidin and 2ndash24 120583gmL for DiosminAbsorbances were plotted versus respective concentrationsand linear regression analysis was performed on the resultantcurves correlation coefficient was found to be 0999 and0999 for hesperidin and Diosmin respectively (see Table 1)
32 Precision To check the degree of repeatability of themethod suitable statistical evaluation was carried out Theconcentrations of the two drugs were measured 3 times onthe same day at intervals of one hour and on three differentdays for intra- and inter-day study The standard deviation(SD) and relative standard deviation (RSD) were calculatedThe results are given in Table 2
33 Accuracy Recovery studies were carried out by applyingthe method to a drug sample to which known amount of
1
2
4
3
220 250 300 350 400 450
(nm)
0
1
2
3
Absorbance
Figure 6 Spectrum of Hesperidin and Diosmin
standard Hesperidin and Diosmin corresponding to 50 100and 150 of label claim had been added At each level of theamount three determinations were performed The resultsare given in Table 3
34 LOD and LOQ The LOD of hesperidin and diosminwas found to be 0139 120583gmL and 0048120583gmL respectivelyand the LOQ was found to be 0042120583gmL and o147 120583gmLrespectively the results are given in Table 4
35 Analysis of Marketed Formulation Twenty tablets(Daflon 500mg) were powdered and the average weightwas calculated The Quantity equivalent to 10mg of drugwas dissolved in 02N NAOH and to it 8mg of (standardaddition method) such that sample contains 9mg of eachHesperidin and Diosmin Absorbances were noted at 285 nmand 268 nm respectively (see Table 5 and Figure 6)
4 Conclusion
This method is considered simple reliable and selectiveproviding satisfactory accuracy precision with lower limitsof detection more specific quantification and sensitivity Thegood recoveries were obtained in all cases and the reliableagreement with the reported procedure proved that theproposedmethod could be applied efficiently for the determi-nation of Hesperidin and Diosmin in oral dosage form withsatisfactory precisionmoreover the shorter duration of anal-ysis of Hesperidin and Diosmin marks the reported methodsuitable for routine analysis in pharmaceutical dosage forms
Authorsrsquo Contribution
Author Srilatha managed the literature survey and analysisof the study and wrote the first draft of the paper AuthorMahesh Nasare designed the study plan and authors Dr VV L N Prasad and Dr Prakash V Diwan provided chemicalsand instruments
4 ISRN Spectroscopy
Table 3 Recovery values of Hesperidin and Diosmin
Drug Recovery RSD50 100 150 50 100 150
Hesperidin 1006 9866 101 0289 0260 0281Diosmin 102 993 100 0280 0204 0239 RSD of three observations
Table 4 LOD and LOQ of Hesperidin and Diosmin
Drug LOD LOQHesperidin 0139 042Diosmin 0048 0147
Table 5 Analysis of formulation and its RSD values
Drug Amount labeled Amount found label claim RSDHesperidin 50 498 996 024Diosmin 450 452 1004 019
Acknowledgment
The authors are grateful to the School of Pharmacy AnuragGroup of Institutions Hyderabad for giving permission tocarry out their research work
References
[1] Z Moldovan A A Bunaciu M Al-Omara and H Y Aboul-Enein ldquoA spectrophotometric method for diosmin determina-tionrdquoThe Open Chemical and Biomedical Methods Journal vol3 pp 123ndash127 2010
[2] V Kuntic N Pejic and S Micic ldquoDirect spectrophotometricdetermination of hesperidin in pharmaceutical preparationsrdquoActa Chimica Slovenica vol 59 pp 436ndash441 2012
[3] S Wei X Ze and L Xin Detection of Hesperidin in OrangeSkin By the UV-VIS Spectrophotometry Chinese Journal ofSpectroscopy Laboratory 2005
[4] A El Bayoumi ldquoModified H-point standard addition methodand logarithmic function for the spectrophotometric and spec-trodensitometric determination of hesperidin and diosmin inmixturesrdquo Analytical Letters vol 32 no 2 pp 383ndash400 1999
[5] Z Moldovan and H Y Aboul-Enein ldquoA sensitive spectropho-tometric method for determination of diosmin using sodiumnitroprusside as a chromogenic reagentrdquo Instrumentation Sci-ence and Technology vol 39 no 2 pp 135ndash148 2011
[6] L A Pavun J M D Markovic P T Durdevic et al ldquoDevel-opment and validation of a fluorometric method for the deter-mination of hesperidin in human plasma and pharmaceuticalformsrdquo Journal of the Serbian Chemical Society vol 77 no 11pp 1625ndash1640 2012
[7] AM El-Shafae andMM El-Domiaty ldquoImproved LCmethodsfor the determination of diosmin andor hesperidin in plantextracts and pharmaceutical formulationsrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 26 no 4 pp 539ndash5452001
[8] M A Campanero M Escolar G Perez E Garcia-QuetglasB Sadaba and J R Azanza ldquoSimultaneous determination of
diosmin and diosmetin in human plasma by ion trap liq-uid chromatography-atmospheric pressure chemical ionizationtandem mass spectrometry application to a clinical phar-macokinetic studyrdquo Journal of Pharmaceutical and BiomedicalAnalysis vol 51 no 4 pp 875ndash881 2010
[9] I Saeidi M R Hadjmohammadi M Peyrovi et al ldquoHPLCdetermination of hesperidin diosmin and eriocitrin in Iranianlime juice using polyamide as an adsorbent for solid phaseextractionrdquo Journal of Pharmaceutical and Biomedical Analysisvol 56 no 2 pp 419ndash422 2011
[10] L S Mazzaferro and J D Breccia ldquoQuantification of hesperidinin citrus-based foods using a fungal diglycosidaserdquo Food Chem-istry vol 134 no 4 pp 2338ndash2344 2012
[11] G Chen L Zhang J Zhao and J Ye ldquoDetermination ofhesperidin and synephrine in Pericarpium Citri Reticulataeby capillary electrophoresis with electrochemical detectionrdquoAnalytical and Bioanalytical Chemistry vol 373 no 3 pp 169ndash173 2002
[12] S A Mir A A Ahangar and A S Bhat ldquoA new simple col-orimetric method for assaying diosmin and flavonoids indaflon tablets and orange peel extractsrdquo International Journalof PharmTech Research vol 5 no 2 pp 341ndash348
[13] Z Janeczko U Hubicka J Krzek and I Podolak ldquoQualitativeand quantitative analysis of diosmin in tablets by thin-layerchromatography with densitometric UV detectionrdquo Journal ofPlanar Chromatography Modern TLC vol 16 no 5 pp 377ndash380 2003
[14] M S El-Shahawi A S Bashammakh and T El-Mogy ldquoDeter-mination of trace levels of diosmin in a pharmaceutical prepa-ration by adsorptive stripping voltammetry at a glassy carbonelectroderdquo Analytical Sciences vol 22 no 10 pp 1351ndash13542006
Submit your manuscripts athttpwwwhindawicom
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Inorganic ChemistryInternational Journal of
Hindawi Publishing Corporation httpwwwhindawicom Volume 2014
International Journal ofPhotoenergy
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Carbohydrate Chemistry
International Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Journal of
Chemistry
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Advances in
Physical Chemistry
Hindawi Publishing Corporationhttpwwwhindawicom
Analytical Methods in Chemistry
Journal of
Volume 2014
Bioinorganic Chemistry and ApplicationsHindawi Publishing Corporationhttpwwwhindawicom Volume 2014
SpectroscopyInternational Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014
Medicinal ChemistryInternational Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Chromatography Research International
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Applied ChemistryJournal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Theoretical ChemistryJournal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Journal of
Spectroscopy
Analytical ChemistryInternational Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Quantum Chemistry
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Organic Chemistry International
ElectrochemistryInternational Journal of
Hindawi Publishing Corporation httpwwwhindawicom Volume 2014
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
CatalystsJournal of
ISRN Spectroscopy 3Ab
sorbance
109080706050403020100 10 20 30 40 50 60
Concentration
y = 0018x minus 0003
R2= 0999
Figure 4 Calibration curve of Hesperidin at 285 nm
Absorbance
Concentration
12
1
08
06
04
02
00 5 10 15 20 25 30
y = 0041x minus 0021
R2= 0999
Figure 5 Calibration curve of Diosmin at 268 nm
Table 1 Linearity values of Hesperidin and Diosmin
Parameter Hesperidin DiosminRegression equation 119884 = 0018119909 minus 0003 119884 = 004119909 + 0021
Linearity 120583gmL 5ndash50 2ndash24Correlation coefficient 0999 0999
Table 2 Precision values of Hesperidin and Diosmin
Drug Concentration(120583gmL)
Intraday( RSD)
Interday( RSD)
Hesperidin 20 021 0055Diosmin 10 014 021
31 Linearity Linearity was established by least squareslinear regression analysis of the calibration curve and thecalibration curves were linear over the concentration rangeof 5ndash50120583gmL for hesperidin and 2ndash24 120583gmL for DiosminAbsorbances were plotted versus respective concentrationsand linear regression analysis was performed on the resultantcurves correlation coefficient was found to be 0999 and0999 for hesperidin and Diosmin respectively (see Table 1)
32 Precision To check the degree of repeatability of themethod suitable statistical evaluation was carried out Theconcentrations of the two drugs were measured 3 times onthe same day at intervals of one hour and on three differentdays for intra- and inter-day study The standard deviation(SD) and relative standard deviation (RSD) were calculatedThe results are given in Table 2
33 Accuracy Recovery studies were carried out by applyingthe method to a drug sample to which known amount of
1
2
4
3
220 250 300 350 400 450
(nm)
0
1
2
3
Absorbance
Figure 6 Spectrum of Hesperidin and Diosmin
standard Hesperidin and Diosmin corresponding to 50 100and 150 of label claim had been added At each level of theamount three determinations were performed The resultsare given in Table 3
34 LOD and LOQ The LOD of hesperidin and diosminwas found to be 0139 120583gmL and 0048120583gmL respectivelyand the LOQ was found to be 0042120583gmL and o147 120583gmLrespectively the results are given in Table 4
35 Analysis of Marketed Formulation Twenty tablets(Daflon 500mg) were powdered and the average weightwas calculated The Quantity equivalent to 10mg of drugwas dissolved in 02N NAOH and to it 8mg of (standardaddition method) such that sample contains 9mg of eachHesperidin and Diosmin Absorbances were noted at 285 nmand 268 nm respectively (see Table 5 and Figure 6)
4 Conclusion
This method is considered simple reliable and selectiveproviding satisfactory accuracy precision with lower limitsof detection more specific quantification and sensitivity Thegood recoveries were obtained in all cases and the reliableagreement with the reported procedure proved that theproposedmethod could be applied efficiently for the determi-nation of Hesperidin and Diosmin in oral dosage form withsatisfactory precisionmoreover the shorter duration of anal-ysis of Hesperidin and Diosmin marks the reported methodsuitable for routine analysis in pharmaceutical dosage forms
Authorsrsquo Contribution
Author Srilatha managed the literature survey and analysisof the study and wrote the first draft of the paper AuthorMahesh Nasare designed the study plan and authors Dr VV L N Prasad and Dr Prakash V Diwan provided chemicalsand instruments
4 ISRN Spectroscopy
Table 3 Recovery values of Hesperidin and Diosmin
Drug Recovery RSD50 100 150 50 100 150
Hesperidin 1006 9866 101 0289 0260 0281Diosmin 102 993 100 0280 0204 0239 RSD of three observations
Table 4 LOD and LOQ of Hesperidin and Diosmin
Drug LOD LOQHesperidin 0139 042Diosmin 0048 0147
Table 5 Analysis of formulation and its RSD values
Drug Amount labeled Amount found label claim RSDHesperidin 50 498 996 024Diosmin 450 452 1004 019
Acknowledgment
The authors are grateful to the School of Pharmacy AnuragGroup of Institutions Hyderabad for giving permission tocarry out their research work
References
[1] Z Moldovan A A Bunaciu M Al-Omara and H Y Aboul-Enein ldquoA spectrophotometric method for diosmin determina-tionrdquoThe Open Chemical and Biomedical Methods Journal vol3 pp 123ndash127 2010
[2] V Kuntic N Pejic and S Micic ldquoDirect spectrophotometricdetermination of hesperidin in pharmaceutical preparationsrdquoActa Chimica Slovenica vol 59 pp 436ndash441 2012
[3] S Wei X Ze and L Xin Detection of Hesperidin in OrangeSkin By the UV-VIS Spectrophotometry Chinese Journal ofSpectroscopy Laboratory 2005
[4] A El Bayoumi ldquoModified H-point standard addition methodand logarithmic function for the spectrophotometric and spec-trodensitometric determination of hesperidin and diosmin inmixturesrdquo Analytical Letters vol 32 no 2 pp 383ndash400 1999
[5] Z Moldovan and H Y Aboul-Enein ldquoA sensitive spectropho-tometric method for determination of diosmin using sodiumnitroprusside as a chromogenic reagentrdquo Instrumentation Sci-ence and Technology vol 39 no 2 pp 135ndash148 2011
[6] L A Pavun J M D Markovic P T Durdevic et al ldquoDevel-opment and validation of a fluorometric method for the deter-mination of hesperidin in human plasma and pharmaceuticalformsrdquo Journal of the Serbian Chemical Society vol 77 no 11pp 1625ndash1640 2012
[7] AM El-Shafae andMM El-Domiaty ldquoImproved LCmethodsfor the determination of diosmin andor hesperidin in plantextracts and pharmaceutical formulationsrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 26 no 4 pp 539ndash5452001
[8] M A Campanero M Escolar G Perez E Garcia-QuetglasB Sadaba and J R Azanza ldquoSimultaneous determination of
diosmin and diosmetin in human plasma by ion trap liq-uid chromatography-atmospheric pressure chemical ionizationtandem mass spectrometry application to a clinical phar-macokinetic studyrdquo Journal of Pharmaceutical and BiomedicalAnalysis vol 51 no 4 pp 875ndash881 2010
[9] I Saeidi M R Hadjmohammadi M Peyrovi et al ldquoHPLCdetermination of hesperidin diosmin and eriocitrin in Iranianlime juice using polyamide as an adsorbent for solid phaseextractionrdquo Journal of Pharmaceutical and Biomedical Analysisvol 56 no 2 pp 419ndash422 2011
[10] L S Mazzaferro and J D Breccia ldquoQuantification of hesperidinin citrus-based foods using a fungal diglycosidaserdquo Food Chem-istry vol 134 no 4 pp 2338ndash2344 2012
[11] G Chen L Zhang J Zhao and J Ye ldquoDetermination ofhesperidin and synephrine in Pericarpium Citri Reticulataeby capillary electrophoresis with electrochemical detectionrdquoAnalytical and Bioanalytical Chemistry vol 373 no 3 pp 169ndash173 2002
[12] S A Mir A A Ahangar and A S Bhat ldquoA new simple col-orimetric method for assaying diosmin and flavonoids indaflon tablets and orange peel extractsrdquo International Journalof PharmTech Research vol 5 no 2 pp 341ndash348
[13] Z Janeczko U Hubicka J Krzek and I Podolak ldquoQualitativeand quantitative analysis of diosmin in tablets by thin-layerchromatography with densitometric UV detectionrdquo Journal ofPlanar Chromatography Modern TLC vol 16 no 5 pp 377ndash380 2003
[14] M S El-Shahawi A S Bashammakh and T El-Mogy ldquoDeter-mination of trace levels of diosmin in a pharmaceutical prepa-ration by adsorptive stripping voltammetry at a glassy carbonelectroderdquo Analytical Sciences vol 22 no 10 pp 1351ndash13542006
Submit your manuscripts athttpwwwhindawicom
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Inorganic ChemistryInternational Journal of
Hindawi Publishing Corporation httpwwwhindawicom Volume 2014
International Journal ofPhotoenergy
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Carbohydrate Chemistry
International Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Journal of
Chemistry
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Advances in
Physical Chemistry
Hindawi Publishing Corporationhttpwwwhindawicom
Analytical Methods in Chemistry
Journal of
Volume 2014
Bioinorganic Chemistry and ApplicationsHindawi Publishing Corporationhttpwwwhindawicom Volume 2014
SpectroscopyInternational Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014
Medicinal ChemistryInternational Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Chromatography Research International
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Applied ChemistryJournal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Theoretical ChemistryJournal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Journal of
Spectroscopy
Analytical ChemistryInternational Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Quantum Chemistry
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Organic Chemistry International
ElectrochemistryInternational Journal of
Hindawi Publishing Corporation httpwwwhindawicom Volume 2014
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
CatalystsJournal of
4 ISRN Spectroscopy
Table 3 Recovery values of Hesperidin and Diosmin
Drug Recovery RSD50 100 150 50 100 150
Hesperidin 1006 9866 101 0289 0260 0281Diosmin 102 993 100 0280 0204 0239 RSD of three observations
Table 4 LOD and LOQ of Hesperidin and Diosmin
Drug LOD LOQHesperidin 0139 042Diosmin 0048 0147
Table 5 Analysis of formulation and its RSD values
Drug Amount labeled Amount found label claim RSDHesperidin 50 498 996 024Diosmin 450 452 1004 019
Acknowledgment
The authors are grateful to the School of Pharmacy AnuragGroup of Institutions Hyderabad for giving permission tocarry out their research work
References
[1] Z Moldovan A A Bunaciu M Al-Omara and H Y Aboul-Enein ldquoA spectrophotometric method for diosmin determina-tionrdquoThe Open Chemical and Biomedical Methods Journal vol3 pp 123ndash127 2010
[2] V Kuntic N Pejic and S Micic ldquoDirect spectrophotometricdetermination of hesperidin in pharmaceutical preparationsrdquoActa Chimica Slovenica vol 59 pp 436ndash441 2012
[3] S Wei X Ze and L Xin Detection of Hesperidin in OrangeSkin By the UV-VIS Spectrophotometry Chinese Journal ofSpectroscopy Laboratory 2005
[4] A El Bayoumi ldquoModified H-point standard addition methodand logarithmic function for the spectrophotometric and spec-trodensitometric determination of hesperidin and diosmin inmixturesrdquo Analytical Letters vol 32 no 2 pp 383ndash400 1999
[5] Z Moldovan and H Y Aboul-Enein ldquoA sensitive spectropho-tometric method for determination of diosmin using sodiumnitroprusside as a chromogenic reagentrdquo Instrumentation Sci-ence and Technology vol 39 no 2 pp 135ndash148 2011
[6] L A Pavun J M D Markovic P T Durdevic et al ldquoDevel-opment and validation of a fluorometric method for the deter-mination of hesperidin in human plasma and pharmaceuticalformsrdquo Journal of the Serbian Chemical Society vol 77 no 11pp 1625ndash1640 2012
[7] AM El-Shafae andMM El-Domiaty ldquoImproved LCmethodsfor the determination of diosmin andor hesperidin in plantextracts and pharmaceutical formulationsrdquo Journal of Pharma-ceutical and Biomedical Analysis vol 26 no 4 pp 539ndash5452001
[8] M A Campanero M Escolar G Perez E Garcia-QuetglasB Sadaba and J R Azanza ldquoSimultaneous determination of
diosmin and diosmetin in human plasma by ion trap liq-uid chromatography-atmospheric pressure chemical ionizationtandem mass spectrometry application to a clinical phar-macokinetic studyrdquo Journal of Pharmaceutical and BiomedicalAnalysis vol 51 no 4 pp 875ndash881 2010
[9] I Saeidi M R Hadjmohammadi M Peyrovi et al ldquoHPLCdetermination of hesperidin diosmin and eriocitrin in Iranianlime juice using polyamide as an adsorbent for solid phaseextractionrdquo Journal of Pharmaceutical and Biomedical Analysisvol 56 no 2 pp 419ndash422 2011
[10] L S Mazzaferro and J D Breccia ldquoQuantification of hesperidinin citrus-based foods using a fungal diglycosidaserdquo Food Chem-istry vol 134 no 4 pp 2338ndash2344 2012
[11] G Chen L Zhang J Zhao and J Ye ldquoDetermination ofhesperidin and synephrine in Pericarpium Citri Reticulataeby capillary electrophoresis with electrochemical detectionrdquoAnalytical and Bioanalytical Chemistry vol 373 no 3 pp 169ndash173 2002
[12] S A Mir A A Ahangar and A S Bhat ldquoA new simple col-orimetric method for assaying diosmin and flavonoids indaflon tablets and orange peel extractsrdquo International Journalof PharmTech Research vol 5 no 2 pp 341ndash348
[13] Z Janeczko U Hubicka J Krzek and I Podolak ldquoQualitativeand quantitative analysis of diosmin in tablets by thin-layerchromatography with densitometric UV detectionrdquo Journal ofPlanar Chromatography Modern TLC vol 16 no 5 pp 377ndash380 2003
[14] M S El-Shahawi A S Bashammakh and T El-Mogy ldquoDeter-mination of trace levels of diosmin in a pharmaceutical prepa-ration by adsorptive stripping voltammetry at a glassy carbonelectroderdquo Analytical Sciences vol 22 no 10 pp 1351ndash13542006
Submit your manuscripts athttpwwwhindawicom
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Inorganic ChemistryInternational Journal of
Hindawi Publishing Corporation httpwwwhindawicom Volume 2014
International Journal ofPhotoenergy
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Carbohydrate Chemistry
International Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Journal of
Chemistry
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Advances in
Physical Chemistry
Hindawi Publishing Corporationhttpwwwhindawicom
Analytical Methods in Chemistry
Journal of
Volume 2014
Bioinorganic Chemistry and ApplicationsHindawi Publishing Corporationhttpwwwhindawicom Volume 2014
SpectroscopyInternational Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014
Medicinal ChemistryInternational Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Chromatography Research International
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Applied ChemistryJournal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Theoretical ChemistryJournal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Journal of
Spectroscopy
Analytical ChemistryInternational Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Quantum Chemistry
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Organic Chemistry International
ElectrochemistryInternational Journal of
Hindawi Publishing Corporation httpwwwhindawicom Volume 2014
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
CatalystsJournal of
Submit your manuscripts athttpwwwhindawicom
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Inorganic ChemistryInternational Journal of
Hindawi Publishing Corporation httpwwwhindawicom Volume 2014
International Journal ofPhotoenergy
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Carbohydrate Chemistry
International Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Journal of
Chemistry
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Advances in
Physical Chemistry
Hindawi Publishing Corporationhttpwwwhindawicom
Analytical Methods in Chemistry
Journal of
Volume 2014
Bioinorganic Chemistry and ApplicationsHindawi Publishing Corporationhttpwwwhindawicom Volume 2014
SpectroscopyInternational Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014
Medicinal ChemistryInternational Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Chromatography Research International
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Applied ChemistryJournal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Theoretical ChemistryJournal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Journal of
Spectroscopy
Analytical ChemistryInternational Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Journal of
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Quantum Chemistry
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
Organic Chemistry International
ElectrochemistryInternational Journal of
Hindawi Publishing Corporation httpwwwhindawicom Volume 2014
Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014
CatalystsJournal of