determinationoffluoroquinolonesinpharmaceutical...

Research ArticleDetermination of Fluoroquinolones in PharmaceuticalFormulations by Extractive Spectrophotometric Methods UsingIon-Pair Complex Formation with Bromothymol Blue

Trung Dung Nguyen 1 Hoc Bau Le1 Thi Oanh Dong1 and Tien Duc Pham 2

1Faculty of Physics and Chemical Engineering Le Quy Don Technical University 236 Hoang Quoc Viet Hanoi Vietnam2Faculty of Chemistry VNU-University of Science Vietnam National University Hanoi 19 Le )anh Tong Hoan KiemHanoi Vietnam

Correspondence should be addressed to Trung Dung Nguyen nguyentrungdung1980gmailcom and Tien Duc Phamtienduchphngmailcom

Received 11 March 2018 Revised 16 June 2018 Accepted 1 August 2018 Published 4 October 2018

Academic Editor Bengi Uslu

Copyright copy 2018 Trung Dung Nguyen et al +is is an open access article distributed under the Creative Commons AttributionLicense which permits unrestricted use distribution and reproduction in any medium provided the original work isproperly cited

In this paper we reported a new simple accurate and precise extractive spectrophotometric method for the determination of flu-oroquinolones (FQs) including ciprofloxacin (CFX) levofloxacin (LFX) and ofloxacin (OFX) in pharmaceutical formulations +eproposed method is based on the ion-pair formation complexes between FQs and an anionic dye bromothymol blue (BTB) in acidicmedium+e yellow-colored complexes whichwere extracted into chloroformweremeasured at thewavelengths of 420 415 and 418nmfor CFX LFX and OFX respectively Some effective conditions such as pH dye concentration shaking time and organic solvents werealso systematically studied Very good limit of detection (LOD) of 0084microgmL 0101microgmL and 0105microgmLwere found for CFX LFXand OFX respectively +e stoichiometry of the complexes formed between FQs and BTB determined by Jobrsquos method of continuousvariation was 1 1 No interference was observed from common excipients occurred in pharmaceutical formulations +e proposedmethod has been successfully applied to determine the FQs in some pharmaceutical products A good agreement between extractivespectrophotometric method with high-performance liquid chromatography mass spectrometry (HPLC-MS) for the determination ofFQs in some real samples demonstrates that the proposed method is suitable to quantify FQs in pharmaceutical formulations

1 Introduction

Fluoroquinolones (FQs) are the important antibiotics usedfor the treatment of Gram-negative bacterial infections inboth human and veterinary medicine+ey are derivatives of4-quinolone which have unsubstituted or substituted pi-perazine ring attached at the 7-position to the central ringsystem of quinoline as well as fluorine atom at the 6-position+e FQs are useful to treat a variety of infections includingsoft-tissue infections respiratory infections urinary tractinfections bone-joint infections typhoid fever prostatitissexually transmitted diseases acute bronchitis community-acquired pneumonia and sinusitis [1ndash3]

Ciprofloxacin (CFX) which is one of the second-generated groups of synthetic FQs can exhibit greater

intrinsic antibacterial activity and make a broader anti-bacterial spectrum Ofloxacin (OFX) is a chiral compoundthat is widely used to treat above infections Levofloxacin(LFX) is the pure (ndash)-(S)-enantiomer of the racemic drugsubstance ofloxacin Figures 1(a)ndash1(c) show the chemicalstructures of CFX LFX and OFX respectively

Several techniques like voltammetry [4] flow injectionelectrogenerated chemiluminescence [5] spectrofluorome-try [6 7] spectrophotometry [8 9] high-performance liquidchromatography [10 11] and liquid chromatography tan-dem mass spectrometry [12 13] have been used for thedetermination of fluoroquinolones in pharmaceutical andbiological products Among them spectrophotometricmethod has several advantages such as simplicity fast andlow cost Spectrophotometry was successfully used for

HindawiJournal of Analytical Methods in ChemistryVolume 2018 Article ID 8436948 11 pageshttpsdoiorg10115520188436948

pharmaceutical analysis involving quality control of com-mercialized product and pharmacodynamic studies Spec-trophotometric methods for the determination offluoroquinolones could be classified according to the dif-ferent reactions (i) charge-transfer complexation based onthe reaction of FQs as electron donors with p-acceptorssuch as 23-dichloro-56-dicyano-q-benzoquinone 7788-tetracyanoquinodimethane q-chloranil q-nitrophenoland tetracyanoethylene [7 14ndash16] (ii) oxidative couplingreaction using oxidative coupling with 3-methyl-2-benzothiazolinonehydrazone hydrochloride and cerium(IV) ammonium sulfate Fe(III)-MBTH tris(o-phenan-throline) iron(II) and tris (bipyridyl) iron(II) [17 18] (iii)ion-pair complex formation with acid-dye reagents such asSudan III methyl orange supracene violet 3B tropaeolin000 bromophenol blue bromothymol blue bromocresolgreen and bromocresol purple [8 14 19 20]+esemethodswere related with some major drawbacks such as havingnarrow linearity range requiring heating and close pHcontrol long time for the reaction to complete and lowstability of the colored product formed

Bromothymol blue (BTB) (Figure 1(d)) is an anionic dyeand that can be protonated or deprotonated to form yellowor blue respectively +e BTB was used to make ion-paircomplex which was applied to determine many pharma-ceutical compounds by extractive spectrophotometricmethods [21ndash30] However the ion-pair complex betweenBTB and FQs has not been studied +e method based onion-pair complexes between analytes and BTB into a suitableorganic solvent is also simple fast and cheap

In the previous study we used sulphonphthalein acidincluding bromophenol blue bromocresol green and bro-mothymol blue to determine ciprofloxacin pharmaceuticalformulations and achieved good results [31]

In this paper for the first time we investigated extractivespectrophotometric method based on the formation of ion-pair complexes between ciprofloxacin levofloxacin and

ofloxacin with BTB subsequent extraction into chloroformSome effective conditions on the formation of complexessuch as pH shaking time organic solvent and the con-centration of dye were systematically studied +e presentmethod was also applied to determine FQs in some phar-maceutical formulations including tablets and infusions

2 Experimental

21 Apparatus A double beam UV-visible spectropho-tometer (SP-60 Biochrom Ltd UK) with 10 cm of pathlength quartz cells was used to measure all sample absor-bances Inolab pH-meter instrument (Germany) was used tomonitor the pH of solutions +ree standard buffers wereused to calibrate the electrode before measuring pH ofsolutions All measurements were conducted at 25 plusmn 2degCcontrolled by air conditional laboratory

22 Materials and Reagents All chemicals used were ofanalytical grade and double-distilled water was used toprepare all solutions in the present study

FQs were purchased from Sigma (Germany with puritygt990) whereas bromothymol blue (BTB) was supplied byMaya-R China with purity gt99 +e organic solventsincluding chloroform dichloromethane carbon tetrachlo-ride dichloroethane benzene toluene and other chemicalsare analytical reagents (Merck Germany)

+e following dosage forms containing FQs were pur-chased from local pharmacy market and employed in thestudy Hasancip and Kacipro tablets equivalent to 500mgciprofloxacin (Hasan-Dermapharm and Dong Nammanufacturing-Trading pharmaceutical Co Ltd Vietnam)Ciprofloxacin infusion equivalent to 200mg cipro-floxacin100ml solution for infusion (Hebei TianchengPharmaceutical Co Ltd and Shandong Hualu Pharma-ceutical Co Ltd China) Stada and DHG tablets equivalent

HN

N

F

N

O O

OH

(a)

H3C

OH

OO

ONCH3

F

N N

(b)

N

F

N

O

O O

NH3C CH3

OH

(c)

CH3CH3

C3H7C3H7

SO2

OHHO

BrBr OC

(d)

Figure 1 Chemical structures of ciprofloxacin (a) levofloxacin (b) ofloxacin (OFX) (c) and bromothymol blue (d)

2 Journal of Analytical Methods in Chemistry

to 500mg levofloxacin (Stada-VN JVCompany and DHGpharmaceutical jointndashstock company Vietnam) Ofloxacin(200mgtablet) was provided by the Mekophar ChemicalPharmaceutical Company (Vietnam)

23 Solution Preparation A stock solution of FQs (1mgmL)in double-distilled water +e working standard solution ofFQs containing 100 microgmL was prepared by appropriate di-lution +e stock solution of BTB (0025) was prepared indouble-distilled water All stock solutions were kept in darkbottle stored in 4degC and could be used within one week

24 Construction of Calibration Curves A series of 125mLseparating funnel the volumes of working solutions of thedrugs in different concentration ranges (CFX (1ndash35 microgmL)LFX (05ndash25 microgmL) and OFX (05ndash25 microgmL) were trans-ferred +en 40mL of 0025 BTB solution was addedbefore thoroughly mixing After that a 10mL of chloroformwas added to each of the separating funnel +e contentswere shaken for 2min and allowed to separate the two layers+e yellow-colored chloroform layer containing the ion-paircomplexes was measured at 420 nm for CFX 415 nm forLFX and 418 nm for OFX against the reagent blanks At eachconcentration the experiment was repeated 6 times +ecolored chromogen complexes are stable for 24 h

25 Sample Preparation Weigh and mix the contents oftwenty tablets of each drug (CFX LFX and OFX) an ac-curately weighed amount of powder equivalent to 01 g ofdrugs transferred into a 100-mL beaker A magnetic stirrerwas used to completely disintegrate the powder in doublydistilled water +en filter through a Whatman paper (No40) and fill up to 100mL with doubly distilled water ina volumetric flask +e working solution of the drugscontaining 100 microgmL was prepared by dilution and de-termined under optimum conditions

26ValidationwithHigh-PerformanceLiquidChromatography-Mass Spectrometry (HPLC-MS) Some real samples of threeFQs were determined by HPLC-MS using HPLC 20 AXL(Shimadzu Japan) coupled with electrospray ionisationtandem mass spectrometric detection ABI 5500 QQQ (Ap-plied BioSystem) +e chromatographic conditions are in-cluding column C18 MRC-ODS (150mmtimes 21mmtimes 35 microm)mobile phase containing acetonitrile (ACN) with formic acid(01) in water under a flow rate of 05mlmin and gradientelution +e inject volume is 10microL

3 Results and Discussion

31 Optimum Reaction Conditions

311 Effect of Extracting Solvent Six organic solvents in-cluding chloroform carbon tetrachloride dichloromethanedichloroethane benzene and toluene were used to study theeffect of solvent to ion-pair formation between FQs andBTB Figure 2 shows that chloroform is the most suitable

solvent for the extraction of three FQs with low blank ab-sorbance highest absorbances and lowest standard de-viations It implies that chloroform is the best extractingsolvent to achieve a good recovery of the complexes with theshortest time to reach the equilibrium processes

312 Effect of pH +e pH of solution plays an importantrole in the complex formations +e effect of pH on theformation of ion pairs was examined by varying the pH from20 to 60 by adjusting 1M HCl and 1M NaOH +emaximum absorbances were observed at pH 33 34 and 35for the complexes of BTB and OFX CFX and LFX re-spectively (Figure 3) +ese pH values correspond to theinitial pH of the examined drug and the dye +erefore it isnot necessary to adjust the pH before extraction

313 Effect of Dye Concentration +e effect of dye con-centrations was studied by adding different volumes of0025 BTB from 10 to 60mL with a fixed concentration ofFQs (10 μgmL) (Figure 4) Figure 4 shows that the maxi-mum absorbance of the complex was achieved with 40mLof 0025 of BTB in each case and excess dye did not affectthe absorbance of the complex +erefore 40mL of 0025of BTB is optimum dye volume and it is kept as constant forfurther studies

314 Effect of Shaking Time +e effect of shaking time onthe formation and stability of the ion-pair complex wasinvestigated by measuring the absorbance of the extract-ed ion associates with increasing time from 0 to 40minFigure 5 shows that the ion-pair complexes were formed

1 2 3 4 5 600

02

04

06

08

10

Abs

orba

nce

Solvent type

OFX-BTBLFX-BTBCFX-BTB

(1) Chloroform(2) Dichloromethane(3) Dichloroethane(4) Benzene(5) Toluene(6) Carbon tetrachloride

Figure 2 +e effect of solvent on the ion-pair complex formation(10 microgmL of fluoroquinolones (FQs) with bromothymol blue(BTB))

Journal of Analytical Methods in Chemistry 3

instantaneously with 20min shaking time +us 20min isthe optimum shaking time and it is fixed for further studies

315 Stoichiometry of Ion-Pair Complexes Jobrsquos method ofcontinuous variation of equimolar solutions was employedto evaluate stoichiometry of the complex A 30times10minus4Mstandard solution of three FQs and 30times10minus4M solution ofBTB were used A series of the solutions were prepared inwhich the total volume of drug and reagent was kept in

10mL whereas the absorbances were measured at 420 415and 418 nm for CFX LFX and OFX respectively +eabsorbances were plotted against the mole fraction of thedrugs +e stoichiometry for each drug-dye ion-pair com-plex was found to be 1 1 (Figure 6)

316 Mechanism of Reaction and Absorption SpectraFluoroquinolones can contain a secondary amino group(CFX) and a tertiary amino group (LFX and OFX) that canbe easily protonated under acidic conditions On the onehand the sulphonic acid group in BTB that is the onlygroup undergoing dissociation in the pH range 1ndash5 +ecolour of BTB is on the basis of lactoid ring and subsequentformation of quinoid group It is suggested that the twotautomers are plausible in equilibrium due to strong acidicnature of the sulphonic acid group +us the quinoid bodymust predominate Finally the protonated fluo-roquinolones form ion pairs with BTB dye that could bequantitatively extracted into chloroform +e possible re-action mechanisms are proposed and given in a scheme inFigure 7

+e absorption spectra of the ion-pair complexes whichwere formed between FQs and BTB were measured in thewavelength range 350ndash500 nm against the blank solutionand shown in Figure 8

Figure 8 shows that absorption maxima for CFX-BTBLFX-BTB and OFX-BTB in chloroform were observed at420 415 and 418 nm respectively +e reagent blanks undersimilar conditions have insignificant absorbances Atwavelengths 420 415 and 418 nm absorption spectrum ofBTB does not affect the absorption spectrum of ion-associatecomplexes of FQs +erefore the selectivity of the proposedmethod for the determination of FQs is guaranteed

0 1 2 3 4 500

02

04

06

08

10

Abs

orba

nce

Shaking time (min)


Figure 5 Effect of shaking time on the ion-pair complexes

1 2 3 4 5 6 700

02

04

06

08

10

Abs

orba

nce

pH


Figure 3 Effect of pH on the absorbances of 10 μgmL of OFXLFX and CFX

0 1 2 3 4 5 6 700

02

04

06

08

10

Abs

orba

nce

Volume of dye (ml)


Figure 4 Effect of the volume of 0025 BTB on the absorbance of10 μgmL of OFX LFX and CFX


317 Association Constants of Ion-Pair Complexes +eequation of association constant of ion-pair complex is

AAm

1minus AAm( 11138571113858 1113859n+2

CM(n)n

(1)

where A and Am are the observed absorbance and the max-imum absorbance value when all the drug present is associatedrespectively CM is the molar concentration of the drugs at themaximum absorbance and n is the stoichiometry in which BTBion associates with drugs +e conditional stability constants(Kf) of the ion-pair complexes according to Britton [32] for thecases of FQs were calculated from the continuous variationdata using the following equation

Kf AAm

1minusAAm1113858 1113859n+2

CM(n)n (2)

+e conditional stability constants (Kf) of the ion-paircomplexes for FQs are indicated in Table 1

Table 1 shows that the log Kf values of ion-pair associatesfor OFX-BTB LFX-BTB and CFX-BTB were 608plusmn 046604plusmn 058 and 591plusmn 032 respectively (numbers of repli-cated experiments n 6) +e obtained results confirmedthat the ion-pair formation complexes are of high stability

32Validationof thePresentMethod +eproposedmethodsare validated according to ICH recommendations Q2(R1)[33] +e parameters that have been investigated are in-dicated below

321 Linearity Sensitivity and Limits of Detection andQuantification A linear relationship between the measured

absorbance and the concentration range studied for eachdrug as shown in Figure 9 and the correlation coefficient (R)of at least 0997 were achieved +e limit of detection (LOD)and quantification (LOQ) of the method are determined by33(SDb) and 10(SDb) respectively where SD is thestandard deviation of blank absorbance values and b is theslope of the calibration curve equation

+e LOD and LOQ values slope and intercept of lineargraphs for all the drugs and analytical parameters are in-dicated in Table 2 +e molar absorptivities and Sandellrsquossensitivity of each methods were calculated and these valuesshowed that the molar absorptivity of ion-pair complexeswas in the order CFX-BTBgt LFX-BTBgtOFX-BTB

322 Accuracy and Precision +e accuracy and precision ofthe methods were determined by preparing solutions ofthree different concentrations of drug and analyzing them insix replicates +e precision of the proposed methods wasevaluated as percentage relative standard deviation (RSD)and accuracy as percentage relative error (RE) +e per-centage relative error was calculated using the followingequation

RE() (foundedndashadded)

added1113890 1113891 times 100 (3)

+e accuracy and precision were summarized in Table 3+e low values of the RSD and RE confirm the high precisionand the good accuracy of the present method

323 Robustness and Ruggedness For the evaluation of themethod robustness some parameters were interchangedpH dye concentration wavelength range and shaking time+e capacity remains unaffected by small deliberate varia-tions Method ruggedness was expressed as RSD of thesame procedure applied by two analysts and using differentinstruments on different days +e results showed no sta-tistical differences between different analysts and in-struments suggesting that the developed methods wererobust and rugged (Table 4)

324 Selectivity and Effect of Interferences +e effect ofcommonly utilized excipients in drug formulation wasstudied +e investigated FQs were studied with variousexcipients such as magnesium stearate glucose lactosestarch and sodium chloride which were prepared in theproportion corresponding to their amounts in the real drugswith a final dosage of 10 microgmL FQ +e effect of excipientson the determination of FQs was evaluated by recovery whendetermining FQs analyzed with the proposed method in thepresence of excipient (Table 5)

+e results in Table 5 show that the recoveries are in therange of 9853ndash10204 demonstrating that there is no in-terference of excipients when FQs in drugs are quantified byextractive spectrophotometric using ion-pair formation withBTB In other words the present method has a high se-lectivity for determining FQs in its dosage forms

00 02 04 06 08 1000

02

04

06

08

10

Abs

orba

nce

(Drug)(drug+BTB)


Figure 6 Jobrsquos method of continuous variation graph for thereaction of drug with acid dyes BTB [drug] [dye] 30times10minus4M


360 400 440 480 52000

02

04

06

08

10

Abs

orba

nce

Wavelength (nm)

1

4

2

3

(1) BTB(2) OFX-BTB(3) LFX-BTB(4) CFX-BTB

Figure 8 Absorption spectrum of ion-associate complexes of fluoroquinolones (10 microgmL) with BTB against reagent blank

CH3

O

BrCH3

H+

C

HO

Br

C3H7

CH3 CH3SO3H

Br

O

C

HO

Br

C3H7 C3H7C3H7

+

F

NNH

N

O

O

O

OH

+

C3H7

CH3SO2

C3H7HO OH

OC BrBrCH3

Bromothymol blue(lactoid ring)

Protonated levofloxacin

Bromothymol blue(lactoid ring) (Quinoid ring)

Anion bromothymol blue

(Quinoid ring)

SO3ndash

+

C3H7 C3H7O

Br

HO

Br CCH3 CH3

SO3ndash

Levofloxacin-bromothymol blue complex

CH3 CH3

+

O

OH

O

NNONH

+

F

C3H7 C3H7HO O

Br+ H+

Br CCH3 CH3

SO3H

Br

OC3H7 C3H7

HO

CBr

C3H7

CH3CH3 SO2

OC

C3H7OHHO

BrBr

SO3ndash

Protonated ofloxacin


HO

C

O

BrBr

C3H7 C3H7

CH3 CH3SO3

ndash

Ofloxacin-bromothymol blue complex

Br CCH3 CH3

C3H7 C3H7

F

N N

O

O

O

OHHO O

Br

NHSO3

+ ndash

F

NO

N

O O

OHHO

C

O

BrBr

NH

C3H7 C3H7

CH3 CH3SO3

ndash+

Figure 7 Proposal mechanism for the reaction between levofloxacin ofloxacin and bromothymol blue


33 Comparison with Other Spectrophotometric Methods+e proposed method compares with other reportedmethods It has been observed that the extractive spectro-photometric method with BTB in the present study is of highsensitivity than other ones (Table 6) It also does not needheating the product is stable for a longer time and theinterferences are minimum

34 Analysis of Pharmaceutical Formulations +e proposedmethod was applied successfully for the determination ofstudied drugs in the pharmaceutical formulations (tablets

0 10 20 30 4000

04

08

12

16

20


Abs

orba

nce

Concentration of drug (μgmiddotmLndash1)

Figure 9 Calibration curves for OFX LFX and CFX at 418 415and 420 nm respectively

Table 2 Analytical characteristics of the proposed methods(n 6)

ParametersProposed methods

Ofloxacin Levofloxacin CiprofloxacinColour Yellow Yellow YellowWavelengths λmax (nm) 418 415 420pH 33 35 34Stability (h) 24 24 24Shaking time (min) 2 2 2Stoichiometric ratio 1 1 1 1 1 1Beerrsquos law range(microgmL) 1ndash35 05ndash25 05ndash25

Limit of detectionLOD (microgmL) 0105 0101 0084

Limit of quantitationLOQ (microgmL) 0315 0303 0252

Molar absorptivity(Lmolcm) 144times104 207times104 209times104

Sandellrsquos sensitivity(microgcm2) 0068 0048 0046

Regression equation (Y bx+ a) where Y is the absorbance a isthe intercept b is the slope and x is the concentration in μgmLSlope (b) 0040 0057 0061Intercept (a) 00165 0072 0089Correlationcoefficient (R) 0998 0997 0998

Table 1 +e conditional stability constants (Kf ) of the ion-pair complexes for FQs

Sample Vdrug (mL) VBTB (mL) A n nandn [1minus (AAm)]n+2 Kf logKf MeanOfloxacin

1 025 225 0202 01111 07834 03116 502047802 47007 6082 05 2 0272 02500 07071 01486 1570489127 519603 075 175 0336 04286 06955 00512 5725077789 575784 1 15 0419 06667 07631 00035 95623298320 698065 125 125 0476 10000 10000 00000 _ _6 15 1 0432 15000 18371 00002 594141776247 777397 175 075 0356 23333 72213 00026 11722461806 60690

Levofloxacin1 025 225 0137 01111 07834 05749 147898587 41700 6042 05 2 0313 02500 07071 01856 1159613839 506433 075 175 0432 04286 06955 00426 7085741920 585044 1 15 0559 06667 07631 00005 677452456475 783095 125 125 0594 10000 10000 00000 _ _6 15 1 053 15000 18371 00004 341653128945 753367 175 075 0426 23333 72213 00042 6828970665 58344

Ciprofloxacin1 025 225 0392 01111 07834 02112 835300520 49218 5912 05 2 0452 02500 07071 01265 1781450100 525083 075 175 0564 04286 06955 00345 8284800305 591834 1 15 0661 06667 07631 00036 85222134055 693065 125 125 0752 10000 10000 00000 _ _6 15 1 0615 15000 18371 00026 45722671248 666017 175 075 0538 23333 72213 00043 6087986008 57845mdash not determined


and infusion) and the results are presented in Table 7 Sixreplicated determinations were measured Table 7 shows thatsatisfactory recovery data were obtained and the recoveryefficiency varies from 9741 to 10120 indicating highaccuracy of the present method in determining real phar-maceutical samples

35ComparisonwithHPLC-MSMethod In order to validatethe experimental data in determining some real drugsamples HPLC-MS was used with the conditions describedon Section 26 according to the previously published paper[13] +e comparison between the results determined by thepresent method with HPLC-MS method was indicated inTable 8

Table 8 shows a good agreement between the proposedmethod and HPLC-MS where the relative differences of twomethods were less than 11 Furthermore the standarddeviation of the proposed method is almost lower thanthat of HPLC-MS Our results indicate that the extractivespectrophotometric determination of FQs using BTB dye in

chloroform is a very good method to quantify the FQ inpharmaceutical formulations

4 Conclusions

We have reported a new method when using BTB as ananionic dyes for the extractive spectrophotometric de-termination of ciprofloxacin (CFX) levofloxacin (LFX) andofloxacin (OFX) in different pharmaceutical drugs (tabletsand infusions) +e methods have the advantages of sim-plicity without heating pH-adjustment and high sensitivity+e limit of detection (LOD) values are 0084 microgmL forCFX 0101 microgmL for LFX and 0105 microgmL for OFX Nointerference from common excipients was confirmed +estoichiometry complexes of FQs and BTB determined byJobrsquos method of continuous variation were found to be 1 1+e developed and validated methods are indicated as theacceptable precision and accuracy and recovery of the drugsand suitable for routine analysis of drugs in pharmaceuticalformulations+e results of some real samples by the presentmethod that were compared with HPLC-MS method with

Table 3 Evaluation of accuracy and precision of the proposed methods (n 6)

Method Additive concentration (μgmL) Found concentration (μgmL) Recovery () RSD () RE ()

Ofloxacin500 511 10219 231 221000 1026 10264 134 261500 1489 9925 088 minus073

Levofloxacin500 516 10270 203 281000 1016 10156 110 161500 1482 9880 050 minus12

Ciprofloxacin500 513 10271 192 261000 974 9741 052 minus261500 1460 9733 057 minus27

Table 4 +e results of analysis of pharmaceutical preparation and standard of fluoroquinolones by two different analysts and instruments(n 6)

MethodDifferent instruments Different analysts

X plusmnSD RSD () X plusmnSD RSD ()Ofloxacin-BTB pure ofloxacin (10 microgmiddotmLminus1)Mekopharm (200mg ofloxacin per tablet)

1021 019 186 993 024 242199 057 029 196 076 039

Levofloxacin-BTB pure levofloxacin (10 microgmiddotmLminus1)Stada (500mg levofloxacin per tablet)

1016 015 148 1019 021 206498 061 012 501 085 017

Ciprofloxacin-BTB pure ciprofloxacin (10 microgmiddotmLminus1)Hasancip (500mg ciprofloxacin per tablet)

985 018 183 1012 025 247502 064 013 497 092 019

Table 5 +e effect of excipients on the determination of fluoroquinolones (10 microgmL)

Recovery ()plusmn SDExcipients Amount of excipient added (μgmL) Ofloxacin Levofloxacin CiprofloxacinMagnesium stearate 500 10204plusmn 012 10123plusmn 0089 9853plusmn 091Glucose 250 10017plusmn 016 9904plusmn 014 9908plusmn 0062Lactose 500 9992plusmn 021 10020plusmn 012 9973plusmn 021Starch 200 10096plusmn 024 9889plusmn 013 10131plusmn 017Sodium chloride 500 10013plusmn 024 10015plusmn 011 9975plusmn 016


Table 6 +e comparison of present study with other spectrophotometric methods

Drug Reagent λmax(nm)

Range ofdetermination

(μgmL)

Molarabsorptivity(Lmolmiddotcm)

Remarks Reference

Ciprofloxacin

Co (II) tetrathiocyanate 623 20ndash240 838times102 Less sensitive [34]Supracene violet 3 575 25ndash30 862times103 Less sensitive [35]

Eosin Y 547 2ndash8 356times104 Less stable colour [36]Merbromin 545 2ndash15 123times104 Addition of CNndashto inhibit Hg+2 ions

Ce(IV)- MBTH 630 10ndash50 mdash Involves shaking time [17]Tris(o-phenanthroline) iron(II) 510 004ndash72 34times104 Involves shaking time and heating [18]

Tris (bipyridyl) iron(II) 522 005ndash9 295times104 Involves shaking time and heatingCL 520 16ndash96 mdash Involves shaking time and heating [16]

TCNE 335 025ndash15 mdash Involves shaking time and heatingSudan II 550 08ndash71 53times104

Narrow linear range [8]Congo red 517 05ndash60 283times104

Gentian violet 585 05ndash10 221times 104

Brilliant blue G 610 05ndash60 286times104 Narrow linear range and requiredpH adjustment [37]

Bromocresol green 412 1ndash20 228times104 Required pH adjustment [14]

BTB 420 05ndash25 209times104Highly sensitive with wide lineardynamic ranges no heating and

no pH adjustment

+isstudy

Levofloxacin

Chloranilic acid 521 15ndash250 12times103 Less sensitive [14]Bromocresol green 411 1ndash20 216times104 Required pH adjustment


Cobalt (II) tetrathiocyanate 623 20ndash240 mdash Less sensitive [34]Bromophenol blue 424 185ndash315 198times104 Required pH adjustment [19]Bromocresol green 428 185ndash25 182times104


no pH-adjustment

+isstudy

Ofloxacin

Supracene violet 3 575 25ndash25 109times104 Less sensitive [35]Tropaeolin 000 485 25ndash30 823times102 Less sensitive

Sudan II 560 08ndash84 297times104



Bromocresol purple 400 10ndash160 2 4times104 Required pH adjustment [38]Bromocresol green 410 10ndash160 196times104 Required pH adjustmentBromophenol blue 410 5ndash25 103times104 Required close pH control and

involved extraction steps [20]Bromothymol blue 415 2ndash15 201times 104

Bromocresol purple 410 2ndash20 164times104

Bromothymol blue 415 1ndash35 144times104Highly sensitive with wide lineardynamic ranges no heating and no

pH-adjustment

+isstudy

Table 7 Determination of the studied drugs in their pharmaceutical preparations using the proposed method (n 6)

Pharmaceuticalpreparation

Hasanciptablet

Kaciprotablet

Shandonginfusion Hebei infusion Levofloxacin

StadaLevofloxacin

DHGOfloxacinmekopharm

Labeled amount (mgform) 500tablet 500tablet 200100mL 200100mL 500tablet 500tablet 200tablet

Recovery ()plusmn SD 9889plusmn023

10120plusmn020 9741plusmn 042 9769plusmn 036 9953plusmn 017 10101plusmn 035 9958plusmn 046

Table 8 Amount of some fluoroquinolone antibiotics determined by the proposed method and HPLC-MS

SampleAmount (mgtablet)

Difference ()Proposed method HPLC -MS

Ciprofloxacin-Hasancip table 49445plusmn 1163 44693plusmn 1584 1063Ofloxacin mekopharm 19916plusmn 085 20200plusmn 272 minus141Levofloxacin DHG 50505plusmn 1733 48055plusmn 5416 510Levofloxacin Stada 49765plusmn 924 48604plusmn 924 239


the relative differences are less than 11 indicating that thepresent method is good for determination of FQs inpharmaceutical formulations

Data Availability

+e data used to support the findings of this study areavailable from the corresponding author upon request

Conflicts of Interest

+e authors declare that they have no conflicts of interest

Acknowledgments

+is work was supported financially by the project of Le QuyDon Technical University under 11HLKT2017 project+e authors would like to thank the National Institute forFood Control (Vietnam) for providing HPLC-MS system tovalidate the present method

References

[1] V Kapetanovic L Milovanovic and M Erceg ldquoSpectro-photometric and polarographic investigation of theOfloxacin-Cu(II) complexesrdquo Talanta vol 43 no 12pp 2123ndash2130 1996

[2] Y Khaliq and G G Zhanel ldquoFluoroquinolone-associatedtendinopathy a critical review of the literaturerdquo ClinicalInfectious Diseases vol 36 no 11 pp 1404ndash1410 2003

[3] G G Zhanel K Ennis L Vercaigne et al ldquoA critical review ofthe fluoroquinolonesrdquo Drugs vol 62 no 1 pp 13ndash59 2002

[4] Y Ni YWang and S Kokot ldquoSimultaneous determination ofthree fluoroquinolones by linear sweep stripping voltammetrywith the aid of chemometricsrdquo Talanta vol 69 no 1pp 216ndash225 2006

[5] H Ma X Zheng and Z Zhang ldquoFlow-injection electro-generated chemiluminescence determination of fluo-roquinolones based on its sensitizing effectrdquo Luminescencevol 20 no 4-5 pp 303ndash306 2005

[6] S T Ulu ldquoSpectrofluorimetric determination of fluo-roquinolones in pharmaceutical preparationsrdquo Spectrochi-mica Acta Part A Molecular and Biomolecular Spectroscopyvol 72 no 1 pp 138ndash143 2009

[7] L M Du A P Lin and Y Q Yang ldquoSpectrofluorimetricdetermination of certain fluoroquinolone through chargetransfer complex formationrdquo Analytical Letters vol 37no 10 pp 2175ndash2188 2004

[8] A S Amin M E Moustafa and R M S El-DosokyldquoSpectrophotometric determination of some fluoroquinolonederivatives in dosage forms and biological fluids using ion-pair complex formationrdquo Analytical Letters vol 41 no 5pp 837ndash852 2008

[9] A M A-E El-Didamony and O Mona ldquoKinetic spectro-photometric method for the determination of some fourthgeneration fluoroquinolones in bulk and in pharmaceuticalformulationsrdquo Journal of Saudi Chemical Society vol 21pp S58ndashS66 2017

[10] M I R M Santoro N M Kassab A K Singh andE R M Kedor-Hackmam ldquoQuantitative determination ofgatifloxacin levofloxacin lomefloxacin and pefloxacin fluo-roquinolonic antibiotics in pharmaceutical preparations byhigh-performance liquid chromatographyrdquo Journal of

Pharmaceutical and Biomedical Analysis vol 40 no 1pp 179ndash184 2006

[11] G Carlucci ldquoAnalysis of fluoroquinolones in biological fluidsby high-performance liquid chromatographyrdquo Journal ofChromatography A vol 812 no 1-2 pp 343ndash367 1998

[12] H Ziarrusta N Val H Dominguez et al ldquoDetermination offluoroquinolones in fish tissues biological fluids and envi-ronmental waters by liquid chromatography tandem massspectrometryrdquo Analytical and Bioanalytical Chemistryvol 409 no 27 pp 6359ndash6370 2017

[13] L Johnston L Mackay and M Croft ldquoDetermination ofquinolones and fluoroquinolones in fish tissue and seafood byhigh-performance liquid chromatography with electrosprayionisation tandem mass spectrometric detectionrdquo Journal ofChromatography A vol 982 no 1 pp 97ndash109 2002

[14] A M El-Brashy M E-S Metwally and F A El-SepaildquoSpectrophotometric Determination of Some Fluo-roquinolone Antibacterials through Charge-transfer and Ion-pair Complexation Reactionsrdquo Bulletin of the KoreanChemical Society vol 25 no 3 pp 365ndash372 2004

[15] L M Du H Y Yao and M Fu ldquoSpectrofluorimetric study ofthe charge-transfer complexation of certain fluoroquinoloneswith 7788-tetracyanoquinodimethanerdquo Spectrochimica ActaPart A Molecular and Biomolecular Spectroscopy vol 61no 1-2 pp 281ndash286 2005

[16] S Mostafa M El-Sadek and E A Alla ldquoSpectrophotometricdetermination of ciprofloxacin enrofloxacin and pefloxacinthrough charge transfer complex formationrdquo Journal ofPharmaceutical and Biomedical Analysis vol 27 no 1-2pp 133ndash142 2002

[17] M Rizk F B F Ibrahim S M Ahmed and N M El-EnanyldquoA simple kinetic spectrophotometric method for the de-termination of certain 4-quinolones in drug formulationsrdquoScientia Pharmaceutica vol 68 no 2 pp 173ndash188 2000

[18] B S Nagaralli J Seetharamappa and M B MelwankildquoSensitive spectrophotometric methods for the determinationof amoxycillin ciprofloxacin and piroxicam in pure andpharmaceutical formulationsrdquo Journal of Pharmaceutical andBiomedical Analysis vol 29 no 5 pp 859ndash864 2002

[19] S Ashour and R Al-Khalil ldquoSimple extractive colorimetricdetermination of levofloxacin by acid-dye complexationmethods in pharmaceutical preparationsrdquo Farmaco vol 60no 9 pp 771ndash775 2005

[20] Y M Issa F M Abdel-Gawad M A Abou Table andH M Hussein ldquoSpectrophotometric determination ofofloxacin and lomefloxacin hydrochloride with some sul-phonphthalein dyesrdquo Analytical Letters vol 30 no 11pp 2071ndash2084 1997

[21] H A Omara and A S Amin ldquoExtractive-spectrophotometricmethods for determination of anti-Parkinsonian drug inpharmaceutical formulations and in biological samples usingsulphonphthalein acid dyesrdquo Journal of Saudi Chemical So-ciety vol 16 no 1 pp 75ndash81 2012

[22] A A Gouda A S Amin R El-Sheikh and A G YousefldquoSpectrophotometric determination of gemifloxacin mesylatemoxifloxacin hydrochloride and enrofloxacin in pharma-ceutical formulations using acid dyesrdquo Journal of AnalyticalMethods in Chemistry vol 2014 Article ID 286379 16 pages2014

[23] S G Nair J V Shah P A ShahM Sanyal and P S ShrivastavldquoExtractive spectrophotometric determination of five selecteddrugs by ion-pair complex formation with bromothymol bluein pure form and pharmaceutical preparationsrdquo CogentChemistry vol 1 no 1 article 1075852 2015


[24] NRahman and SNHejaz-Azmi ldquoExtractive spectrophotometricmethods for determination of diltiazem HCl in pharmaceuticalformulations using bromothymol blue bromophenol blue andbromocresol greenrdquo Journal of Pharmaceutical and BiomedicalAnalysis vol 24 no 1 pp 33ndash41 2000

[25] H E Abdellatef ldquoExtractive-spectrophotometric determinationof disopyramide and irbesartan in their pharmaceutical for-mulationrdquo Spectrochimica Acta Part A Molecular and Bio-molecular Spectroscopy vol 66 no 4 pp 1248ndash1254 2007

[26] N Rahman S K Manirul Haque S N H Azmi andH Rahman ldquoOptimized and validated spectrophotometricmethods for the determination of amiodarone hydrochloridein commercial dosage forms using N-bromosuccinimide andbromothymol bluerdquo Journal of Saudi Chemical Society vol 21no 1 pp 25ndash34 2017

[27] D Taskın G Erensoy and S Sungur ldquoOptimized and vali-dated spectrophotometric determination of butamirate citratein bulk and dosage forms using ion-pair formation withmethyl orange and bromothymol bluerdquo Farmacia vol 65pp 761ndash765 2017

[28] P Govardhan Reddy V Kiran Kumar V Appala RajuJ Raghu Ram and N Appala Rraju ldquoNovel spectrophoto-metric method development for the estimation of boceprevirin bulk and in pharmaceutical formulationsrdquo ResearchJournal of Pharmacy and Technology vol 10 p 4313 2017

[29] A Sakur and S Affas ldquoDirect spectrophotometric determinationof sildenafil citrate in pharmaceutical preparations via complexformation with two sulphonphthalein acid dyesrdquo ResearchJournal of Pharmacy and Technology vol 10 p 1191 2017

[30] K N Prashanth K Basavaiah and K B Vinay ldquoSensitive andselective spectrophotometric assay of rizatriptan benzoate inpharmaceuticals using three sulphonphthalein dyesrdquo ArabianJournal of Chemistry vol 9 pp S971ndashS980 2016

[31] T D Nguyen L Bau L Q +ao and N Dang Dat ldquoEx-tractive spectrophotometric methods for determination ofciprofloxacin in pharmaceutical formulations using sulfo-nephthalein acid dyesrdquo Vietnam journal of chemistry vol 55no 6 pp 767ndash774 2017

[32] H T S BrittonHydrogen Ions Chapman amp Hall 4th edition1952

[33] I T Q (R1) Validation of Analytical Procedures Text andMethodology (CPMPICH28195) ICH Secretariat GenevaSwitzerland 2010

[34] A M El-Brashy M E-S Metwally and F A El-SepaildquoSpectrophotometric determination of some fluoroquinoloneantibacterials by ion-pair complex formation with cobalt (II)tetrathiocyanaterdquo Journal of the Chinese Chemical Societyvol 52 no 1 pp 77ndash84 2005

[35] C S P Sastry K R Rao and D S Prasad ldquoExtractivespectrophotometric determination of some fluoroquinolonederivatives in pure and dosage formsrdquo Talanta vol 42 no 3pp 311ndash316 1995

[36] A M El-Brashy M El-Sayed Metwally and F A El-SepaildquoSpectrophotometric determination of some fluoroquinoloneantibacterials by binary complex formation with xanthenedyesrdquo Farmaco vol 59 no 10 pp 809ndash817 2004

[37] B G Gowda and J Seetharamappa ldquoExtractive spectro-photometric determination of fluoroquinolones and anti-allergic drugs in pure and pharmaceutical formulationsrdquoAnalytical Sciences vol 19 no 3 pp 461ndash464 2003

[38] K N Prashanth K Basavaiah and M S Raghu ldquoSimple andselective spectrophotometric determination of ofloxacin inpharmaceutical formulations using two sulphonphthalein aciddyesrdquo ISRN Spectroscopy Article ID 357598 9 pages 2013


TribologyAdvances in

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Advances inPhysical Chemistry

Hindawiwwwhindawicom

Analytical Methods in Chemistry

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Volume 2018

Bioinorganic Chemistry and ApplicationsHindawiwwwhindawicom Volume 2018

SpectroscopyInternational Journal of


Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom

The Scientific World Journal

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Medicinal ChemistryInternational Journal of


NanotechnologyHindawiwwwhindawicom Volume 2018

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Applied ChemistryJournal of



Biochemistry Research International


Enzyme Research


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SpectroscopyAnalytical ChemistryInternational Journal of


MaterialsJournal of



BioMed Research International Electrochemistry

International Journal of


Na

nom

ate

ria

ls


Journal ofNanomaterials

Submit your manuscripts atwwwhindawicom

pharmaceutical analysis involving quality control of com-mercialized product and pharmacodynamic studies Spec-trophotometric methods for the determination offluoroquinolones could be classified according to the dif-ferent reactions (i) charge-transfer complexation based onthe reaction of FQs as electron donors with p-acceptorssuch as 23-dichloro-56-dicyano-q-benzoquinone 7788-tetracyanoquinodimethane q-chloranil q-nitrophenoland tetracyanoethylene [7 14ndash16] (ii) oxidative couplingreaction using oxidative coupling with 3-methyl-2-benzothiazolinonehydrazone hydrochloride and cerium(IV) ammonium sulfate Fe(III)-MBTH tris(o-phenan-throline) iron(II) and tris (bipyridyl) iron(II) [17 18] (iii)ion-pair complex formation with acid-dye reagents such asSudan III methyl orange supracene violet 3B tropaeolin000 bromophenol blue bromothymol blue bromocresolgreen and bromocresol purple [8 14 19 20]+esemethodswere related with some major drawbacks such as havingnarrow linearity range requiring heating and close pHcontrol long time for the reaction to complete and lowstability of the colored product formed

Bromothymol blue (BTB) (Figure 1(d)) is an anionic dyeand that can be protonated or deprotonated to form yellowor blue respectively +e BTB was used to make ion-paircomplex which was applied to determine many pharma-ceutical compounds by extractive spectrophotometricmethods [21ndash30] However the ion-pair complex betweenBTB and FQs has not been studied +e method based onion-pair complexes between analytes and BTB into a suitableorganic solvent is also simple fast and cheap

In the previous study we used sulphonphthalein acidincluding bromophenol blue bromocresol green and bro-mothymol blue to determine ciprofloxacin pharmaceuticalformulations and achieved good results [31]

In this paper for the first time we investigated extractivespectrophotometric method based on the formation of ion-pair complexes between ciprofloxacin levofloxacin and

ofloxacin with BTB subsequent extraction into chloroformSome effective conditions on the formation of complexessuch as pH shaking time organic solvent and the con-centration of dye were systematically studied +e presentmethod was also applied to determine FQs in some phar-maceutical formulations including tablets and infusions

2 Experimental

21 Apparatus A double beam UV-visible spectropho-tometer (SP-60 Biochrom Ltd UK) with 10 cm of pathlength quartz cells was used to measure all sample absor-bances Inolab pH-meter instrument (Germany) was used tomonitor the pH of solutions +ree standard buffers wereused to calibrate the electrode before measuring pH ofsolutions All measurements were conducted at 25 plusmn 2degCcontrolled by air conditional laboratory

22 Materials and Reagents All chemicals used were ofanalytical grade and double-distilled water was used toprepare all solutions in the present study

FQs were purchased from Sigma (Germany with puritygt990) whereas bromothymol blue (BTB) was supplied byMaya-R China with purity gt99 +e organic solventsincluding chloroform dichloromethane carbon tetrachlo-ride dichloroethane benzene toluene and other chemicalsare analytical reagents (Merck Germany)

+e following dosage forms containing FQs were pur-chased from local pharmacy market and employed in thestudy Hasancip and Kacipro tablets equivalent to 500mgciprofloxacin (Hasan-Dermapharm and Dong Nammanufacturing-Trading pharmaceutical Co Ltd Vietnam)Ciprofloxacin infusion equivalent to 200mg cipro-floxacin100ml solution for infusion (Hebei TianchengPharmaceutical Co Ltd and Shandong Hualu Pharma-ceutical Co Ltd China) Stada and DHG tablets equivalent

HN

N

F

N

O O

OH

(a)

H3C

OH

OO

ONCH3

F

N N

(b)

N

F

N

O

O O

NH3C CH3

OH

(c)

CH3CH3

C3H7C3H7

SO2

OHHO

BrBr OC

(d)

Figure 1 Chemical structures of ciprofloxacin (a) levofloxacin (b) ofloxacin (OFX) (c) and bromothymol blue (d)














1 2 3 4 5 600

02

04

06

08

10

Abs

orba

nce

Solvent type











0 1 2 3 4 500

02

04

06

08

10

Abs

orba

nce

Shaking time (min)



1 2 3 4 5 6 700

02

04

06

08

10

Abs

orba

nce

pH



0 1 2 3 4 5 6 700

02

04

06

08

10

Abs

orba

nce

Volume of dye (ml)





AAm

1minus AAm( 11138571113858 1113859n+2

CM(n)n

(1)


Kf AAm

1minusAAm1113858 1113859n+2

CM(n)n (2)









added1113890 1113891 times 100 (3)





00 02 04 06 08 1000

02

04

06

08

10

Abs

orba

nce

(Drug)(drug+BTB)




360 400 440 480 52000

02

04

06

08

10

Abs

orba

nce

Wavelength (nm)

1

4

2

3



CH3

O

BrCH3

H+

C

HO

Br

C3H7

CH3 CH3SO3H

Br

O

C

HO

Br

C3H7 C3H7C3H7

+

F

NNH

N

O

O

O

OH

+

C3H7

CH3SO2

C3H7HO OH

OC BrBrCH3





(Quinoid ring)

SO3ndash

+

C3H7 C3H7O

Br

HO

Br CCH3 CH3

SO3ndash


CH3 CH3

+

O

OH

O

NNONH

+

F

C3H7 C3H7HO O

Br+ H+

Br CCH3 CH3

SO3H

Br

OC3H7 C3H7

HO

CBr

C3H7

CH3CH3 SO2

OC

C3H7OHHO

BrBr

SO3ndash



HO

C

O

BrBr

C3H7 C3H7

CH3 CH3SO3

ndash


Br CCH3 CH3

C3H7 C3H7

F

N N

O

O

O

OHHO O

Br

NHSO3

+ ndash

F

NO

N

O O

OHHO

C

O

BrBr

NH

C3H7 C3H7

CH3 CH3SO3

ndash+





0 10 20 30 4000

04

08

12

16

20


Abs

orba

nce













1 025 225 0202 01111 07834 03116 502047802 47007 6082 05 2 0272 02500 07071 01486 1570489127 519603 075 175 0336 04286 06955 00512 5725077789 575784 1 15 0419 06667 07631 00035 95623298320 698065 125 125 0476 10000 10000 00000 _ _6 15 1 0432 15000 18371 00002 594141776247 777397 175 075 0356 23333 72213 00026 11722461806 60690

Levofloxacin1 025 225 0137 01111 07834 05749 147898587 41700 6042 05 2 0313 02500 07071 01856 1159613839 506433 075 175 0432 04286 06955 00426 7085741920 585044 1 15 0559 06667 07631 00005 677452456475 783095 125 125 0594 10000 10000 00000 _ _6 15 1 053 15000 18371 00004 341653128945 753367 175 075 0426 23333 72213 00042 6828970665 58344







4 Conclusions




Ofloxacin500 511 10219 231 221000 1026 10264 134 261500 1489 9925 088 minus073






1021 019 186 993 024 242199 057 029 196 076 039


1016 015 148 1019 021 206498 061 012 501 085 017


985 018 183 1012 025 247502 064 013 497 092 019







(μgmL)


Remarks Reference

Ciprofloxacin











no pH adjustment

+isstudy

Levofloxacin





no pH-adjustment

+isstudy

Ofloxacin









pH-adjustment

+isstudy



Hasanciptablet

Kaciprotablet


StadaLevofloxacin











Data Availability




Acknowledgments


References















































Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls
















1 2 3 4 5 600

02

04

06

08

10

Abs

orba

nce

Solvent type











0 1 2 3 4 500

02

04

06

08

10

Abs

orba

nce

Shaking time (min)



1 2 3 4 5 6 700

02

04

06

08

10

Abs

orba

nce

pH



0 1 2 3 4 5 6 700

02

04

06

08

10

Abs

orba

nce

Volume of dye (ml)





AAm

1minus AAm( 11138571113858 1113859n+2

CM(n)n

(1)


Kf AAm

1minusAAm1113858 1113859n+2

CM(n)n (2)









added1113890 1113891 times 100 (3)





00 02 04 06 08 1000

02

04

06

08

10

Abs

orba

nce

(Drug)(drug+BTB)




360 400 440 480 52000

02

04

06

08

10

Abs

orba

nce

Wavelength (nm)

1

4

2

3



CH3

O

BrCH3

H+

C

HO

Br

C3H7

CH3 CH3SO3H

Br

O

C

HO

Br

C3H7 C3H7C3H7

+

F

NNH

N

O

O

O

OH

+

C3H7

CH3SO2

C3H7HO OH

OC BrBrCH3





(Quinoid ring)

SO3ndash

+

C3H7 C3H7O

Br

HO

Br CCH3 CH3

SO3ndash


CH3 CH3

+

O

OH

O

NNONH

+

F

C3H7 C3H7HO O

Br+ H+

Br CCH3 CH3

SO3H

Br

OC3H7 C3H7

HO

CBr

C3H7

CH3CH3 SO2

OC

C3H7OHHO

BrBr

SO3ndash



HO

C

O

BrBr

C3H7 C3H7

CH3 CH3SO3

ndash


Br CCH3 CH3

C3H7 C3H7

F

N N

O

O

O

OHHO O

Br

NHSO3

+ ndash

F

NO

N

O O

OHHO

C

O

BrBr

NH

C3H7 C3H7

CH3 CH3SO3

ndash+





0 10 20 30 4000

04

08

12

16

20


Abs

orba

nce













1 025 225 0202 01111 07834 03116 502047802 47007 6082 05 2 0272 02500 07071 01486 1570489127 519603 075 175 0336 04286 06955 00512 5725077789 575784 1 15 0419 06667 07631 00035 95623298320 698065 125 125 0476 10000 10000 00000 _ _6 15 1 0432 15000 18371 00002 594141776247 777397 175 075 0356 23333 72213 00026 11722461806 60690

Levofloxacin1 025 225 0137 01111 07834 05749 147898587 41700 6042 05 2 0313 02500 07071 01856 1159613839 506433 075 175 0432 04286 06955 00426 7085741920 585044 1 15 0559 06667 07631 00005 677452456475 783095 125 125 0594 10000 10000 00000 _ _6 15 1 053 15000 18371 00004 341653128945 753367 175 075 0426 23333 72213 00042 6828970665 58344







4 Conclusions




Ofloxacin500 511 10219 231 221000 1026 10264 134 261500 1489 9925 088 minus073






1021 019 186 993 024 242199 057 029 196 076 039


1016 015 148 1019 021 206498 061 012 501 085 017


985 018 183 1012 025 247502 064 013 497 092 019







(μgmL)


Remarks Reference

Ciprofloxacin











no pH adjustment

+isstudy

Levofloxacin





no pH-adjustment

+isstudy

Ofloxacin









pH-adjustment

+isstudy



Hasanciptablet

Kaciprotablet


StadaLevofloxacin











Data Availability




Acknowledgments


References















































Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls










0 1 2 3 4 500

02

04

06

08

10

Abs

orba

nce

Shaking time (min)



1 2 3 4 5 6 700

02

04

06

08

10

Abs

orba

nce

pH



0 1 2 3 4 5 6 700

02

04

06

08

10

Abs

orba

nce

Volume of dye (ml)





AAm

1minus AAm( 11138571113858 1113859n+2

CM(n)n

(1)


Kf AAm

1minusAAm1113858 1113859n+2

CM(n)n (2)









added1113890 1113891 times 100 (3)





00 02 04 06 08 1000

02

04

06

08

10

Abs

orba

nce

(Drug)(drug+BTB)




360 400 440 480 52000

02

04

06

08

10

Abs

orba

nce

Wavelength (nm)

1

4

2

3



CH3

O

BrCH3

H+

C

HO

Br

C3H7

CH3 CH3SO3H

Br

O

C

HO

Br

C3H7 C3H7C3H7

+

F

NNH

N

O

O

O

OH

+

C3H7

CH3SO2

C3H7HO OH

OC BrBrCH3





(Quinoid ring)

SO3ndash

+

C3H7 C3H7O

Br

HO

Br CCH3 CH3

SO3ndash


CH3 CH3

+

O

OH

O

NNONH

+

F

C3H7 C3H7HO O

Br+ H+

Br CCH3 CH3

SO3H

Br

OC3H7 C3H7

HO

CBr

C3H7

CH3CH3 SO2

OC

C3H7OHHO

BrBr

SO3ndash



HO

C

O

BrBr

C3H7 C3H7

CH3 CH3SO3

ndash


Br CCH3 CH3

C3H7 C3H7

F

N N

O

O

O

OHHO O

Br

NHSO3

+ ndash

F

NO

N

O O

OHHO

C

O

BrBr

NH

C3H7 C3H7

CH3 CH3SO3

ndash+





0 10 20 30 4000

04

08

12

16

20


Abs

orba

nce













1 025 225 0202 01111 07834 03116 502047802 47007 6082 05 2 0272 02500 07071 01486 1570489127 519603 075 175 0336 04286 06955 00512 5725077789 575784 1 15 0419 06667 07631 00035 95623298320 698065 125 125 0476 10000 10000 00000 _ _6 15 1 0432 15000 18371 00002 594141776247 777397 175 075 0356 23333 72213 00026 11722461806 60690

Levofloxacin1 025 225 0137 01111 07834 05749 147898587 41700 6042 05 2 0313 02500 07071 01856 1159613839 506433 075 175 0432 04286 06955 00426 7085741920 585044 1 15 0559 06667 07631 00005 677452456475 783095 125 125 0594 10000 10000 00000 _ _6 15 1 053 15000 18371 00004 341653128945 753367 175 075 0426 23333 72213 00042 6828970665 58344







4 Conclusions




Ofloxacin500 511 10219 231 221000 1026 10264 134 261500 1489 9925 088 minus073






1021 019 186 993 024 242199 057 029 196 076 039


1016 015 148 1019 021 206498 061 012 501 085 017


985 018 183 1012 025 247502 064 013 497 092 019







(μgmL)


Remarks Reference

Ciprofloxacin











no pH adjustment

+isstudy

Levofloxacin





no pH-adjustment

+isstudy

Ofloxacin









pH-adjustment

+isstudy



Hasanciptablet

Kaciprotablet


StadaLevofloxacin











Data Availability




Acknowledgments


References















































Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls





AAm

1minus AAm( 11138571113858 1113859n+2

CM(n)n

(1)


Kf AAm

1minusAAm1113858 1113859n+2

CM(n)n (2)









added1113890 1113891 times 100 (3)





00 02 04 06 08 1000

02

04

06

08

10

Abs

orba

nce

(Drug)(drug+BTB)




360 400 440 480 52000

02

04

06

08

10

Abs

orba

nce

Wavelength (nm)

1

4

2

3



CH3

O

BrCH3

H+

C

HO

Br

C3H7

CH3 CH3SO3H

Br

O

C

HO

Br

C3H7 C3H7C3H7

+

F

NNH

N

O

O

O

OH

+

C3H7

CH3SO2

C3H7HO OH

OC BrBrCH3





(Quinoid ring)

SO3ndash

+

C3H7 C3H7O

Br

HO

Br CCH3 CH3

SO3ndash


CH3 CH3

+

O

OH

O

NNONH

+

F

C3H7 C3H7HO O

Br+ H+

Br CCH3 CH3

SO3H

Br

OC3H7 C3H7

HO

CBr

C3H7

CH3CH3 SO2

OC

C3H7OHHO

BrBr

SO3ndash



HO

C

O

BrBr

C3H7 C3H7

CH3 CH3SO3

ndash


Br CCH3 CH3

C3H7 C3H7

F

N N

O

O

O

OHHO O

Br

NHSO3

+ ndash

F

NO

N

O O

OHHO

C

O

BrBr

NH

C3H7 C3H7

CH3 CH3SO3

ndash+





0 10 20 30 4000

04

08

12

16

20


Abs

orba

nce













1 025 225 0202 01111 07834 03116 502047802 47007 6082 05 2 0272 02500 07071 01486 1570489127 519603 075 175 0336 04286 06955 00512 5725077789 575784 1 15 0419 06667 07631 00035 95623298320 698065 125 125 0476 10000 10000 00000 _ _6 15 1 0432 15000 18371 00002 594141776247 777397 175 075 0356 23333 72213 00026 11722461806 60690

Levofloxacin1 025 225 0137 01111 07834 05749 147898587 41700 6042 05 2 0313 02500 07071 01856 1159613839 506433 075 175 0432 04286 06955 00426 7085741920 585044 1 15 0559 06667 07631 00005 677452456475 783095 125 125 0594 10000 10000 00000 _ _6 15 1 053 15000 18371 00004 341653128945 753367 175 075 0426 23333 72213 00042 6828970665 58344







4 Conclusions




Ofloxacin500 511 10219 231 221000 1026 10264 134 261500 1489 9925 088 minus073






1021 019 186 993 024 242199 057 029 196 076 039


1016 015 148 1019 021 206498 061 012 501 085 017


985 018 183 1012 025 247502 064 013 497 092 019







(μgmL)


Remarks Reference

Ciprofloxacin











no pH adjustment

+isstudy

Levofloxacin





no pH-adjustment

+isstudy

Ofloxacin









pH-adjustment

+isstudy



Hasanciptablet

Kaciprotablet


StadaLevofloxacin











Data Availability




Acknowledgments


References















































Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls




360 400 440 480 52000

02

04

06

08

10

Abs

orba

nce

Wavelength (nm)

1

4

2

3



CH3

O

BrCH3

H+

C

HO

Br

C3H7

CH3 CH3SO3H

Br

O

C

HO

Br

C3H7 C3H7C3H7

+

F

NNH

N

O

O

O

OH

+

C3H7

CH3SO2

C3H7HO OH

OC BrBrCH3





(Quinoid ring)

SO3ndash

+

C3H7 C3H7O

Br

HO

Br CCH3 CH3

SO3ndash


CH3 CH3

+

O

OH

O

NNONH

+

F

C3H7 C3H7HO O

Br+ H+

Br CCH3 CH3

SO3H

Br

OC3H7 C3H7

HO

CBr

C3H7

CH3CH3 SO2

OC

C3H7OHHO

BrBr

SO3ndash



HO

C

O

BrBr

C3H7 C3H7

CH3 CH3SO3

ndash


Br CCH3 CH3

C3H7 C3H7

F

N N

O

O

O

OHHO O

Br

NHSO3

+ ndash

F

NO

N

O O

OHHO

C

O

BrBr

NH

C3H7 C3H7

CH3 CH3SO3

ndash+





0 10 20 30 4000

04

08

12

16

20


Abs

orba

nce













1 025 225 0202 01111 07834 03116 502047802 47007 6082 05 2 0272 02500 07071 01486 1570489127 519603 075 175 0336 04286 06955 00512 5725077789 575784 1 15 0419 06667 07631 00035 95623298320 698065 125 125 0476 10000 10000 00000 _ _6 15 1 0432 15000 18371 00002 594141776247 777397 175 075 0356 23333 72213 00026 11722461806 60690

Levofloxacin1 025 225 0137 01111 07834 05749 147898587 41700 6042 05 2 0313 02500 07071 01856 1159613839 506433 075 175 0432 04286 06955 00426 7085741920 585044 1 15 0559 06667 07631 00005 677452456475 783095 125 125 0594 10000 10000 00000 _ _6 15 1 053 15000 18371 00004 341653128945 753367 175 075 0426 23333 72213 00042 6828970665 58344







4 Conclusions




Ofloxacin500 511 10219 231 221000 1026 10264 134 261500 1489 9925 088 minus073






1021 019 186 993 024 242199 057 029 196 076 039


1016 015 148 1019 021 206498 061 012 501 085 017


985 018 183 1012 025 247502 064 013 497 092 019







(μgmL)


Remarks Reference

Ciprofloxacin











no pH adjustment

+isstudy

Levofloxacin





no pH-adjustment

+isstudy

Ofloxacin









pH-adjustment

+isstudy



Hasanciptablet

Kaciprotablet


StadaLevofloxacin











Data Availability




Acknowledgments


References















































Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls






0 10 20 30 4000

04

08

12

16

20


Abs

orba

nce













1 025 225 0202 01111 07834 03116 502047802 47007 6082 05 2 0272 02500 07071 01486 1570489127 519603 075 175 0336 04286 06955 00512 5725077789 575784 1 15 0419 06667 07631 00035 95623298320 698065 125 125 0476 10000 10000 00000 _ _6 15 1 0432 15000 18371 00002 594141776247 777397 175 075 0356 23333 72213 00026 11722461806 60690

Levofloxacin1 025 225 0137 01111 07834 05749 147898587 41700 6042 05 2 0313 02500 07071 01856 1159613839 506433 075 175 0432 04286 06955 00426 7085741920 585044 1 15 0559 06667 07631 00005 677452456475 783095 125 125 0594 10000 10000 00000 _ _6 15 1 053 15000 18371 00004 341653128945 753367 175 075 0426 23333 72213 00042 6828970665 58344







4 Conclusions




Ofloxacin500 511 10219 231 221000 1026 10264 134 261500 1489 9925 088 minus073






1021 019 186 993 024 242199 057 029 196 076 039


1016 015 148 1019 021 206498 061 012 501 085 017


985 018 183 1012 025 247502 064 013 497 092 019







(μgmL)


Remarks Reference

Ciprofloxacin











no pH adjustment

+isstudy

Levofloxacin





no pH-adjustment

+isstudy

Ofloxacin









pH-adjustment

+isstudy



Hasanciptablet

Kaciprotablet


StadaLevofloxacin











Data Availability




Acknowledgments


References















































Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls








4 Conclusions




Ofloxacin500 511 10219 231 221000 1026 10264 134 261500 1489 9925 088 minus073






1021 019 186 993 024 242199 057 029 196 076 039


1016 015 148 1019 021 206498 061 012 501 085 017


985 018 183 1012 025 247502 064 013 497 092 019







(μgmL)


Remarks Reference

Ciprofloxacin











no pH adjustment

+isstudy

Levofloxacin





no pH-adjustment

+isstudy

Ofloxacin









pH-adjustment

+isstudy



Hasanciptablet

Kaciprotablet


StadaLevofloxacin











Data Availability




Acknowledgments


References















































Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls







(μgmL)


Remarks Reference

Ciprofloxacin











no pH adjustment

+isstudy

Levofloxacin





no pH-adjustment

+isstudy

Ofloxacin









pH-adjustment

+isstudy



Hasanciptablet

Kaciprotablet


StadaLevofloxacin











Data Availability




Acknowledgments


References















































Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls





Data Availability




Acknowledgments


References















































Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls

























Journal of

Chemistry





Journal of

Volume 2018






Volume 2018




Journal of






Enzyme Research


Journal of



MaterialsJournal of






Na

nom

ate

ria

ls




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