A COMPARATIVE STUDY ON THE IN VITRO

DISSOLUTION PROFILES OF COMMERCIALLY

AVAILABLE CLARITHROMYCIN ORAL DOSAGE

FORMS IN NAIROBI COUNTY, KENYA

By:Manani O. Rebecca

U59/81391/2012

Department of Pharmaceutical ChemistrySchool of PharmacyUniversity of Nairobi

November 13, 2014

OUTLINE

Introduction

Study justification

Study objectives

Methodology

Results and discussion

Conclusion

Recommendations

Acknowledgements2

INTRODUCTION: MACROLIDES

Natural and semi-synthetic antibiotics characterised by macrocyclic rings with attached sugars

Source: Streptomyces and micromonospora species

Classification

14– erythromycin, clarithromycin, roxithromycin

15 – semi-synthetic azalides: azithromycin, gamithromycin

16 – josamycin, spiramycin, rokitamycin3

MACROLIDES (CONT’D) Mode of Action

Inhibit protein synthesis; prevent transpeptidation Spectrum of Activity

G+ve: S. aureus, Str. pyogenes, Str. pneumoniae

G-ve: H. influenzae, N. meningitidis, H. pylori

Atypical: M. pneumoniae, C. trachomatis, Mycobacteria, Toxoplasma, Borrelia spp.

Limitations of Natural Macrolides

Acid instability, GIT side-effects, resistance, narrow spectrum

4

CLARITHROMYCIN

Chemistry

2nd generation semi-synthetic macrolide

5

O

CH3O

H3CHO

HO

H3CH5C2

O

CH3

CH3

OCH3

O

O CH3

OHOCH3H3C

OCH3

O CH3

NCH3H3C

HO

Clarithromycin

Erythromycin A

1

3

6

9

12

O

CH3O

H3CHO

HO

H3CH5C2

O

CH3

CH3

OH

O

O CH3

OHOCH3H3C

OCH3

O CH3

NCH3H3C

HO

5

STABILITY OF CLARITHROMYCIN

Oxidative; basic; acidic conditions6

O

CH3O

H3CHO

HO

H3CH5C2

O

CH3

CH3

OCH3OCH3

O CH3

NCH3H3C

HO

OH

Decladinosylclarithromycin

O

CH3O

H3C

HO

H3CH5C2

O

CH3

CH3

OCH3

O

O CH3

OHOCH3H3C

OCH3

O CH3

NCH3H3C

HO

10,11-Anhydroclarithromycin

9,12-Hemiketal

O

CH3

H3C

H3CH5C2

O

CH3

CH3

OCH3

O

O CH3

OHOCH3H3C

OCH3

O CH3

NCH3H3C

HO

O

O

CH3O

H3CHO

HO

H3CH5C2

O

CH3

CH3

OCH3

O

O CH3

OHOCH3H3C

OCH3

O CH3

NH3C

HO

CH3

O

Clarithromycin N-oxide

STUDY JUSTIFICATION Quality of drugs crucial in treatment outcomes Previous reports of poor quality products in 3rd

world countries Clarithromycin is in EML - Kenya, WHO BCS classification

Tablets – class II (Low aq. solubility, low permeability) Suspensions – class IV (Low aq. solubility, low

permeability) Generics used as a cost cutting measure No bioequivalence testing centres in Kenya No previous PMS studies on CLA in Kenya

7

STUDY OBJECTIVES

General Objective

To conduct comparative in vitro dissolution studies on oral clarithromycin products in Nairobi County

Specific Objectives

To carry out product sampling in Nairobi County

To carry out identification and assay tests

To carry out comparative dissolution testing – innovator, generics

To determine pharmaceutical equivalence 8

METHODOLOGY I: SAMPLING Study population – 125 mg/5 mL susp, 500 mg

tablets

Time frame – January to March 2014

Stocking patterns revealed uneven distribution

Sites –11 outlets in Nairobi, 1 in England, UK

16 samples – 12 tabs and 4 suspensions (67% stocking rate in Nairobi)

Sample size – 60 tablets and 500 mL suspensions 9

METHODOLOGY II: METHOD OPTIMIZATION

MOA – published method (Abuga et al. 2001)

Method optimization

Detection wavelength, Sample injection volume, Mobile phase flow rate

Quenching conditions - dissolution pH 1.2

Optimum conditions: mp - ACN-0.2 M phosphate buffer, pH 6.80-water (40:3.5:56.5, v/v/v), flow rate -1.5 mL/min, sp - XTerra RP C18, 5 m (250 mm x 4.6 mm ID), temp - 56 oC detection 205 nm, quenching – 3 mL of 0.2 M NaOH 10

METHOD OPTIMIZATION (CONT’D)

Fig.1 - Typical assay chromatogram for clarithromycin working standard under the optimum chromatographic conditions

11

0.0 5.0 10.0 15.0 20.0 min

-75

-50

-25

0

25

50

75

100

125

mAU205nm,4nm (1.00)

CLA

METHODOLOGY III: ASSAY AND DISSOLUTION

USP and BP methods for sample preparation

Tabs – uniformity of weight (BP), assay

Granules – RD, extraction (USP), assay

Dissolution carried out at pH 1.2, 4.5, 6.8

Run time - 60 min. tabs, 90 min. suspensions

6 sampling time points

Dissolution profiles generated and f2 factors calculated

f2= 50×log {[1+ (1/n) Σ t=1n (Rt-Tt) 2] -0.5 ×100} 12

RESULTS I: ASSAY

Table 1 (a) – Assay Results for CLA Tablets

Table 1 (b) – Assay Results for CLA Suspensions

13

Sample Code

C1 C2 C3 C4 C5 C6 C7

% Content 100.9 100.2 100.1 102.1 98.8 99.3 98.7

C8 C9 C10 C11 C12 C17

101.5 102.0 98.8 98.4 105.9 103.5

Sample Code

C13 C14 C15 C16 C18

% Content 107.8 108.8 99.5 109.6 110.1

RESULTS II: ACIDIC DEGRADATION

CLA std incubation - 37 oC in 0.1 M H3PO4 (pH 1.4) and 0.1 M HCl (pH 1.2)

Table 2 – Acid Degradation Parameters for CLA Std

14

Degradation Parameters

pH 1.4 pH 1.2

Half life (min) 185 36.7

Rate constant (min-1) 3.7 x 10-3 19 x 10-3

% API degradation in 40 min

13.8 60

RESULTS III: DISSOLUTION AT PH 1.2

Fig. 2 – Chrm for dissolution at pH 1.2, 30 min.

Dissolution proceeded with concurrent degradation – profiles created using Total Clarithromycins

0.0 5.0 10.0 15.0 20.0 min

-25

0

25

50

75

100

125

150

mAU 205nm,4nm (1.00)

DECL

CLA

15

RESULTS IV: F2 FACTORS AT PH 1.2

Fig. 3 – Similarity factors at pH 1.2

Compliance rate 50% (6/12)16

RESULTS V: F2 FACTORS AT PH 4.5

Fig. 4 – Similarity factors at pH 4.5

Compliance rate 67% (8/12)17

RESULTS VI: F2 FACTORS AT PH 6.8

Fig. 5 – Similarity factors at pH 6.8

Compliance rate 50% (6/12) 18

F2 FACTORS – COMPLIANT SAMPLES

Table 3: Similarity factors for compliant samples

Overall compliance rate 25% (4/16 products)19

Sample Code

Dosage Form

f2 factors (%)

pH 1.2 pH 4.5 pH 6.8

C1 Tablet 80.4 93.6 54.8

C6 Tablet 54.6 52.4 64.4

C11 Tablet 64.7 80.9 61.8

C17 Tablet 72.2 75.2 55.2

F2 FACTORS – NON-COMPLIANT SAMPLES

Table 4: Similarity factors for non-compliant samples

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Sample Code

Dosage Form

f2 Factors (%)

pH 1.2 pH 4.5 pH 6.8

C2 Tablet 46.0 30.0 54.8

C3 Tablet 40.2 53.7 35.5

C4 Capsule 69.2 16.9 26.5

C5 Tablet 28.8 61.5 33.3

C7 Tablet 37.9 25.3 27.6

C8 Tablet 58.9 69.5 45.4

C9 Tablet 44.8 34.2 34.7

C10 Tablet 22.2 56.6 79.7

C13 Susp. ND ND 42.4

C14 Susp. ND ND 14.3

C15 Susp. ND ND 15.2

C18 Susp. ND ND 42.0

DISCUSSION

Variability in product performance

Table 5 – Inconsistent Products

Gastric pH 0.5 - 2 adults, 1.5 - 3 children, 2 sick state

Gastric residence time 0.5 – 2 hrs Significant degradation noted at 30 min, pH 1.2

Product Code

f2 factors (%)

pH 1.2 pH 4.5 pH 6.8

C2 46.0 30.0 55.8

C8 58.9 69.5 45.4

C10 22.2 56.6 79.7

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DISCUSSION (CONT’D) Variability in performance of innovator products

in different markets:

Significant differences not expected due to change in site – GMP, SUPAC-related guidelines

Minimal API release for suspensions C14 and C15 – implications of sub-optimal product dissolution

22

pH 1.2 4.5 6.8

f2 Value (Tabs) 72 75 55

f2 Value (Susp.) ND ND 42

CONCLUSION

Not all available CLA products meet quality standards

Proof of quality and pharmaceutical equivalence requires more than assay and single point dissolution tests

Role of DRAs critical in QA of pharmaceuticals

Pre-registration

Pharmacovigilance - effective, sustained, targeted

Staff – proper education, training, experience23

RECOMMENDATION

In vitro dissolution profiling to be included in routine QC and post-marketing surveillance tests

Sustained PMS and PV activities by DRAs

Sufficiently deterrent measures for non-compliant products by DRAs to discourage circulation of poor quality products

Further studies on other antibiotics in circulation, special attention paid to macrolides 24

ACKNOWLEDGEMENTS

Supervisors - Dr. K. O. Abuga

- Dr. H. K. Chepkwony

NQCL management and staff

University of Nairobi

Ministry of Health

Technical staff – C. Rotich, D. Nyamweya, J.

Kalama, H. Mugo, O. King’ondu, J. Nguyo 25

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