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`Journal Copy
`DOCLINE:
`Hindustan antibiotics bulletin
`Hindustan Antibiot Bull
`1974 May;16(4):175-84
`Stability studies on hamycin and tetracycline hydrochloride
`Trivedi BM;Rupare!iya MT
`2985097R Verify: PubMed
`4549311
`0018-1935 (Print)
`Any format
`Dr M 3 Thirumalachar, Pimpri :
`Copyright Compliance Guidelines
`JAMIE FRIDAY
`
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`This material may be protected by copyright law (TITLE 17,U.S. CODE)
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`

`

` .;
`
`[
`
`110.
`
`ihnol
`
`ows:
`
`Stability Studies on Hamycin and
`Tetracycline Hydrochloride with Selected Diluents
`t-
`Stability Studies on Hamycin and
`i;
`Tetracycline Hydrochloride with Selected Diluents
`BHANOO M. TRIVEDI AND M. T. RUPARELIYA
`l
`
`L. M. College of Pharmacy, Ahmedabad-380 009
`BHANOO M. TRIVEDI AND M. T. RUPARELIYA
`
`L. M. College of Pharmacy, Ahmedabad-380 009
`Tetracyclines are one of the widely used
`in patients subjected to intensive tetracy-
`I
`I' broad spectrum antibiotics. An extensive
`cline therapy and has been resulted in death
`in patients subjected to intensive tetracy-
`Tetracyclines are one of the widely used
`i use of tetracyclines has resulted in emer-
`in some cases. Amongst many annoying
`.
`cline therapy and has been resulted in death
`broad spectrum antibiotics. An extensiVe
`I gence of certain problems. Many of the
`symptoms the staphylococcal take-over is
`in some cases. Amongst many annoying
`use of tetracyclines has resulted in emer-
`! side effects are believed to result from an
`perhaps the most serious and number of
`symptoms the staphylococcal
`take-over is
`gence of certain problems. Many of the
`f outgrowth of Candida albicans resulting
`such cases have been terminated fatally.
`perhaps the most serious and number of
`side effects are believed to result from an
`'
`in candidiasis (Stone and Mersheimer, 1956),
`Patients treated with pneumonia with these
`such cases have been terminated fatally.
`outgrowth of Candida albicans
`resulting
`|
`antibiotics have been cured of the primary
`Patients treated with pneumonia with these
`in candidiasis(Stone and Mersheimer, 1956).
`antibiotics have been cured of the primary
`infection, only to die of systemic staphyloco-
`Among the undesirable organisms that
`infection, only to die of systemic staphyioco-
`. proliferate in the intestinal tract during or
`cosis. Death often
`results also
`from
`Among the undesirable organisms that
`cosis. Death often
`results also
`from
`proliferate in the intestinal tract during or
`following extended or shorter treatment,
`superficial necrosis of large area of the
`superficial necrosis of large area of the
`following extended or shorter
`treatment,
`. with tetracyclines are Candida albicans and mucosa of the small intestine which lead to
`mucosa of the small intestine which lead to
`with tetracyclines are Candida albicans and
`i other yeast and fungi ; abnormal coliform,
`profuse watery diarrhoea, dehydration and
`profuse watery diarrhoea, dehydration and
`other yeast and fungi ; abnormal coliform,
`such as species of Proteus, Pseudomonas
`circulatory collapse.
`circulatory collapse.
`such as species of Proteus, Pseudomonas ..
`. and resitant staphylococci. The emergence
`and resitant staphylococd. The emergence
`of these organisms in the mouth and gas­
`of these organisms in the mouth and gas-
`trointestinal
`tract of patients receiving
`trointestinal
`tract of patients
`receiving
`broad spectrum antibiotics has been ascribed
`broad spectrum antibiotics has been ascribed
`to suppression of normal microbial inhabi-
`to suppression of normal microbial inhabi-
`tant that ordinarily overgrow, and prevent
`tant that ordinarily overgrow, and prevent
`rapid proliferation of Candida and of other
`rapid proliferation of Candida and of other
`yeast and fungi
`through their antibiotic
`yeast and fungi through their antibiotic
`action.
`action.
`Another contributing factor may be the
`Another contributing factor may be the
`deprivation of microbially produced nutri-
`deprivation of microbially produced nutri­
`tional
`factor
`to which the host may be
`tional factor to which the host may be
`subjected during tetracyclines
`treatment
`subjected during
`tetracyclines
`treatment
`resulting in
`the weakening of mucous
`resulting
`in
`the weakening of mucous
`membrane and thus becoming susceptible
`portal of entry for pathogenic organisms,
`! membrane and thus becoming susceptible
`normally unable to penetrate intact, healthy
`;
`portal of entry for pathogenic organisms,
`mucosa. Some are of the opinion that
`)
`normally unable to penetrate intact, healthy
`broad spectrum antibiotics may actually
`mucosa. Some are of the opinion that
`stimulate growth of yeast and fungi. These
`broad spectrum antibiotics may actually
`organisms, probably are responsible for
`stimulate growth of yeast and fungi. These
`anal
`itching and cause of pruritis vulvae.
`organisms, probably are responsible for
`anal itching and cause of pruritis vulvae.
`Occasionally saprophytes and semi sapro-
`phytes have emerged as invansive pathogens
`Occasionally saprophytes and semi sapro­
`May. 1974
`phytes have emerged as invansive pathogens
`
`0W3:
`
`
`
`...4;st
`
`\
`
`Various attempts have been made to
`Various attempts have been made to
`tackle this problem, one of which being
`tackle this problem, one of which being
`combining tetracyclines with antifungal anti­
`combining tetracyclines with antifungal anti-
`biotics. Nystatin and amphoterecin-B have
`biotics. Nystatin and amphoterecin-B have
`been already employed for these purpose.
`been already employed for these purpose.
`In case of nystatin though the number of
`In case of nystatin though the number of
`Candida popuiation is reduced during treat-
`Candida population is reduced during treat­
`ment, it increases very rapidly after ceasation
`ment, it increases very rapidly after ceasation
`of the treatment. As regards amphotere-
`of the treatment. As regards amphotere-
`cin—B, the effect of reducing candidial popu-
`cin-B, the effect of reducing candidial popu­
`lation in gastro intestinal tract is of short
`lation in gastro intestinal tract is of short
`duration (0501, et al. 1967).
`duration (Osol, et al. 1967).
`Hamycin, a polyeue antifungai antibiotic
`has been claimed to show in viva, antifungai
`Hamycin, a polyene antifungal antibiotic
`activity on large number of fungi, patho-
`has been claimed to show in vivo, antifungal
`gens,
`saprophytes yeast
`like
`fungi
`and
`activity on large number of fungi, patho­
`Candida
`albr'cans. Hamyein
`has
`been
`gens, saprophytes yeast
`like fungi and
`proved very promising in the treatment of
`Candida albicans. Hamycin has
`been
`candidiasis as compared to nystatin and
`proved very promising in the treatment of
`amphoterecin-B (Vad and Thirumalachar,
`candidiasis as compared to nystatin and
`1968].
`amphoterecin-B (Vad and Thirumalachar,
`1968).
`With this in view, stability studies were
`planned for an oral formulation of tetracy-
`With this in view, stability studies were
`planned for an oral formulation of tetracy-
`
`l I
`
`l 1
`
`:
`;
`'
`i
`
`.
`
`•
`
`'
`
`1 May, 1974
`
`Apotex Exhibit 1011
`Page 2 of 11
`
`

`

`176
`
`HINDUSTAN ANTIBIOTICS BULLKTIN
`
`the
`cline hydrochloride with hamycin in
`employing log conc. in mcg/ml against
`176
`HINDUSTAN ANTIBIOTICS BULLETIN
`form of capsules. The investigation was
`diameters of zones of inhibition in mm.
`aimed at the stability studies of both anti­
`cline hydrochloride with hamycin in the
`biotics in the presence of another along
`form of capsules.
`The investigation was
`with selected diluents (Rupareliya, 1973).
`aimed at the stability studies of both anti-
`biotics in the presence of another along
`MATERULS AND METHODS
`with selected diluents (Rupareliya, 1973).
`
`employing log cone.
`in meg/ml against
`Microbiological Estimation of Tetracyline
`diameters of zones of inhibition in mm.
`Hydrochloride
`Microbiological Estimation of Tezracyfine
`Medium and test organism used in estima­
`Hydrochloride
`tion were same as mentioned in LP.
`Medium and test organism used in estima.
`Medium (120 ml) was inoculated with
`tion were same as mentioned in LP.
`spore suspension (1.8 ml) of B, pumilus
`which was
`transferred
`in glass plate
`Medium (120 ml) was
`inoculated with
`spore suspension (1.8 ml) of B. pumilus
`(24 x 24 cms). In the solidified medium
`which was
`transferred
`in
`glass
`plate
`cups of 8 mm.
`diameter were bored,
`(24x24 ems).
`In the solidified medium
`which were filled with the selected dilution
`cups of 8 mm.
`diameter were
`bored,
`of tetracycline (2, 4, 6, 8, 10, 12, 14, 16, 18
`which Were filled with the selected dilution
`and 20 mcg/ml). After allowing diffusion
`of tetracycline (2, 4, 6, 8, 10, 12, I4, 16, 18
`of tetracycline hydrochloride in the medium
`and 20 meg/ml). After allowing diffusion
`the plates were incubated at 37° for 24
`of tetracycline hydrochloride in the medium
`hours.
`the plates were incubated
`at 37° for 24
`hours.
`After incubation period zones of inhibi­
`tion were recorded and a standard graph
`After incubatiora period .zones of inhibi-
`tion were recorded and a standard graph
`was plotted employing log conc.
`in meg/
`was plotted employing log cone.
`in 111ch
`ml against diameter of zones of inhibition
`ml against diameter of zones of inhibition '
`in mm.
`in mm.
`
`Preparation of dessicators for exposing the
`Preparation of dessicatars for exposing the
`capsules at varying experimental conditions :
`capsules or varying experimental conditions .'
`
`As shown below the saturated salts solu­
`As shown below the saturated salts solu-
`tions were filled
`in the dessicators to get
`tions were filled in the dessicators to get
`30% and 45% relative humidities at 5°,
`30% and 45% relative humidities at 5°,
`37° and 50°. Capsules were also exposed
`37° and 50°. Capsules were also exposed
`to room temperature.
`to room temperature.
`———-.—_._.__._.__...._..____,________
`
`Materials :
`MATERIALS AND METHODS
`All chemicals employed in this investiga­
`Materials :
`tion were of the best available quality.
`All chemicals employed in this invostiga-
`Hamycin was received from Hindustan
`tion were of the best available quality.
`Antibiotics, Pimpri, Poona.
`Hamycin was
`received from Hindustan
`Antibiotics, Pimpri, Poona.
`Tetracycline hydrochloride was received
`from Cyanamid (India) Ltd., Bulsar.
`Tetracycline hydrochloride was received
`from Cyanamid (India) Ltd., Bulsar.
`Glass distilled water was used for prepa­
`ring solutions whenever required. Hard
`Glass distilled water was used for prepa-
`gelatin capsules of No, 0 were employed
`ring solutions whenever
`required. Hard
`gelatin capsules of No. 0 were employed
`for filling
`the antibiotics-diluent mixtures.
`for filling the antibiotics-diluent mixtures.
`Assay Methods ;
`Assay Methods :
`I . Microbiological Estimation of Hamycin
`l. Microbiological Estimation of Hamycin
`Medium—Modified Sabourad's Agar
`Medium—Modified
`Sabourad’s Agar
`adjusted
`to pH 5.5 was sterilized by
`adjusted
`to pH 5.5 was
`sterilized by
`autoclaving.
`autoclaving.
`
`Test organism : Paecelomyces variotm
`Test organism: PaeCelomyces variotm
`spores in sterile normal saline containing
`spores in sterile normal saline containing
`1
`:5000 tween
`80
`and having 30%
`1
`: 5000
`tween 80
`and having 30%
`transmittance constituted the test organism
`transmittance constituted the test organism
`for the assay of Hamycin.
`for the assay of Hamycin.
`Plotting of a standard graph :
`Plotting of a standard graph :
`Modified Sabourad’s agar (120 ml) was
`Modified Sabourad's agar (120 ml) was
`inoculated with spore suspension (2 ml),
`inoculated with spore suspension (2 ml),
`which was
`transferred
`in
`glass plates
`(24 x 24. cms).
`In the solidified medium
`which was
`transferred
`in glass plates
`cups of 8 mm diameter were bored which
`(24 x 24. cms).
`In the solidified medium
`were filled with the selected dilutions of
`cups of 8 mm diameter were bored which
`Hamycin (0.5, 0.6, 0.7, 0.8, 09,1016,
`were filled with the selected dilutions of
`2.0, 2.5, 3.0, 3.5, 4.0, 4.5, 5.0, 5.5 and
`Hamycin (0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.5,
`6.0 meg/ml). After allowing diffusion of
`2.0, 2.5, 3.0, 3.5, 4.0, 4.5, 5.0, 5.5 and
`Hamycin in the medium the plates were
`6 . 0 mcg/ml). After allowing diffusion o f
`incubated at
`37° for
`24 hours. After
`Hamycin in the medium the plates were
`incubation period zones of inhibition Were
`incubated at 37° for 24 hours. After
`recorded and a standard graph was plotted
`incubation period zones of inhibition were
`recorded and a standard graph was plotted
`
`Tempe- Approx.
`Tempe- Appro*,
`rature
`percen-
`rature
`percen-
`in DC)
`tage
`relative
`in 0C
`tage
`humidity
`relative
`——-._.__._.__.____________
`humidity
`l.
`Nacl + KN03 + Na—
`N03
`1. Nad + KNOg -j- Na—
`NOs
`CrOB
`.
`.
`.
`.
`.
`.
`2. CrOa
`Na 1 21-120
`3. Na I 2H2O
`Cro3‘
`
`Saturated solutions of salts
`Saturated solutions of salts
`
`2.
`
`3.
`
`4.
`
`5"
`
`5°
`
`37°
`
`37°
`
`30%
`5°
`45%
`5*
`30%
`37°
`45%
`
`5.
`
`so‘1
`
`30%
`370
`
`Na I ango
`..
`CrOs
`5. NailHaO
`50°
`6.
`45%
`CrOs
`50°
`'—-“—--——————_..—__________
`6, CrOs
`
`50«
`Vol. 16. No. 4
`
`Vol. 16. No. 4
`
`Apotex Exhibit 1011
`Page 3 of 11
`
`30%
`
`30%
`45%
`
`30%
`
`•1
`•
`j
`
`I
`
`1
`
`I
`
`i
`
`\
`
`{
`
`)
`
`

`

`STABILITY STUDIES ON HAMYCIN
`
`177
`
`Formulation :
`
` l
`
`i
`
`in capsules
`These mixtures were filled
`177
`STABILITY STUDIES ON HAMYCIN
`which were exposed to 5 ± 1° in polythene
`Each capsule contained antibiotics and
`bags, to study the influence of diluents
`These mixtures were filled in capsules
`diluents in the following proportion by
`themselves on the individual antibiotics.
`Formulation .-
`which were exposed to 5 i 1° in polythene
`weight :
`bags,
`to- study the influence of diluents
`Each capsule contained antibiotics and
`themselves on the individual antibiotics.
`Selection of diluents:
`diluents in the following proportion by
`Quantity
`% of
`Ingredients
`weight :
`ingredient
`__________________...—————-————
`Tetracycline hydrochloride .. 100 mg.
`33.3%
`Ingredients
`Quantity
`3/; of
`ingredient
`4 mg.
`1-33%
`! Hamycin
`Tetracycline hydrochloride .. 100 mg.
`33.3%
`.. 196 mg.
`65.34%
`l Diluent
`4 mg.
`1.33%
`Hamycin
`
`UQtt
`HQ}.
`
`'line
`tinst
`11m.
`
`ma-
`'iine
`
`ma-
`nth
`ilus
`late
`rith
`um
`Hus
`ed,
`late
`tun
`ion
`18
`ion
`:oa
`i8
`am
`ion
`24
`um
`
`24
`
`bi«
`ph
`'g/
`on
`
`he
`
`Q-
`et
`
`•4
`
`65.34%
`.. 196 mg.
`Diluent
`Blending of Antibiotics
`_________,,_______..._._——-—-——-—
`
`i
`
`25.0 gm, of Tetracycline and 1.0 gm. of
`I
`Blending of Antibiotics
`} Hamycin' were thoroughly blended
`by
`25.0 gm. of Tetracycline and 1.0 gm. of
`trituration to have uniform distribution
`)
`Hamycin' were thoroughly
`blended
`by
`in each other. This mixture was employed
`trituration to have uniform distribution
`for mixing with the following diluents in
`in each other. This mixture was employed
`order to study the stability of both antibio­
`for mixing with the following diluents in
`tics in the presence of diluents.
`order to study the stability of both antibio-
`tics in the presence of diluents.
`1. Ascorbic acid
`2. Potassium dihydrogen phosphate
`l. Ascorbic acid
`2. Potassium dihydrogen phosphate
`3. Mannitol
`3. Mannitol
`4. Kaolin
`4. Kaolin
`5. Talc
`Taic
`6. Light magnesium oxide
`Light magnesium oxide
`7. Lactose
`Lactose
`8. Starch
`Starch
`9. Sodium bisulphite.
`Sodium bisulphite.
`
`399°.‘49‘P‘
`
`Mixing of Antibiotics with Diluents :
`Mixing of Antibiotics with Diluems .'
`65. 34% w/w of each diluent was thorou—
`65.34% w/w of each diluent was thorou­
`ghly mixed with 34.66%J w/w mixture of
`ghly mixed with 34.66% w/w mixture of
`tetracycline ”hydrochloride and hamycin.
`tetracycline hydrochloride and hamycin.
`The resultant blended mixture was filled
`The resultant blended mixture was filled
`in capsules using hand operated capsule
`in capsules using hand operated capsule
`filling machine.
`filling machine.
`Blanks .-
`Blanks:
`Hamycin (4.0 mg) with each selected
`diluents 296 mg. constituted hamycin Blank.
`Hamycin (4.0 mg) with each selected
`Similarly
`tetracycline hydrochloride
`100
`diluents 296 mg. constituted hamycin Blank.
`mg with each selected diluents (200 mg)
`Similarly
`tetracycline hydrochloride 100
`constituted
`etracycline
`hydrochloride
`mg with each selected diluents (200 mg)
`Blank.
`etracycline
`hydrochloride
`constituted
`Blank.
`May, 1974
`
`:ke'
`
`May, 1974
`
`ttfci
`
`tetracycline hydrochlo­
`The antibiotics
`Selection of diluents .'
`ride (100 mg) and Hamycin (4 nag) were
`mixed with following diluents (196 mg) to
`The antibiotics
`tetracycline hydrochlo-
`have an uniform blending.
`ride (100 mg) and Hamycin (4 mg) were
`mixed with following diluents (196 mg) to
`have an uniform blending.
`L Ascorbic acid
`2. Potassium dihydrogen phosphate
`Ascorbic acid
`3. Mannitol
`Potassium dihydrogen phosphate
`4. Kaolin
`Mannitol
`5. Talc
`Kaolin
`6. Light Magnesium Oxide
`Talc
`7. Lactose
`Light Magnesium Oxide
`8. Starch
`Lactose
`9. Sodium bisulphite
`Starch
`10. Calcium Carbonate
`Sodium bisulphite
`10. Calcium Carbonate
`11. Stearic acid
`ll. Stearic acid
`12. Magnesium Stearate
`12. Magnesium Stearate
`13. Citric acid
`i3. Citric acid
`14. Methyl Cellulose
`14.
`rMethyl Cellulose
`
`PWFF‘VP‘P’P?
`
`These mixtures were fjlled in capsules
`These mixtures were filled in capsules
`and were exposed to 65 % humidity at 50°
`and were exposed to 65% humidity at 50°
`for a period of 45 days (extreme condition
`for a period of 45 days (extreme condition
`was selected). Observations were made at
`was selected). Observations were made at
`varying intervals and last
`five diluents
`varying intervals and last five
`diluents
`were rejected on the basis of certain reasons.
`were rejected on the basis of certain reasons.
`Remaining nine were employed for
`the
`Remaining nine were employed for the
`investigation.
`investigation.
`
`STABILITY STUDIES ON HAMYCIN
`STABILITY STUDIES ON
`HAMYCIN
`IN COMBINATION WITH TETRACY-
`IN COMBINATION WITH TETRACY­
`CLINE HYDROCHLORIDE WITH SE-
`CLINE HYDROCHLORIDE WITH SE­
`LECTED DILUENTS
`LECTED DILUENTS
`(i) Procedure of
`testing the capsules
`contents for potency. of Hamycin.
`(0 Procedure of testing the capsules
`contents for potency of Hamycin.
`The capsules containing antibiotics and
`diiuents, prepared as described earlier were
`The capsules containing antibiotics and
`exposed to varying temperatures (5°. 37°,
`diluents, prepared as described earlier were
`50°, and room temperature)
`at
`selected
`exposed to varying temperatures (5°, 37°,
`humidities (30% and 45% RH).
`50°, and room
`temperature) at selected
`humidities (30% and 45% RH).
`
`Apotex Exhibit 1011
`Page 4 of 11
`
`

`

`178
`
`HINDUSTAN ANTIBIOTICS BULLETIN
`
`*
`
`
`
` l'
`
`storage at an interval of 3, 6, 9, 12, 15,
`initial content of Hamycin was
`The
`HINDUSTAN ANTIBIOTICS BULLETIN
`I78
`30, 45 and 60 days along with blank.
`estimated raicrobiologically
`immediately
`Standard Graph (Fig. I) was employed
`after preparation of capsules and further
`The
`initial
`content of Harnycin was
`for estimating potency of antibiotic.
`after exposing to selected conditions of
`storage at an interval of 3, 6, 9,
`12,
`15,
`estimated microbiologically
`immediately
`ad
`30. 45 and 60 days along with blank
`after preparation of capsules and further
`Standard Graph (Fig.
`l) was
`employed
`after exposing to selected conditions of
`for estimating potency of antibiotic.
`0.5.
`0.6.
`0.4.
`0-5
`
`0.4
`
`(3.
`ex
`
`(3.
`CK
`
`fo
`in
`gr
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`1.1.
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`1.0.
`
`5.9
`10
`
`11
`
`f
`‘14-
`15
`16
`T
`17 18
`T
`.20 .21 2a :25
`.24 25 25 at
`19
`19 Z0 ai $3. £3 24 55 3L6 &
`12 13 14 15 16 17 18
`ZONE 0F INHIBITION IN mm.
`ZONE OF imiBirm IN mm.
`. Standard graph of hamycin capsules with tetracycline He! at
`_
`nd selected diluents. Log Cone. meg/ml
`versus zone of inhibition m min (common for all the diluents).
`Fig. I. Standard graph of hamycin capsules with tetracycline Hcl and selected diluents. Log Cone, incg/nil
`versus zone of mhibition in mm (commoTi for all the diluents).
`Capsules content equivalent to 2 mg. of
`Initial content was considered as 100%
`Hamycin was exactly weighed and was
`Capsules content equivalent to 2 mg. of
`Initial content was considered as 100%
`retention (no initial destruction has been
`dissolved or suspended in sterile distilled
`observed). Percentage retention of Harm-
`Hamycin was exactly weighed and was
`retention (no initial destruction has been
`water of pH 8. Dilutions were filled in
`cin at selected conditions and time intervals
`dissolved or suspended in sterile distilled
`observed). Percentage retention of Hamy­
`triplicate along with their respective blanks
`were ascertained and recorded.
`water of pH 8. Dilutions were filled
`in
`cin at selected conditions and time intervals
`in the cups. After observing the standard
`The procedure for calculating Haif Life Of
`triplicate along with their respective blanks
`were ascertained and recorded.
`conditions of incubations,
`the diameter of
`Hamycin capsules formulations—
`in the cups. After observing the standard
`zone
`inhibition
`for
`each
`dilution was
`The procedure for calculating Half Life of
`Log of percentage retention of hamycin
`measured and recorded.
`conditions of incubations, the diameter of
`Hamycin capsules formulations—
`were plotted against storage time in days
`zone
`inhibition for each dilution was
`Log of percentage retention of hamycin
`measured and recorded.
`were plotted against storage time in days
`
`
`
`Apotex Exhibit 1011
`Page 5 of 11
`
`999ELL)“
`
`1.06CONS.OFHAMVCINmflgm/mti3SL-SE;E
`
`
`
`
`
`s
`0.2.
`£
`^ O . U
`Z
`^ 0.0,
`‘53.0
`g 1.9.
`£
`5 f.8.
`u.
`0 17.
`1 1.6.
`o
`15.
`^5
`O
`^ 1.4.
`U .
`:e-.-*83“
`1.2
`
`

`

`
`
`STABILITY STUDIES ON HAMYCIN
`
`179
`
`5,
`c.
`d
`
`; |
`
`Mill.“
`
`I
`
`aL
`
`0.693
`
`179
`
`T/2
`
`(3, 6, 9, 12, 15, 30, 45 and 60) at all selected
`smnlu'rv STUDIES ON HAMYCIN
`experimental conditions, viz.
`5±r and 30% relative humidity
`(3, 6, 9, 12, 15, 30, 45 and 60) at all selected
`5± 1° and 45% relative humidity
`experimental conditions, viz.
`Room temperature (R.T.).
`5il° and 30% relative humidity
`37° and 30% relative humidity
`Sil" and 45% relative humidity
`37° and 45% relative humidity
`Room temperature (R.T.).
`50° and 30% relative humidity
`37° and 30% relative humidity
`50° and 45% relative humidity
`37° and 45%1 relative humidity
`50° and 30% relative humidity
`The rate of decomposition of Hamycin
`50° and 45% relative humidity
`followed first
`order reaction which was
`The rate of decomposition of Hamycin
`Only one
`indicated by straight line graphs,
`followed first order
`reaction which was
`graph (Fig.2) of starch as a diluent is given.
`indicated by straight line graphs. Only one
`Other diluents followed the same pattern.
`graph (Fig.2) of starch as a diluent is given.
`From these graphs slope value' for each
`Other diluents followed the same pattern.
`storage condition was calculated, followed
`From these graphs slope value‘ for each
`by K values.
`storage condition was calculated, followed
`(— k = — slope X 2.303)
`by K values.
`Half life in each case was calculated em-
`(4- k : -- slope >< 2.303)
`Half life in each case was calculated em-
`ploying the formula.
`ploying the formula.
`2.0
`
`3.0
`1,9.
`1.9
`
`k
`0.693
`where T/2 = Half life in days. These re­
`T/2 r— —--——
`sults are recorded and compared in Table I.
`k
`STABILITY STUDIES ON TETRACY­
`where 172 = Half life in days. These re-
`sults are recorded and compared in Table I.
`CLINE HYDROCHLORIDE WITH HA­
`MYCIN WITH SELECTED DILUENTS.
`STABILITY STUDIES ON TETRACY-
`CLINE HYDROCHLORIDE WITH HA-
`( / / ) P r o c e d u r e o f t e s t i n g t h e c a p s u l e c o n t e n t
`MYCIN WITH SELECTED DILUENTS.
`for potency of tetracycline hydrochloride.
`(ii) Procedure of testing the capsule content
`Capsules were prepared and exposed to
`for potency of tetracycline hydrochloride.
`earlier mentioned experimental conditions.
`Capsules were prepared and exposed to
`The initial content of tetracycline hydro­
`earlier mentioned experimental conditions.
`chloride was estimated microbiologically
`The initial content of tetracycline hydro-
`immediately after preparation of capsules
`chloride was estimated microbiologically
`and further after exposing to selected condi­
`immediately after preparation of capsules
`tions of storage at an interval of 3, 6, 9, 12,
`and further after exposing to selected condi-
`15, 30, 45 and 60 days along with blank.
`tions of storage at an interval of 3, 6, 9, 12,
`Standard graphs (Fig. 3) were employed
`15, 30, 45 and 60 days along with blank.
`for estimating potency of antibiotic.
`Standard graphs (Fig. 3) were employed
`for estimating potency of antibiotic.
`,— S'ldOlRH
`p» 57502, RH
`-5m/45y.RH
`r
`- 5 745% RH
`
`2; - Room TmpfRt)
`- Roam Temp (RT)-
`-37730% KM
`-37°/30% RH
`1 , 8 .
`1.8
`
`"^-SO'lZQtRH
`~ 507.307. RH
`i
`
`U .
`1.7
`L--3/745 7«/?W
`- 57745 7.1m
`
`l
`
`
`1. 6 .
`1.6
`1.5
`5 1.5.
`5 1.4,
`1.4
`K
`
`UJ
`C* 1.3.
`
`2“- it:
`/.a.
`u. o
`
`
`M.
`— - 507451, av
`
`i“ C)
`-j i . Q .
`-- 50745% RH
`
`{.060FZRETENTION ia
`.0990010:6
`
`0 . 9 .
`
`O.&.
`
`0 . 7 .
`
`
`
`10
`20
`50
`40
`5O
`60
`
`0.6
`TIME IN DAYS
`T
`0
`10
`30
`60
`40
`50
`Fig. 2. Log of percentage retention of hamycin versus time in days in capSules of different conditions of
`storage with tetracycline hydrochlonde and starch as a diluent.
`T I M E J N d A Y S
`Fig. 2. Log of percentage retention of hamycin versus time in days in capsules of different conditions of
`storage with tetracycline hydrochloride and starch as a diluent.
`
`so
`
`Apotex Exhibit 1011
`Page 6 of 11
`
`

`

`HINDUSTAN ANTIBIOTICS BULLETIN
`
`HINDUSTAN ANTIBIOTICS BULLETIN
`
`1
`
`t
`
`]
`
`1
`
`2
`
`
`
`180
`
`180
`
`31
`30.
`29.
`38.
`£ 7 .
`
`£ 2 6 .
`5
`2; 3,5.
`P09»:
`2: o
`S9 23.
`3:
`5 aa.
`LL o a* J
`Uj
`p ao.
`Nl
`
`ZONE0FINHIBITIONINmm.
`
`1 9 .
`
`soc»‘00:):
`must:#01me
`a33:5Pa;323’3::
`
`1 8 .
`1 7 .
`1 6 .
`--L U!.
`1 5 .
`1 4
`r
`1.2, 4.3 4.4
`0.2 05 0.4 0.5 0.5 0.7 as 0.9 {.0 {:1
`r
`i
`i
`f
`i
`t
`1
`T
`0.2. 03 0.4 0.5 0.6 0.7 0-8 0.9 1.0 i:i
`f.& 1.3 14
`LOG COHC. meg I ml Of TETZACYCLINE HCL
`
`14
`
`Fig. 3. Standard graphs of tetracycline capsules with hamycin and selected diluents. Log conc. meg per
`ml Versus zone of inhibition in mm. (1) for potassium dihydrogen phosphate, talc, lactose, sodium
`bisulfite (2) for ascorbic acid, mannitol, kaolin, light magnesium oxide and starch.
`- V01. 16. No 4.
`
`
`
`• Vol. 16. No 4.
`
`Apotex Exhibit 1011
`Page 7 of 11
`
`1
`
`e
`
`N
`
`

`

`STABILITY STUDIES ON HAMYCIN
`
`181
`
`.
`
`days (3, 6, 9, 12, 15, 30, 45 and 60) at all
`Capsule content equivalent to 50 mg. of
`STABILITY STUDIES ON HAMYCIN
`181
`selected conditions.
`tetracycline hydrochloride was exactly
`The rate of decomposition of Tetracy­
`weighed and was dissolved or suspened in
`days (3, 6, 9, 12, 15, 30, 4S and 60) at all
`Capsule content equivalent to 50 mg. of
`cline hydrochloride followed first order
`sterile buffer of pH 4.5. Dilutions were
`selected conditions.
`tetracycline
`hydrochloride was
`exactly
`reaction which was indicated by straight
`filled in triplicate along with their respective
`The rate of decomposition of Tetracy-
`weighed and was dissolved or suspened in
`line graphs. Only one graph of Lactose as
`blanks in the cups. After observing the
`cline hydrochloride followed first order
`sterile buffer of pH 4.5. Dilutions were
`diluent (Fig. 4)
`is given. Other diluents
`standard conditions of incubation the dia­
`reaction which was indicated by straight
`filled in triplicate along with their respective
`followed the same pattern.
`meter of zone of inhibition for each dilution
`line graphs. Only one graph of Lactose as
`blanks in the cups. After observing the
`From these graphs, slope value for each
`was measured and recorded.
`diluent
`(Fig. 4)
`is given. Other diluents
`standard conditions of incubation the dia-
`storage condition was calculated, followed
`followed the same pattern.
`meter of zone of inhibition for each dilution
`From these graphs, slope value for each
`The procedure for calculating half life of
`by K values. Finally employing K values
`was measured and recorded.
`storage condition was calculated, followed
`half life of the antibiotic was calculated as
`Tetracycline in capsule formulation.
`by K values.
`Finally employing K values
`The procedure for calculating half life of
`Log of percentage retention of
`tetracy­
`shown in case of Hamycin. The results
`half life of the antibiotic was calculated as
`Tetracycline in capsule formulation.
`cline was plotted against storage time in
`are recorded and compared in Table 11.
`shown in case of Hamycin. The results
`Log of percentage retention of
`tetracy-
`*,0
`are recorded and compared in Table II.
`cline was plotted against storage time in
`
`2,0 .
`
`
`
`‘3‘
`.
`
`l’
`
`t
`i
`
`9‘.
`
`
`[.060F%RETENTION ;
`
`f . 9 .
`1.9
`
`1.8.
`
`o
`K:
`
`Uj
`
`iij -
`
`u. o
`ta o
`
`- - B l a n k
`
`- - 50/ 3 0 % R H
`- - P / 4 5 % R H
`
`--/rr
`- - 577507. RH
`- - 37730% RH
`
`1.7
`1.7.
`
`
`— — 577457, RH
`- - 37e/45% RH
`_ .. 50°/50% RH
`
`50*130% RH
`— — ctr/45% RH
`
`> - - 5 Q ' / 4 5 % R H
`4O
`50
`60
`20
`60 '“
`f0
`T
`t
`?
`0
`60
`10
`HO
`3 0
`4 0
`5 0
`TIME iN DAYS
`TIME IN DAY3
`Fig. 4. Log of percentage retention of tetracycline hydrochloride versus time in days.
`different conditions of storage with Hamycin and lactose as a diluent
`In capsules at
`Fig. 4. Log of percentage retention of tetracycline hydrochloride versus time in days,
`different conditions of storage with Hamycin and lactose as a diluent
`CONCLUSION
`Hamycin,
`in Refrigerator at 30% RH.
`mannitol,
`lactose and starch showed the
`Hamycin, in Refrigerator at 30 % R.H.
`same half-life of 1505 days but at 45 °,{, R.H.
`mannitol, lactose and starch showed the
`mannitol is found best followed by starch
`same half-life of 1505 days but at 45% R.H.
`mannitol is found best followed by starch
`
`1.6
`1.6
`
`0
`
`In capsules at
`
`CONCLUSION
`Perusal at the grading given in Table I for
`each diluent with reference to half life of
`Perusal at the grading given in Table I for
`each diluent with reference to half life of
`May, 1974
`
`May, 1974
`
`Apotex Exhibit 1011
`Page 8 of 11
`
`

`

`m
`
`3. Mannitol
`
`BS
`
`4. Kaolin
`
`B
`
`5. Talc
`
`“—04.55%nomzoEDonm.
`
`Sr.
`No.
`
`Diluents
`
`T/2 of
`Hamy-
`cinin
`diluents
`after 60
`dziys flt
`50±r
`Blank
`
`only
`
`At 5°
`
`Room Temp. At I T
`
`30% R.H.
`
`45% R.H.
`
`T/2 G
`
`T/2 G
`
`T/2
`
`G
`
`T/2 G
`
`1. Ascorbic Acid
`2. Potassium dihydrogen
`
`phosphate
`
`.. 1777.0 978.1 5
`
`677.2
`
`6
`
`623.00 4
`
`238.5 6
`
`.. 0758.0 256.6
`
`9
`
`144.4 9
`
`84.26
`
`9
`
`39.1 9
`
`1777.0 1505.0 1
`
`1204.0
`
`1
`
`872.8
`
`2
`
`300.9 3
`
`0995.0 967.4 6
`
`871.2 4
`
`406.3
`
`6
`
`270.0
`
`4
`
`.. 0995.0 1053.0 4
`
`862.4
`
`an
`
`6. Light magnesium oxide .. 0995.0 371.1 8
`
`248.3
`
`5
`
`8
`
`571.9 5
`
`270.9
`
`5
`
`139.5
`
`8
`
`106.0 7
`
`TS
`
`7. Lactose
`
`8. Starch
`
`1777.0 1505.0
`
`3
`
`902.0 3
`
`902.8
`
`1
`
`339.8
`
`2
`
`1777.0 1505.0
`
`2
`
`1053.0 2
`
`662.1
`
`3
`
`601.9 1
`
`9. Sodium-bi-su!phite
`
`.. 0995.0
`
`600.6
`
`7
`
`271.0
`
`7
`
`248.3
`
`7
`
`40.61
`
`8
`
`<
`o
`f""
`
`Z
`Q
`
`132
`
`It
`
`s
`
`% R = Percent Retention
`T/2 = Half life in days
`
`G
`RH = Relative Humidity.
`
`= Grade at particular experimental condition
`
`TABLE n—COMPARISON OF HALF LIFE OF TETRACYCLINE HYDROCHLORIDE AT
`
`_x...._:.u...,..2
`
`
`
`mN3.2w92.m2.:n3.2w:2m23..m:2938..”285385232235
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`
`
`
`64.71
`
`4
`
`145.8
`
`3
`
`mw3.2v3;;n8.2v22”928¢2.~45a:323%..5.22.v
`
`82.69 3
`
`144.4 4
`
`127.8 5
`
`25.59 6
`
`
`
`21.07 8
`
`22.57 9
`
`
`
`3303s22.2N:22322:8m:3n9.22N2822::22%.mmm3.2222.a33N:2H:8m225m2822R:
`
`
`
`57.18
`
`5
`
`73.9 6
`
`mmRdn:2w2.8mn.8,”Nwas_$.322232SE3:532.m
`
`.NH.m8.2aSean2.3w3mmw8.5w”.5n2.2m2::353.82.2
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`196.6
`
`2
`
`156.5 1
`
`
`53333wm«.3m2.2m8.5.m2%a3.:a3.:m.:322.5:23982W59:35“...
`
`
`
`129.4 2
`
`270.9 1
`
`if
`
`
`
`o”a.0mi0NP03.0«a..0NF0NF03%.gamma“.
`
`252
`
`18.8 9
`
`33.78 7
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`9.78
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`7
`
`3:0
`
`8.52%..
`
`
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`
`VARYING CONDITIONS OF STORAGE
`
`-- E^jj-rrf
`
`'•
`
`
`
`Eru<>b4NGESEUOMG>Emzflomugmh.nous3nozagmonI—H35C.
`
`TABLE I—COMPARISON OF HALF LIFE OF H^MCYIN AT VARYING CONDITIONS OF STORAGE
`
`1'ill.
`
`
`
`$235230235MRmy.4
`
`At 50°
`
`
`
`45% R.H.
`
`30% R.H.
`
`45% R.H.
`
`
`
`cchaou5585333332mm2“.3:30HUM.
`
`30% R.H.
`
`£35oh:mamHNE.6.
`
`isW
`
`
`
`5:3quEvoke...“Hd..—XH.
`
`T/2 G
`
`T/2 G
`
`T/2 G
`
`
`
`26.59
`
`7
`
`24.07 8
`
`
`
`na:h2.2a”.2m5.2.F2%h9:...”h068238efiagméénfiom.m
`
`10.03
`
`7.32
`
`5
`
`9
`
`9.27
`
`8
`
`10.27 4
`
`10.03 6
`
`12.04 2
`
`10.98
`
`3
`
`19.55 1
`
`oo
`to
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`s z
`o
`C 09
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`Z
`
`Apotex Exhibit 1011
`Page90f11
`
`
`
`

`

`agfc.v
`
`•<
`
`£ P3
`
`[83
`
`--J
`4^
`
`Sr.
`No.
`
`Diluents
`
`At 5° ± 1°
`T/2 of Tetra-
`cycline at in
`diluent only
`after 60 days