CHAPTER 10
DARUNAVIR
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10.1. DRUG PROFILE
Darunavir (brand name Prezista, formerly known as TMC114) is in the protease inhibitor
class used to treat human immunodeficiency virus (HIV) in adults and children 6 years of age
and older. It was approved by the Food and Drug Administration (FDA) on June 23, 2006.[1]
To
overcome the problems with early protease inhibitor (PIs) like severe side effects and drug
toxicities, require a high therapeutic dose, are costly to manufacture, and show a disturbing
susceptibility to drug resistant mutations a second generation protease inhibitor (PIs) Darunavir
is discovered.
Darunavir is used with ritonavir (Norvir) and other medications to treat human
immunodeficiency virus (HIV). It works by slowing the spread of HIV in the body. Darunavir
does not cure HIV infection and may not prevent you from developing HIV-related illnesses.
Darunavir does not prevent you from spreading HIV to other people.
Figure 10.A: Darunavir
Systematic (IUPAC) name : (1R, 5S, 6R)-2, 8-dioxabicyclo [3.3.0] oct-6-yl] N
[(2S, 3R)-4- [(4-aminophenyl) sulfonyl- (2 methylpropyl)
amino]-3-hydroxy-1-phenyl- butan-2 yl] carbamate
Formula : C27H37N3O7S
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Mol. Mass : 547.665 g/mol
Routes : Oral
Darunavir was designed to form robust interactions with the protease enzyme from many
strains of HIV, including strains from treatment-experienced patients with multiple resistance
mutations to PIs.[2][3]
It blocks HIV protease, an enzyme which is needed for HIV to multiply.
When used with other anti-HIV medicines. It may reduce the amount of HIV in your blood
(called "viral load") and increase your CD4 (T) cell count. HIV infection destroys CD4 (T) cells,
which are important to the immune system. The immune system helps fight infection. Reducing
the amount of HIV and increasing the CD4 (T) cell count may improve your immune system
and, thus, reduce the risk of death or infections that can happen when your immune system is
weak (opportunistic infections).
List of brand names of Darunavir
Table: 10.1
S.NO. BRAND
NAME
FORMULATION COMBINATION AVAILABLE
STRENGTH
Mg
MANUFACTURER
1 PREZISTA Tab Ritonavir 75 JOLLC
2 PREZISTA Tab Ritonavir 150
3 PREZISTA Tab Ritonavir 400
4 PREZISTA Tab Ritonavir 600
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10.2. LITREATURE SURVEY
Several analytical methods have been reported for the determination of Darunavir in pure
drug, pharmaceutical dosage forms and in biological samples using spcetrophotometry liquid
chromatography, electro kinetic chromatography high performance thin layer chromatography
either in single or in combined forms.
Raveendra B.Ganduri et al [4]
has develop a simple, rapid, sensitive, accurate, precise and
reproducible high performance liquid chromatographic method for the determination of
Darunavir in tablet dosage form. Waters, Symmetry shield RP18 (250X4.6mm, 5µm)
column0.1% orthophosphoric acid and acetonitrile (50:50 % v/v) as mobile phase, detection
wavelength of 265 nm, flow rate of 1.0 mLmin-1. The method is linear from 25µg mL-1 to
100µg mL-1, accuracy was found to be 99.54%, mean inter and intraday assay relative standard
deviation (RSD) were less than 1.0%. The method is simple, accurate, specific and precise, can
be used for the determination of Darunavir.
Masaaki Takahashi et al [5]
has described a HPLC method to validate the determination
of plasma DRV concentrations, a simple procedure for simultaneous determination of seven HIV
protease inhibitors and efavirenz. The calibration curve was linear (range of 0.13 to 10.36
μg/ml). The average accuracy ranged from 100.7 to 105.6%. Both the interday and intraday
coefficients of variation were less than 6.7%, which was similar to or much lower than
previously reported values by the LC/MS/MS method. It is concluded that HPLC can be used to
determine plasma DRV concentrations and routinely in the clinical setting; thus, this HPLC
method enables further study of DRV pharmacokinetics in conventional hospital laboratories
Lauriane Goldwirt et al [6]
has developed and validated a precise and accurate high-
performance liquid chromatography (HPLC) method with UV detection for Darunavir, a peptidic
protease inhibitor. An internal standard, methylclonazepam, was added to 100 μL of plasma
before a solid-phase extraction on C18 Bond Elute column. The eluted solutions were evaporated
to dryness and reconstituted with 100 μL of mobile phase before being injected in the
chromatographic system. The separation was performed on a C8 column using an acetonitrile
270
and ultrapure water mixture (40:60, v/v) as mobile phase. All compounds were detected at a
wavelength of 266 nm. The method was linear and validated over a concentration range of 0.25–
20 mg/L. The within-day precision, ranged from 3.0 to 7.9%, while the within-day accuracy
ranged from −11.4 to 0.5%. The between day precision and accuracy were respectively less than
13.7 and −11.4%. The mean recovery was 75.7% for Darunavir and 66.7% for
methylclonazepam. This method provides a useful tool for therapeutic drug monitoring in HIV
patients.
Ivy Song et al [7]
has conducted a pharmacokinetic work to determine the Effect of
Lopinavir/Ritonavir and Darunavir/Ritonavir on the HIV Integrase Inhibitor S/GSK1349572 in
Healthy Participants. S/GSK1349572 is an unboosted, once-daily integrase inhibitor with a novel
resistance profile. As standard of care for patients infected with HIV is combination therapy, the
potential interaction between S/GSK1349572 and ritonavir-boosted protease inhibitors was
evaluated. In an open-label, repeat-dose, 2-period, 2-sequence crossover study in healthy
participants, S/GSK1349572 was administered at 30 mg once daily for 5 days, followed by
randomization to lopinavir/ritonavir 400/100 mg twice daily or Darunavir/ritonavir 600/100 mg
twice daily co administered with S/GSK1349572 30 mg once daily for 14 days. There was no
washout between periods. Serial pharmacokinetic (PK) samples and safety assessments were
obtained throughout the study. Thirty of 31 participants completed the study (15 participants per
group). Treatment comparisons of steady-state S/GSK1349572 PK parameters demonstrated that
co administration of lopinavir/ritonavir had no significant effect on steady-state PK of
S/GSK1349572. Coadministration of Darunavir/ritonavir resulted in a nonclinically significant
reduction in steady-state plasma S/GSK1349572 exposures. Plasma S/GSK1349572 AUC (0-τ),
Cmax, and Cτ decreased by 22%, 11%, and 38%, respectively, on average. S/GSK1349572 was
well tolerated with no serious adverse events (AEs) or withdrawals due to drug-related AEs. The
most frequent drug-related AEs were diarrhea, dizziness, and headache. No dosage adjustment
for S/GSK1349572 is required when used with lopinavir/ritonavir or Darunavir/ritonavir.
D'Avolio A et al[8]
has developed and validated a new method using high performance
liquid chromatography coupled with electrospray mass spectrometry (HPLC-MS) for the
quantification of plasma concentration of the new protease inhibitors Darunavir (DRV) and other
11 antiretroviral agents (ritonavir, amprenavir, atazanavir, lopinavir, saquinavir, indinavir,
271
nelfinavir and its metabolite M-8, nevirapine, efavirenz and tipranavir). A simple protein
precipitation extraction procedure was applied on 50 microl of plasma aliquots and
chromatographic separation of drugs and Internal Standard (quinoxaline) was achieved with a
gradient (acetonitrile and water with formic acid 0.05%) on a C-18 reverse phase analytical
column with 25 min of analytical run. Calibration curves were optimized according to expected
ranges of drug concentrations in patients, and correlation coefficient (r2) was higher than 0.998
for all analytes. Mean intra- and inter-day precision (relative standard deviation %) for all
compounds were 8.4 and 8.3%, respectively, and mean accuracy (% of deviation from nominal
level) was 3.9%. Extraction recovery ranged within 93 and 105% for all drugs analyzed. This
novel HPLC-MS methodology allows a specific, sensitive and reliable determination of DRV
and 11 other antiretrovirals. In our hand, it was used to measure DRV and ritonavir plasma
concentration in HIV-positive patients, and it is now successfully applied for routine therapeutic
drug monitoring and pharmacokinetics studies.
Bhavini N. Patel et al
[9] has developed and validated a quantitative high-performance
thin-layer chromatography (HPTLC) method for determination of Darunavir ethanolate (DRV)
in tablets. DRV from the formulations was separated and identified on silica gel 60 F254 HPTLC
plates with toluene-ethyl acetate-methanol 7.0:2.0:1.0 (ν/ν) as mobile phase. The plates were
developed to a distance of 8 cm. Quantification was performed at λ = 267 nm. Well-resolved
bands were obtained for DRV. The method was validated for specificity, precision, robustness,
and accuracy. The calibration plot for DRV standard was linear in the range 250–1750 ng per
band with r = 0.9994, slope = 0.4253, and intercept = 44.81. The limits of detection and
quantification were 15.28 and 45.84 ng per band, respectively. The method is selective, sensitive,
and specific, with potential application in pharmaceutical analysis.
Jose Molto et al[10]
has described a Open-label, randomized pilot study in HIV-infected
patients on Darunavir/ritonavir 600/100 mg twice daily (viral load <50 copies/ml; Darunavir vIQ
>2). Thirty patients were randomized to Darunavir/ritonavir 900/100 mg once daily (once-daily
group, n=15) or 600/100 mg twice daily (twice-daily group, n=15). Viral load, blood chemistry,
and Darunavir and ritonavir trough plasma concentrations (Ctrough) were determined up to 48
weeks. If the Darunavir vIQ fell to <1.5, the dosage was switched to 600/100 mg twice daily.
The primary end point was the percentage of 48-week treatment failure. Patients had taken a
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mean 11.6 (SD ±3.9) antiretroviral regimens before Darunavir/ritonavir administration. The
proportion of patients without 48-week treatment failure was 86.7% in both groups. The median
(interquartile range [IQR]) Darunavir Ctrough decreased from 3.09 mg/l (IQR 2.43–3.93) at
baseline to 1.60 mg/l (IQR 1.25–2.04) at week 48 (P=0.001) in the once-daily group. Three
once-daily group patients switched to Darunavir/ritonavir 600/100 mg twice daily. Fewer
patients had triglyceride levels >200 mg/dl at week 48 in the once-daily group (20.0%) than in
the twice-daily group (20.0% versus 57.1%; P=0.046). Treatment simplification to Darunavir/-
ritonavir 900/100 mg once daily guided by the Darunavir vIQ in treatment-experienced HIV-
infected patients receiving Darunavir/ritonavir 600/100 mg twice-daily seems to be safe enough
to be tested in adequately powered clinical trials.
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10.3. EXPERIMENTAL
10.3.1. Instrumentation
Peak HPLC containing LC 20AT pump and variable wavelength programmable UV-
Visible detector and Rheodyne injector was employed for investigation. The chromatographic
analysis was performed on a Chromosil C18 column (250 mm × 4.6 mm, 5µm). Degassing of the
mobile phase was done using a Loba ultrasonic bath sonicator. A Denwar Analytical balance was
used for weighing the materials.
10.3.2. Chemicals and Solvents
The reference sample of Darunavir (API) was obtained from Jollc. The Formulation
PREZISTA (Darunavir) was purchased from the local market. Methanol, Water used were of
HPLC grade and purchased from Merck Specialties Private Limited, Mumbai, India.
10.3.3. The mobile phase
A mixture of Acetontrile: Methanol in the ratio of 90:10 v/v was prepared and used as
mobile phase.
10.3.4. Standard solution of the drug
For analysis 100 ppm standard solution was prepared, required concentrations were
obtained from 100 ppm solution by appropriate dilution.
10.3.5. Sample (Capsule) solution
The formulation tablets of Darunavir (PREZISTA - 75 mg) were crushed to give finely
powdered material. From the Powder prepared a 30 ppm solution with mobile phase and then
filtered through Ultipor N66 Nylon 6, 6 membrane sample filter paper.
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10.4. METHOD DEVELOPMENT
For developing the method (as described in chapter 1 and 2), a systematic study of the
effect of various factors was undertaken by varying one parameter at a time and keeping all other
conditions constant. Method development consists of selecting the appropriate wave length and
choice of stationary and mobile phases. The following studies were conducted for this purpose.
10.4.1. Detection wavelength
The spectrum of 10ppm solution of the Darunavir in methanol was recorded separately
on UV spectrophotometer. The peak of maximum absorbance wavelength was observed. The
spectra of Darunavir were showed maximum absorbance at 271nm.
10.4.2. Choice of stationary phase
Preliminary development trials have performed with octadecyl columns with different
types, configurations and from different manufacturers. Finally the expected separation and peak
shapes were obtained on Chromosil C18 (250 mm x 4.6 mm, 5μm) column.
10.4.3. Selection of the mobile phase
In order to get sharp peak, low tailing factor and base line separation of the separation of
the components, a number of experiments were carried out by varying the composition of
various solvents and flow rate. To have an ideal separation of the drug under isocratic conditions,
mixtures of solvents like methanol, water and Acetonitrile with or without different buffers
indifferent combinations were tested as mobile phases on a Chromosil C18 column. A mixture of
Acetonitrile : Methanol in the ratio of 90:10 v/v was proved to be the most suitable of all the
combinations since the chromatographic peak obtained was better defined and resolved and
almost free from tailing.
10.4.4. Flow rate
Flow rates of the mobile phase were changed from 0.5 – 1.5 mL/min for optimum
separation. A minimum flow rate as well as minimum run time gives the maximum saving on the
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usage of solvents. It was found from the experiments that 1.0 mL/min flow rate was ideal for the
successful elution of the analyte.
10.4.5. Optimized chromatographic conditions
Chromatographic conditions as optimized above were shown in Table: 10.2 these
optimized conditions were followed for the determination of Darunavir in bulk samples and in its
Formulations. The chromatogram of standard (3ppm) shown in Figure 10.B
Mobile phase Acetonitrile : Methanol 90:10 v/v
Pump mode Isocratic
Mobile phase PH 6.2
Diluent Mobile phase
Column Chromosil C18 column (250 mm x 4.6
mm, 5μ)
Column Temp Ambient
Wavelength
271 nm
Injection Volume 20 μl
Flow rate 1.0 mL/min
Run time 5 min
Retention Time 2.59 min
Table 10.2: Optimized chromatographic conditions for estimation Darunavir
276
Figure10.B: Chromatogram of standard solution
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10.5. VALIDATION OF THE PROPOSED METHOD
The proposed method was validated (as described in the chapter 1 and 2) as per ICH
guidelines . The parameters studied for validation were specificity, linearity, precision, accuracy,
robustness, system suitability, limit of detection, limit of quantification, and solution stability.
10.5.1. Specificity
The specificity of method was performed by comparing the chromatograms of blank,
standard and sample (Prepared from Formulation). It was found that there is no interference due
to excipients in the tablet formulation and also found good correlation between the retention
times of standard and sample. The specificity results are shown in Table 10.3
NAME OF THE SOLUTION Retention Time in Min
Blank No peaks
Darunavir (Standerd) 2.59
Darunavir (Sample) 2.56
Table 10.3: Specificity study
10.5.2 Linearity
Linearity was performed by preparing mixed standard solutions of Darunavir at different
concentration levels including working concentration mentioned in experimental condition i.e.
30ppm. Twenty micro liters of each concentration was injected in duplicate into the HPLC
system. The response was read at 271 nm and the corresponding chromatograms were recorded.
From these chromatograms, the mean peak areas were calculated and linearity plots of
concentration over the mean peak areas were constructed individually. The regressions of the
plots were computed by least square regression method. Linearity results were presented in Table
10.4.
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Level Concentration of Darunavir
in ppm
Mean peak area
Level -1 10 66073
Level -2 20 112573.0
Level -3 30 169196.8
Level -4 40 214545.7
Level -5 50 281291.2
Range: 10-30ppm Slope
Intercept
Correlation coefficient
5324.09
9013.21
0.997
Table 10.4: Linearity results
Figure 10.C: On X axis concentration of sample, On Y axis peak area response
10.5.3. Precision Precision is the degree of repeatability of an analytical method under normal Operational
conditions. Precision of the method was performed as intraday precision, Inter day precision.
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10.5.3.1. Intraday precision
To study the intraday precision, six replicate standard solutions (50ppm) of Darunavir
were injected. The percent relative standard deviation (% RSD) was calculated and it was found
to be 1.068, which are well within the acceptable criteria of not more than 2.0. Results of system
precision studies are shown in Table 10.5
SAMPLE
CONC(PPM) INJECTION
no.
PEAKS
AREA
R.S.D(Acceptance
criteria ≤ 2.0%)
Darunavir
50
1 280344.9
1.068
2 281291.2
3 287555
4 286832.8
5 286435.8
6 284917.2
Table 10.5: System Precision (Intra Day)
10.5.3.2. Inter Day precision
To study the interday precision, six replicate standard solution (50ppm) of Darunavir was
injected on third day of sample preparation. The percent relative standard deviation (% RSD)
was calculated and it was found to be 0.560, which are well within the acceptable criteria of not
more than 2.0. Results of system precision studies are shown in Table 10.6
SAMPLE
CONC(PPM) INJECTION
No.
PEAKS AREA R.S.D(Acceptance
criteria ≤ 2.0%)
Darunavir
50
1 288032.7
0.560
2 287629.1
3 290361.9
4 290419.7
5 291211.5
6 287647.5
Table 10.6: System Precision (Inter Day)
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10.5.4. Accuracy
The accuracy of the method was determined by standard addition method. A known
amount of standard drug was added to the fixed amount of pre-analyzed tablet solution. Percent
recovery was calculated by comparing the area before and after the addition of the standard drug.
The standard addition method was performed at 50%, 100% and 150% level of 20ppm. The
solutions were analyzed in triplicate at each level as per the proposed method. The percent
recovery and % RSD was calculated and results are presented in Table 10.7 Satisfactory
recoveries ranging from 99.0 to 102.0 were obtained by the proposed method. This indicates that
the proposed method was accurate.
Level Amount of Darunavir
spiked (ppm)
Amount of Darunavir
recovered(ppm)
% Recovery % RSD
50 %
30 29.95 99.83
0.440 30 29.9 99.66
30 29.7 99.00
100%
40 39.8 99.5
0.318 40 40.02 100.05
40 39.8 99.5
150%
50 49.5 99.0
0.696 50 49.5 99.0
50 50.1 100.2
Mean % of
recovery
99.52
Mean
RSD =
0.484
Table 10.7: Percentage RSD and % Recovery
10.5.5. Robustness
The robustness study was performed by slight modification in flow rate of Mobile phase,
pH of the buffer and composition of the mobile phase. Darunavir at 40 ppm concentration was
analyzed under these changed experimental conditions. It was observed that there were no
marked changes in chromatograms, which demonstrated that the developed method was robust in
nature. The results of robustness study are shown in Table 10.8
281
Condition Mean area % assay % difference
Unaltered 214545.7 100.0 0.0
Flow rate at 0.8 mL/min
Flow rate at 1.2mL/min
215328.6
215124.8
100.36
100.26
0.36
0.26
Mobile phase:
MEOH: Water
75% 25%
85% 15%
213845.5
214853.0
99.67
100.14
0.33
0.14
pH of mobile phase at 7.3 2140537.5 99.77 0.23
pH of mobile phase at 7.7 213452.9 99.49 0.51
Table 10.8: Robustness
10.5.6. System suitability
System suitability was studied under each validation parameters by injecting six
replicates of the standard solution 2 ppm). The results obtained were within acceptable limits
(Tailing factor ≤2 and Theoretical plate’s ≥2000) and are represented in Table 10.9. Thus, the
system meets suitable criteria.
Parameter Tailing factor Theoretical plates
Specificity study 1.23 5069.89
Linearity study 0.95 5368.86
Precision study 1.21 4709.73
Table 10.9: System Suitability
10.5.7. Limit of detection and Limit of quantification
Limit of detection (LOD) is defined as the lowest concentration of analyte that gives a
detectable response. Limit of quantification (LOQ) is defined as the lowest Concentration that
can be quantified reliably with a specified level of accuracy and Precision. For this sample was
dissolved by using Mobile Phase and injected until peak was disappeared. After 0.6ppm dilution,
Peak was not clearly observed. So it confirms that 0.6ppm is limit of Detection and 0.198ppm
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dilution is Limit of Quantification. For this study six replicates of the analyte at lowest
concentration were Measured and quantified. The LOD and LOQ of Darunavir are given in
Table 10.10
parameter Measured volume
Limit of Quantification 0.198ppm
Limit of Detection 0.6ppm
Table 10.10: LOD And LOQ
Formulation:
For assay Darunavir (PREZISTA - 75 mg) 20 tablets were weigh and calculate the
average weight. Accurately weigh and transfer the sample equivalent to 10mg of Darunavir in to
a 10ml volumetric flask. Add diluent and sonicate to dissolve it completely and make volume up
to the mark with diluents. Mix well and filter through 0.45um filter. Further pipette 1ml of the
above stock solution into a 10ml volumetric flask and dilute up to mark with diluents and finally
10 ppm were prepared. Mix well and filter through 0.45um filter. An aliquot of this solution was
injected into HPLC system. Peak area of Darunavir was measured for the determination.
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10.6. RESULTS AND DISCUSSION
Having optimized the efficiency of a chromatographic separation the quality of the
chromatography was monitored by applying the system suitability tests. It was assessed by
replicate analysis of two injections of the drug at a concentration of 30ppm. The acceptance
criterion was ± 2% for the percent coefficient of variation (%CV) for the peak area and retention
times for Darunavir. The number of theoretical plates should not be less than 2500 and the tailing
factor should not be more than 2.0. The peak purity of Darunavir was assessed by comparing the
retention time (Rt) of standard and the sample. Good correlation was obtained between the Rt of
standard and sample. A chromatogram obtained from reference substance solution is presented.
System suitability parameters were shown in Table: 10.9
Linearity was studied by preparing standard solutions at different concentration
levels. The linearity y ranges for Darunavir found to be 10-50ppm. The regression equation for
LP and AT were found to be y = 9013x+5324 with coefficient of correlation, (r) 0.997. The
results of Linearity studies are shown in Table: 10.4 and the linearity curve were shown in
Figure: 10.C. Precision was evaluated by carrying out six independent sample preparation of a
single lot of formulation. The sample solution was prepared in the same manner as described in
sample preparation. Percentage relative standard deviation (%RSD) was ( for Intradey-1.068, For
Interday-0.560) found to be less than 2% for within a day and day to day variations, which
proves that method is precise.
Known amounts of standard Darunavir added to pre-analyzed samples and were
subjected to the proposed HPLC method at 50%, 100% and 150% to evaluate the degree of
accuracy. . Results of recovery studies are shown range 99.00-101.45%. The mean recovery data
obtained for each level as well as for all levels combined (Table 10.7) were within 2.0% of the
label claim for the active substance with an R.S.D. < 2.0%, which satisfied the acceptance
criteria set for the study.
To evaluate the robustness of the developed RP-HPLC method, small deliberate
Variations in the optimized method parameters were done. The effect of change in flow rate, pH
and mobile phase ratio on the regent ion time and tailing factor were studied. The values for
284
proposed method are well within acceptance limits of 98-102%, with a RSD of less than 2.0%.
Above experiments indicated that the method is rugged and provides consistent and reliable
results. Limit of detection (LOD) and Limit of quantification (LOQ) were 0.65ppm and 24 ng/ml
respectively. Tablets were evaluated for the amount of Darunavir present in the formulation.
Each sample was analyzed in triplicate and the amount of was Darunavir 21.2 %.
A rapid, specific isocratic HPLC method has been developed for the determination of
Darunavir using a UV detector. The method was validated for accuracy, precision, linearity,
specificity & stability, limit of detection & limit of quantization and robustness & ruggedness.
The method uses a simple mobile phase composition, easy to prepare with little or no variation.
The rapid run time of 10 min and the relatively low flow rate allows the analysis of large number
of samples with less mobile phase that proves to be cost-effective. Efficient UV detection at 271
nm was found to be suitable without any interference from injectable solution excipients or
solvents. Statistical analysis of the results has been carried out revealing that the proposed
method is simple, sensitive and reproducible and hence can be used in routine for simultaneous
determination of Darunavir in bulk as well as in pharmaceutical preparations.
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10.7. BIBILOGRAPHY
1. Rodger D MacArthura; “Darunavir: promising initial results”, doi:10.1016/S0140-
6736(07)60499-1
2. Ghosh AK, Dawson ZL, Mitsuya H "Darunavir, “A conceptually new HIV-1
protease inhibitor for the treatment of drug-resistant HIV". Bioorg. Med. Chem;
2007; 15(24):7576–80.
3. “Darunavir-ritonavir more effective than Lopinavir-ritonavir in HIV infected,
treatment-experienced patients”, The Lancet; 2007, 370, article URL
4. “New RP-HPLC method for the determination of Darunavir in Tablet dosage
form”. Asian J. Pharm. Res; 2011; 1(1): 10-14.
5. “The Validation of Plasma Darunavir Concentrations Determined by the HPLC
Method for Protease Inhibitors”, Biological & Pharmaceutical Bulletin; 2007;30(
10): 1947-1949.
6. “Quantification of Darunavir (TMC114) in human plasma by high-performance
liquid chromatography with ultra-violet detection”; Journal of Chromatography
B” 2007; 857(2, 1): 327-331.
7. “The Effect of Lopinavir/Ritonavir and Darunavir/Ritonavir on the HIV Integrase
Inhibitor S/GSK1349572 in Healthy Participants”; J Clin Pharmacol; 2011; 51(2):
237-242.
8. “HPLC-MS method for the simultaneous quantification of the new HIV protease
inhibitor Darunavir, and 11 other antiretroviral agents in plasma of HIV-infected
patients”; J Chromatogr B Analyt Technol Biomed Life Sci; 2007; 859(2): 234-40.
9. “A simple and sensitive HPTLC method for quantitative analysis of Darunavir
ethanolate tablets”; JPC - Journal of Planar Chromatography - Modern TLC;
2011; 24: 232-235.
10. “Treatment simplification to once daily Darunavir/ritonavir guided by the
Darunavir inhibitory quotient in heavily pretreated HIV-infected patients”,
Antiviral Therapy; 2010; 15:219-25.