Journal of Pharmaceutical Investigation (2013) 43:251257

DOI 10.1007/s40005-013-0075-2

NOTES

Bioequivalence of Samchundang Berastolin tablet to Jeil Berasil

tablet (beraprost sodium 20 lg)
Hyun-Ah Kang Hwa Yoon Yong-Bok Lee

Received: 23 February 2013 / Accepted: 26 April 2013 / Published online: 10 May 2013
The Korean Society of Pharmaceutical Sciences and Technology 2013

Abstract Beraprost sodium, sodium ()-(1R*,2R*,3aS*,

8bS * )-2,3,3a,8b-tetrahydro-2-hydroxy-1-[(E)-(3S * )-3hydroxy-4-methyl-1-octen-6-ynyl]-1H-cyclopenta[b]benzofuran-5-butyrate), an orally absorbable prostacyclin
derivative (PGI2), has marked ischemic symptom treatments
like ulcer and pain with chronic arterial occlusion. The
purpose of the present study was to evaluate the bioequivalence of two beraprost sodium tablets, Samchundang Berastolin tablet (Samchundang Pharm. Co., Ltd.) and Jeil
Berasil tablet (Jeil Pharm. Co., Ltd.), according to the
guidelines of the Korea Food and Drug Administration
(KFDA). The in vitro release of beraprost from the two
beraprost sodium formulations was tested using KP IX
Apparatus II method with various dissolution media. Thirtytwo healthy Korean male volunteers, 23.44 1.48 years in
age and 65.95 8.94 kg in body weight, were divided into
two groups and a randomized 2 9 2 crossover study was
employed. After single administration, three tablets containing 20 lg as beraprost sodium, blood samples were
taken at predetermined time intervals and the concentrations
of beraprost in serum were determined using a LC/MS/MS
method with multiple reaction-monitoring. The dissolution
profiles of two formulations were similar in all tested dissolution media. The pharmacokinetic parameters such as

Hyun-Ah Kang and Hwa Yoon contributed equally to this study.

H.-A. Kang
Clinical Development Team, CJ Cheiljedang Corp.,
Seoul 100-400, Korea
H. Yoon
Y.-B. Lee (&)
Institute of Bioequivalence and Bridging Study, College of
Pharmacy, Chonnam National University, Gwangju 500-757,
Korea
e-mail: leeyb@chonnam.ac.kr

AUCt, Cmax and Tmax were calculated, and computer programs (Equiv Test and K-BE Test 2002) were utilized for
the statistical analysis of the parameters using logarithmically transformed AUCt, Cmax and un-transformed Tmax. The
results showed that the differences between two formulations based on the reference drug, Jeil Berasil tablet, were
2.12, 0.15 and 4 % for AUCt, Cmax, and Tmax, respectively.
There were no sequence effects between two formulations in
these parameters. The 90 % confidence intervals using
logarithmically transformed data were within the acceptance
range of log 0.8log 1.25 (e.g., log 0.9114log 1.0912 and
log 0.8471log 1.1253 for AUCt and Cmax, respectively).
Thus, the criteria of the KFDA bioequivalence guideline
were satisfied, indicating Samchundang Berastolin tablet
was bioequivalent to Jeil Berasil tablet.
Keywords Beraprost sodium
Berastolin tablet

Berasil tablet
Bioequivalence
LC/MS/MS

Introduction
Beraprost sodium (Fig. 1a), a prostacyclin derivative
(PGI2), has marked platelet-aggregation inhibition and
vasodilation activities and relatively week toxicity in animal models. These characteristics make promise therapeutic agent for the treatment of peripheral vascular
diseases. The mechanism of action of beraprost is inhibition of adenylate-cyclase dependent platelet aggregation
(Moriya et al. 2010). Beraprost sodium is an equimolar
mixture of two diastereoisomers. Each of these compounds
has two optical isomers (APS-314 and APS-315). The
inhibitory effects of APS-314d isomer on ADP-induced
platelet aggregation were 18 times more potent than those
of APS-315d isomer on ex vivo human platelets. These

123

252

antiplatelet effects persisted after 10 days of repeated oral

administration. The time to reach the maximum plasma
concentration (Tmax) of beraprost sodium and the half life
(t1/2) of the plasma beraprost sodium have been reported to
be 0.38 0.54 and 0.81 0.24 h after oral administration, respectively (Demolis et al. 1993; Nony et al. 1996).
The present study was conducted to determine the
pharmacokinetics and bioequivalence of two formulations
of beraprost sodium 20 lg tablet, reference (Jeil Berasil
tablet) and test (Samchundang Berastolin tablet) formulation, for the purpose of generic substitution. The test
included thirty-two subjects of healthy Korean male volunteers was performed by latin square design. Volunteers
were randomly assigned to receive a single dose of beraprost sodium tablet 60 lg (three tablets). Beraprost sodium
in serum was measured using LC/MS/MS spectrometer.
The two formulations were compared in terms of standard
pharmacokinetic parameters such as area under the curve
(AUCt), the maximum serum concentration (Cmax), and
Tmax according to the guidelines of Korea Food and Drug
Administration (KFDA) (KFDA 2012).

Materials and methods

Materials and instruments
Each of the study tablets contained 20 lg of beraprost
sodium. The test formulation (Samchundang Berastolin
tablet, Samchundang Pharm. Co., Ltd., lot no. 5L1) and
reference formulation (Jeil Berasil tablet, Jeil Pharm. Co.,
Ltd., lot no. BRF502) manufactured in accordance with the

H.-A. Kang et al.

Korean Good Clinical Practice (KGCP) guidelines (KFDA

2009) were supplied as tablet.
Methanol and acetonitrile (HPLC grade) were purchased
from Fisher Scientific (Fair Lawn, NJ, USA) and the other
chemicals were of HPLC grade or highest quality available.
HPLC grade water was obtained from a Milli-Q water
purification system (Millipore Co., Milford, MA, USA) and
used throughout the study. The mobile phase components
such as acetonitrile and ammonium acetate were filtered
through a 0.22 lm pore size membrane filter prior to
mixing and ultrasonically degassed after mixing.
The HPLC system consisted of a model LC-20AD isocratic pump (Shimadzu, Kyoto, Japan) and a model SLC20AC autosampler (Shimadzu, Kyoto, Japan). The separation was performed on a Capcell Pak C18 UG120 V
column (5 lm particle size, 150 9 2.0 mm I.D.; Shiseido,
Tokyo, Japan) with security guard cartridge (Phenomenex
C18, 4 9 3.0 mm I.D.; Phenomenex, Torrance, CA, USA)
using a mixture of acetonitrile-1 mM ammonium acetate
(60:40, v/v, adjusted to pH 5.0 with formic acid) at a flow
rate of 300 lL/min with a column inlet pressure of about
10 MPa.
The detection was performed using an API 4000 Q
TRAP mass spectrometer (Applied Biosystems-MDS
SCIEX, Toronto, Canada) equipped with a turbo ion spray
interface. Negative ion multiple reaction-monitoring
(MRM) mode was used. Nitrogen was used as auxiliary,
curtain and collision gas. Gas flow rates, source temperature, ion spray voltages and collision energies were optimized for every compound by infusion of 10 ng/mL
standards solutions in mobile phase at 300 lL/min and by
flow-injection analysis at LC flow rate. The main working
parameters of the mass spectrometer were described in
Table 1. The Analyst 1.4.1 software (Applied Biosystems)
was used for data processing.
In vitro dissolution test

Fig. 1 Chemical structures of a beraprost sodium and b limaprost

(internal standard)

123

In vitro dissolution testing was performed using Korean

Pharmacopoeia (KP) IX dissolution Apparatus II (paddle
method) and 900 mL of dissolution solution (pH 1.2, 4.0,
6.8 buffer and water) at 50 rpm at 37 0.5 C (The
Korean Pharmacopoeia IX 2007). Drug-release testing was
conducted on 12 individual dosage units of the 2 formulations. Samples were removed at 5, 10, 15 and 30 min,
after which they were filtered and assayed by HPLC with
UV/Vis detection at 285 nm. The dissolved beraprost
content was expressed as a percentage of the labeled beraprost content.
The acceptance criteria for assessment of equivalence of
dissolution profiles between two preparations are as follows. If mean dissolution of the reference formulation
reached 85 % within 15 min, then mean dissolution of the

Bioequivalence of Samchundang Berastolin tablet

Table 1 Tandem mass spectrometer working parameters
Parameter

Value

Source temperature (C)

700

Dwell time per transition (ms)

250

Ion source gas 1 (psi)

70

Ion source gas 2 (psi)

70

Curtain gas (psi)

10

Collision gas (psi)

Ion spray voltage (V)

-4,500

Entrance potential (V)

-10

Declustering potential (V)

-100 (analyte) and -90 (I.S.)

Collision energy (V)

-24 (analyte) and -16 (I.S.)

Collision cell exit potential (V)

Resolution

-7 (analyte) and -9 (I.S.)

Unit

Mode of analysis

Negative

Ion transition for beraprost (m/z)

397.2/268.9

Ion transition for limaprost (m/z)

379.2/343.0

test formulation should also reach 85 % within 15 min. If

mean dissolution from the reference formulation reached
85 % after [15 min, mean dissolution from the test formulation should not deviate by [15 % from that of the
reference formulation at 2 time points (KFDA 2012).
Selection of volunteers
The study population consisted of thirty-two healthy male
Korean volunteers with an average age of 23.44
1.48 years and an average weight of 65.95 8.94 kg.
Before enrollment, all subjects underwent clinical screening, including a physical examination and laboratory tests
(blood analysis: hemoglobin, hematocrit, RBC, WBC,
platelet, differential counting of WBC, total protein, albumin, sGOT, sGPT, alkaline phosphatase, total bilirubin,
cholesterol, creatinine, blood urea nitrogen, and glucose
fasting and urine analysis; specific gravity, color, pH,
sugar, albumin, bilirubin, RBC, WBC, and cast).
Subjects were excluded if they had a history of hepatic,
renal, respiratory, endocrine, or cardiovascular illness; or
had ingested alcohol or medications, including over-thecounter drugs, within 4 weeks before the study. This was
done to ensure that existing degree of variation would not
be due to an influence of illness or other medications. The
sample size (n = 15), calculated from the results of a
preliminary study by posterior power analysis, had sufficient statistical power ([80 %) to detect 20 % differences
in the pharmacokinetic parameters between the two preparations (a = 0.05). Finally, thirty-two subjects were
selected, including extra subjects for unexpected withdrawals or discontinuation. This calculation was performed
using nQuery Advisor Version 3 (Statistical Solutions Ltd.,

253

London, England). Written informed consent was obtained

from all subjects after the nature and purpose of the study
had been explained, in accordance with the KFDA guidelines for bioequivalence test (KFDA 2012).
Blood sampling from volunteers
The study protocol was approved by the Institutional
Review Board of the Institute of Bioequivalence and
Bridging Study, Chonnam National University. The study
was performed in accordance with the revised Declaration
of Helsinki (World Medical Association Declaration of
Helsinki 2000) and the Good Clinical Practice guidelines
(KFDA 2009).
All of the volunteers avoided taking other drugs for at
least 4 weeks prior to the study and until its completion.
They also refrained from consuming xanthine-containing
foods, alcoholic beverage for 12 h prior to each dosing and
until the collection of the last blood sample. The study had
a single-dose, randomized, two-treatment, two-period
crossover design. Subjects were stayed at the hospital at
8:00 p.m. on the day before the study and fasted for 12 h
before and 4 h after drug administration. At 8:00 a.m., a
cannula (JELCOTM, 22G, Johnson & Johnson Medical,
Pomezia, Italia) was inserted into the median cubital vein
and the cannula was flushed with 0.3 mL heparinized
normal saline solution for injection (150 U/mL) to prevent
clotting. Each subject was randomly assigned to receive a
single dose of the reference or test formulation (three of
20 lg beraprost sodium tablet; beraprost sodium 60 lg)
with 240 mL of water at 8:40 a.m.. Subjects received
standardized meals at 4 h after drug administration. After a
washout of 7 days, subjects received the alternative
formulation.
After 2 mL of blood was discarded, an aliquots of 5 mL
of blood was drawn from the indwelling cannula into a
5 mL Vacutainer tube (BectonDickinson and Company,
Franklin Lakes, New Jersey) before administration (to
serve as a control) and at 0.25, 0.5, 0.75, 1, 1.25, 1.5, 1.75,
2, 3, 4 and 6 h after administration. After sampling, the
cannula was flushed with 0.3 mL of heparinized normal
saline solution for injection. The samples were centrifuged
at 3,000 rpm, 20 min and the serum sample was transferred
to polyethylene tube and stored at -80 C until assayed.
Subjects were continuously monitored by hospital staff
throughout the study period. Vital signs (temperature,
blood pressure, and heart rate) were measured before and
6 h after drug administration.
Determination of serum beraprost concentration
To prepare the sample for assay, stock solutions of beraprost
at concentration of 1 mg/mL and limaprost (Fig. 1b, internal

123

254

standard) at concentration of 0.1 mg/mL were prepared in

water and kept at 4 C. Calibration standard serum samples
of beraprost were prepared at concentrations of 10, 20, 50,
100, 200, 500, 1000 and 2000 pg/mL in drug-free pooled
serum, In the same manner, quality control (QC) samples at
low (50 pg/mL), medium (200 pg/mL) and high (1,000 pg/
mL), respectively, were prepared to evaluate accuracy and
precision. To 800 lL of blank serum, calibration standards
and QC samples, 80 lL of I.S. (limaprost, 5 ng/mL in water),
and 80 lL of 3 N HCl solution were added to clean test
tubes. The samples were mixed by vortex-mixing for 10 s
and centrifuged at 12,000 rpm for 5 min. An Oasis HLB
cartridge (Milford, MA, USA) was conditioned with
sequential washing with 1 mL methanol followed by 1 mL
Milli-Q water. The supernatant (0.9 mL) was loaded onto the
cartridges followed by elution with 1.6 mL of methanol. The
extract was evaporated under a N2 gas stream. The residue
was dissolved in 80 lL of 20 % acetonitrile and then 20 lL
of the solution was injected directly onto the LC/MS/MS
system. The interference by endogenous compounds was
assessed by analyzing standards of beraprost drug-free
serum samples, serum spiked with beraprost, and serum
samples obtained from subjects given beraprost sodium
tablets. Beraprost was quantitated by weighted linear
regression analysis of the peak area ratio versus concentrations of added beraprost using 1/concentration as the
weighting factor. The calibration curves were linear from 10
to 2,000 pg/mL. The lower limit of quantitation (LLOQ) was
defined as the lowest concentration at 10 times the signal-tonoise ratio that yielded a precision of \20 % coefficients of
variation (CV) and an accuracy between 80 and 120 % of the
theoretical value. The LLOQ was 10 pg/mL in five replicate
samples. In order to assess the intra- and inter-day precision
and accuracy of the assay, QC samples were prepared as
described above. The intra-day precision of the assay was
assessed by calculating the CV for the analysis of QC samples in five replicates, and inter-day precision was determined through the analysis of QC samples on five
consecutive days. The precision of the assay was evaluated
based on the criterion that the relative standard deviation
(SD) for each concentration level should not exceed
15 %, with the exception of the LLOQ, which should not
exceed 20 %. Accuracy was determined by comparing
the calculated concentration using calibration curves to
known concentration. The criterion for accuracy was that the
SD for the mean value should not exceed the normal concentration by more than 15 %, except for the LLOQ, for
which the limit was 20 %.
Statistical analysis of pharmacokinetic parameters
Each volunteer received an oral dose of 60 lg of beraprost
sodium in a standard 2 9 2 crossover method in a

123

H.-A. Kang et al.

randomized order. Pharmacokinetic parameters such as

AUCt, Cmax and Tmax were calculated by non-compartmental analysis of serum concentrationtime curve data
using WinNonlin software (Pharsight Corporation, CA,
USA 19981999). The peak concentration (Cmax) and the
time to reach Cmax (Tmax) were determined by the
inspection from individual serum concentrationtime profiles for beraprost. The AUCt was calculated by the linear
trapezoidal rule from 0 to 6 h. The ratios (test/reference)
using log-transformed data, were analyzed with the analysis of variance (ANOVA) that performed with the Equiv
Test (Statistical Solutions Ltd. 2001) and K-BE Test
program (Lee et al. 2000) at a significant level of 0.05.
The bioequivalence of two beraprost tablets was estimated
by AUCt and Cmax. Tmax used as a reference value.

Result and discussion

Dissolution testing
Accordance of KP IX dissolution Apparatus II method,
dissolution testing was done to each of 12 test and reference tablets. Both formulations released [85 % of beraprost sodium within 15 min in all test dissolution media
(pH 1.2, 4.0, 6.8 buffer and water) and had similar release
profiles. So, two formulations had no difference in dissolution testing (Fig. 2).
Analysis of beraprost in serum samples
The method was validated according to FDA guidance and
international guidelines US Department of Health and
Human Services, Food and Drug Administration, Center
for Drug Evaluation and Research, Center for Verternary
Medicine 2001. Figure 3 shows MRM chromatograms of
blank, spiked beraprost containing limaprost and serum
sample from a healthy subject obtained 1 h after oral
administration of beraprost sodium 60 lg. No interference
from endogenous substances was observed in human
serum. The retention time for beraprost and IS (limaprost)
was about 1.81.9 min (Fig. 3b). In this method, beraprost
and limaprost were well separated from the biological
background under the described chromatographic condition, respectively. These peaks were of good shape, completely resolved one. The calibration curve, established by
plotting the peak area ratio (y) versus concentration (x),
was linear over the range from 10 to 2,000 pg/mL with the
following regression equation: y = 0.00162x ? 0.00677
(r = 0.9987, p \ 0.01). The LLOQ of beraprost in human
serum was 10 pg/mL; at this concentration, the CV for
accuracy was 112 %, and the CV for precision below

Bioequivalence of Samchundang Berastolin tablet

255

110

Table 2 Precision and accuracy for the analysis of beraprost concentration in human serum

100

Released beraprost (%)

90

Concentration
(pg/mL)

80

50
40
30

Accuracy
(%, n = 5)

Intra-day CV (%)
(n = 5)

Inter-day CV (%)
(n = 5)

10

8.43

4.67

112

50

7.76

3.13

99.5

200

5.23

3.31

95.5

1,000

3.80

4.50

98.6

70
60

Precision CV (%)

20

CV Coefficient of variation = 100 9 SD/mean

10
0

10

15

20

25

30

Time (min)

Fig. 2 Dissolution profiles of beraprost in water from reference

(filled circle) and test (unfilled circle) formulation. Vertical bars
represent the mean SD (n = 12)

8.43 %. During validation, the CV for accuracy ranged

from 98.6 to 112 %, whereas intra- and inter-day CVs for
precision remained blow 8.43 and 4.67 %, respectively.
These results indicated that the present method has a satisfactory accuracy and precision (Table 2).
Pharmacokinetic analysis

Serum concentraion of beraprost (pg/mL)

0
1000
900
800
700
600
500
400
300
200
100
0
0

Time (min)

LC/MS/MS method was successfully applied to a bioequivalence test in which serum concentrations of beraprost in thirty-two healthy male volunteers were

Fig. 4 Mean (SD, n = 32) serum concentrationtime curves of

beraprost after single oral administration of the reference (filleed
circle) and the test (unfilled circle) beraprost sodium tablets as
beraprost sodium 60 lg

Fig. 3 Chromatograms of a blank human serum; Mass(es): 397.2/

268.9 amu b blank human serum spiked with beraprost (500 pg/mL)
containing limaprost (internal standard); Mass(es): 397.2/268.9 amu

and c a human serum sample from a healthy Korean male volunteer at

1 h after administration of a single oral dose of beraprost sodium
60 lg; Mass(es): 397.2/268.9 amu

123

256

H.-A. Kang et al.

determined up to 6 h after the oral administration of 60 lg

beraprost sodium. Figure 4 shows the mean serum concentrationtime curves of beraprost following single oral
administration of test and reference tablets, and descriptive
statistics of the derived pharmacokinetic parameters such
as AUCt, Cmax, and Tmax for two formulations are summarized in Table 3.
The mean (SD) AUCt was 852.08 359.40 pg h/mL
for the test formulation and 834.36 369.05 pg h/mL for

the reference formulation. Mean (SD) Cmax values for the

test and the reference formulation were 644.31 290.93
and 643.34 252.79 pg/mL, with mean (SD) Tmax values of 0.61 0.34 and 0.59 0.36 h, respectively
(Table 3). The differences of the means of the test to reference medication for AUCt and Cmax were 2.12 and
0.15 %, respectively, which are generally accepted if the
differences of mean values for AUCt and Cmax lie
within 20 % (Table 4).

Table 3 Bioavailability parameters in normal and logarithmic scales for each volunteer obtained after oral administration of Samchundang
Berastolin and Jeil Berasil tablets at the beraprost sodium dose of 60 lg
Subjects

Parameter
AUCt (pg h/mL)
Reference
Value

Cmax (pg/mL)
Test

Log

Value

Tmax (h)

Reference

Test

Reference

Test

Log

Value

Log

Value

Log

Value

Value
0.25

A1

826.25

6.72

979.49

6.89

936

6.84

1,440

7.27

0.25

A2

1,059.83

6.97

976.58

6.88

754

6.63

752

6.62

0.75

0.25

A3

312.75

5.75

236.30

5.47

479

6.17

402

6.00

0.25

0.50

A4

861.63

6.76

1,037.68

6.94

623

6.43

711

6.57

1.00

0.75

A5

923.28

6.83

1,067.28

6.97

357

5.88

428

6.06

2.00

1.75

A6

855.16

6.75

325.65

5.79

1,220

7.11

253

5.53

0.25

0.50

A7

915.98

6.82

843.90

6.74

798

6.68

562

6.33

0.50

0.75

A8

407.54

6.01

382.05

5.95

334

5.81

191

5.25

0.75

1.00

A9

932.70

6.84

1,100.97

7.00

979

6.89

538

6.29

0.25

1.00

A10

574.99

6.35

332.23

5.81

515

6.24

293

5.68

0.25

0.50

A11

622.83

6.43

1,154.58

7.05

404

6.00

1,350

7.21

0.50

0.25

A12

850.93

6.75

864.93

6.76

449

6.11

504

6.22

0.75

1.00

A13

1,154.68

7.05

1,243.90

7.13

492

6.20

940

6.85

0.75

0.25

A14
A15

655.54
616.65

6.49
6.42

484.41
659.48

6.18
6.49

482
369

6.18
5.91

507
416

6.23
6.03

0.75
1.00

0.50
0.75

A16

522.78

6.26

413.94

6.03

331

5.80

359

5.88

1.00

0.25

B1

935.93

6.84

899.65

6.80

956

6.86

844

6.74

0.25

0.25

B2

756.80

6.63

801.00

6.69

473

6.16

677

6.52

0.50

0.50

B3

768.03

6.64

917.08

6.82

454

6.12

552

6.31

0.75

1.00

B4

998.48

6.91

1,555.58

7.35

635

6.45

886

6.79

0.75

0.50

B5

645.53

6.47

522.38

6.26

679

6.52

844

6.74

0.50

0.25

B6

1,313.10

7.18

1,415.28

7.26

859

6.76

992

6.90

0.50

0.75

B7

913.60

6.82

1,129.25

7.03

948

6.85

654

6.48

0.50

1.00

B8

795.51

6.68

647.17

6.47

712

6.57

501

6.22

0.25

0.75

B9

730.05

6.59

839.73

6.73

535

6.28

751

6.62

0.50

0.50

B10

785.05

6.67

936.23

6.84

657

6.49

553

6.32

0.50

0.75

B11

437.15

6.08

487.98

6.19

509

6.23

539

6.28

0.25

0.25

B12

424.06

6.05

701.23

6.55

355

5.87

426

6.05

0.50

0.50

B13
B14

1,009.23
1,190.33

6.92
7.08

838.75
1,240.35

6.73
7.12

670
985

6.51
6.89

460
902

6.13
6.80

0.25
0.50

0.50
0.75

B15

2,379.25

7.77

1,650.35

7.41

1,190

7.08

959

6.87

1.00

0.25

B16

523.88

6.26

581.10

6.36

448

6.10

435

6.08

0.25

0.75

Mean

834.36

6.65

852.08

6.65

643.34

6.39

644.31

6.37

0.59

0.61

SD

369.05

0.39

359.40

0.48

252.79

0.38

290.93

0.46

0.36

0.34

123

Bioequivalence of Samchundang Berastolin tablet

257

Table 4 Statistical results of bioequivalence evaluation between two beraprost sodium tablets
Parameters
AUCt

Cmax

Tmax

Difference

2.12 %

0.15 %

4%

FaG

2.3018

3.0368

1.6085

Test/reference point estimate

0.9973

0.9764

0.0234

Log 0.9114 B d B log 1.0912

Log 0.8471 B d B log 1.1253

-14.43 % B d B 22.43 %

Confidence interval (d)

The AUCt and Cmax values were calculated on the basis of in-transformed data, and the Tmax values on the basis of un-transformed data
a

a = 0.05, F (1, 30) = 4.17

a = 0.05

Bioequivalence analysis
No significant sequence, subject, formulation or period
effects were detected for any pharmacokinetic parameters.
The point estimates for the mean ratio of the test to the
reference formulation for the AUCt, Cmax were 0.9973,
0.9764, respectively (Table 4). The parametric 90 % confidence intervals were in the range of log 0.9114log
1.0912 and log 0.8471log 1.1253, respectively (Table 4),
which were entirely within the regulatory acceptance limits
for bioequivalence (80125 %) (KFDA 2011). The results
obtained from the two programs (Equiv Test and K-BE
test) were not different. This proved that there was no
significant difference between the bioavailability of reference and test formulations.

Conclusion
This validated LC/MS/MS method was sensitive, reproducible and accurate for the determination of beraprost in
human serum samples collected for bioequivalence studies.
Using this method, the bioequivalence of two different
beraprost sodium tablets was examined at the dose of
60 lg in thirty-two healthy normal male volunteers. No
significant differences in AUCt or Cmax were found
between the test and reference formulations and the calculated 90 % confidence intervals for the ratios of mean
AUCt and Cmax were within the regulatory acceptance
range for bioequivalence (80125 %).
Acknowledgments This study was supported by a contract between
Samchundang Pharm. Co. Ltd. and the Institute of Bioequivalence
and Bridging Study of Chonnam National University. The authors

have indicated that they have no conflicts of interest regarding the

content of this article.

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