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ISSN Online: 3007-1941 ISSN P in : 3007-1933
IN-VITRO EVALUATIONS OF LINCOMYCIN HYDROCHLORIDE
CAPSULES: DEVELOPMENT, VALIDATION, AND COMPATIBILITY
A icle De ails
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Keywo ds:
In-Vi o compa a i e e alua ion,
Uni ed S a e Pha macopoeia (U.S.P.),
pha macopeia excipien selec ion
Muhammad Im an
Facul y o Sciences, The Supe io
Uni e si y Laho e, Pakis an.
Ali Im an Mallhi
Facul y o Sciences, he Supe io Uni e si y
Laho e, Pakis an
Muhammad A i * (Co esponding au ho )
Facul y o Sciences, he Supe io Uni e si y
Laho e, Pakis an
Email:
muhammad.a i . sd@supe io .edu.pk
Wajiha Khalid
Facul y o Pha maceu ical Sciences (FPS),
Riphah In e na ional Uni e si y, Islamabad
Rabia Fa idi
Depa men o Plan B eeding and Gene ics,
Uni e si y o Ag icul u e Faisalabad,
38000. Pakis an
Muhammad Rizwan
Facul y o Sciences, he Supe io Uni e si y
Laho e, Pakis an
Ma yam Aslam
Depa men o Chemis y, Go e nmen
College Women Uni e si y Faisalabad
This s udy epo s o pha maceu ical o mula ion de elopmen , analy ical
alida ion, and In-Vi o compa a i e e alua ion o gene ic P oduc
Lincomycin 500mg ha d gela in capsules. The o mula ion was designed
using Lincomycin as he ac i e pha maceu ical ing edien , along wi h
pha macopeia excipien s selec ed based on excipien compa ibili y s udies
conduc ed unde accele a ed condi ions. The inished p oduc me Uni ed
S a e Pha macopoeia (U.S.P.) Speci ica ions o assay, dissolu ion,
uni o mi y o dosage, and wa e con en . Analy ical me hod alida ion was
pe o med using high-pe o mance liquid ch oma og aphy (HPLC), and he
me hod ul illed all ICH and USP equi emen s o sys em sui abili y,
speci ici y, accu acy, p ecision ( epea abili y and in e media e), obus ness,
and spiking eco e y. Compa a i e dissolu ion s udies we e ca ied ou in
0.1N HCl (pH 1.2), ace a e bu e (pH 4.5), and phospha e bu e (pH 6.8)
using USP Appa a us II. The es p oduc demons a ed mo e han 85% d ug
elease wi hin 15 minu es in all media, and exhibi ed simila i y ac o ( 2)
alues g ea e han 50 and di e ence ac o ( 1) alues below 15 when
compa ed wi h he e e ence p oduc No elink 500 mg Capsule. These esul s
con i m he pha maceu ical equi alence o he es o mula ion and suppo
i s sui abili y o local p oduc ion and egula o y app o al.
Muhammad Im an1, Ali Im an Mallhi2, Muhammad A i *3, Wajiha Khalid4, Rabia Fa idi5,
Muhammad Rizwan6, Ma yam Aslam7
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1. INTRODUCTION
Lincomycin HCl is a semisyn he ic de i a i e o he Lecin Base amily ha ing chemical name is me hyl-[7-
chlo o-6,7,8- ideoxy-6- ans-(1-me hyl-4-p opyl-l-2-py ollidin-ca boxamido)-1- hio-l- h eo-α-d-galac o-
oc apy anoside Compendia name is Lincomycin wi h molecula o mula is C₁₈H₃₄N₂O₆S· HCl and molecula
weigh is 443.00 g/mol, syn hesized h ough he chemical modi ica ion o Lincomycin by coupling i wi h L-
lysine and o maldehyde, which enhances i s solubili y and bioa ailabili y [1, 2] [3]. Classi ied as a second-
gene a ion Lincomycin, Lincomycin was de eloped o o e come he limi a ions o ea lie agen s like
Lincomycin and chlo e acycline, pa icula ly in e ms o abso p ion, issue dis ibu ion, and side e ec
p o ile [4]. The second-gene a ion Lincomycin, including clindamycin and, we e in oduced o p o ide
b oade clinical u ili y and imp o e ole abili y [5].
The his o y o Lincomycin begins wi h chlo e acycline (au eomycin), disco e ed in he la e 1940s om
S ep omyces au eo aciens, ma king he ad en o a new e a in an imic obial he apy [6]. Lincomycin quickly
gained p ominence due o hei abili y o inhibi a wide ange o pa hogens. O e ime, s uc u al modi ica ions
led o newe agen s like clindamycin, , and Lincomycin, each wi h imp o ed pha macokine ic and
pha macodynamic cha ac e is ics [7]. These d ugs a e now used no only o in ec ious diseases bu also o
de ma ological applica ions, such as acne and osacea, due o hei dual an imic obial and an i-in lamma o y
ac ions [8]. Mechanis ically, Lincomycin —ac s as a bac e ios a ic agen by inhibi ing bac e ial p o ein
syn hesis. I binds e e sibly o he 30S ibosomal subuni , he eby p e en ing he a achmen o aminoacyl-
RNA o he A si e o he ibosome, e ec i ely hal ing pep ide elonga ion. This in e up ion dis up s bac e ial
eplica ion and g ow h, accoun ing o i s e icacy agains a b oad spec um o g am-posi i e and g am-
nega i e o ganisms[9].
Lincomycin demons a es ac i i y agains se e al clinically ele an pa hogens, including Cu ibac e ium
acnes, Chlamydia achoma is, Mycoplasma pneumoniae, and Ricke sia species [10]. This b oad
an imic obial spec um unde pins i s use in managing condi ions like espi a o y in ec ions, sexually
ansmi ed in ec ions, and de ma ologic diso de s such as acne ulga is[11].
One o he dis inc i e ad an ages o Lincomycin lies in i s pha macokine ics. I exhibi s highe wa e
solubili y compa ed o i s pa en compound, esul ing in mo e e icien o al abso p ion and g ea e
bioa ailabili y [12]. Once inges ed, Lincomycin unde goes apid hyd olysis in he gas oin es inal ac o
elease Lincomycin, which is he pha macologically ac i e moie y, con i ming i s ole as a p od ug [13].
Plasma concen a ion s udies e eal ha only e acycline is p esen sys emically a e o al adminis a ion o
Lincomycin, suppo ing his me abolic ans o ma ion[14].
Addi ionally, Lincomycin p o ides be e issue pene a ion and has a longe hal -li e han olde Lincomycin,
which allows o once-daily o wice-daily dosing and imp o ed pa ien compliance [15]. Gas oin es inal
side e ec s such as nausea and abdominal discom o , commonly obse ed wi h Lincomycin, appea o occu
less equen ly wi h Lincomycin, making i mo e sui able o long- e m use[16]. A p ima y he apeu ic
applica ion o Lincomycin is in he ea men o mode a e o mode a ely se e e acne ulga is. Acne
pa hogenesis in ol es ollicula occlusion, sebum o e p oduc ion, C. acnes coloniza ion, and in lamma ion
[17]. Lincomycin helps educe C. acnes le els and exe s an i-in lamma o y e ec s by inhibi ing neu ophil
chemo axis and he elease o in lamma o y cy okines [18]. This dual ac ion enhances i s e ec i eness in
in lamma o y acne, especially when opical ea men s a e insu icien [19].
Beyond acne, Lincomycin has been explo ed in he managemen o hid adeni is suppu a i a (HS)—a ch onic
in lamma o y skin diso de cha ac e ized by ecu en nodules, abscesses, and sinus ac s. Lincomycin,
including Lincomycin, a e among he i s -line sys emic ea men s ecommended in Eu opean S1 guidelines
o mild o mode a e HS [20]. Thei e icacy is a ibu ed o bo h an imic obial e ec s and modula ion o he
immune esponse, al hough high-quali y andomized ials a e s ill needed o s eng hen he e idence base
[21]. Despi e hese bene i s, Lincomycin, like all Lincomycin, ca ies a isk o ad e se e ec s, no ably
pho osensi i i y eac ions. Pho o oxici y a ises when he d ug abso bs ul a iole ligh , esul ing in oxida i e
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damage o he skin [22]. While a e, his eac ion necessi a es p e en i e counseling o pa ien s on sun
exposu e and pho op o ec ion measu es. In e es ingly, Lincomycin appea s o ha e a lowe incidence o such
eac ions compa ed o ea lie Lincomycin, al hough comp ehensi e epidemiological da a a e s ill e ol ing
[23]. Con inued esea ch may u he expand i s applica ion in o a eas such as esis an bac e ial in ec ions
and immune-media ed in lamma o y diso de s.
Figu e 01 Chemical S uc u e o Lincomycin Hyd ochlo ide
2. EXPERIMENTAL
2.1 Ma e ials
The o mula ion de elopmen o Lincomycin Hyd ochlo ide (HCl) ha d gela in capsules was ca ied ou using
aw ma e ials p ocu ed om au hen ic and eliable sou ces. The ac i e pha maceu ical ing edien (API),
Lincomycin Hyd ochlo ide, was ob ained om Pha maceu ical G ade, supplied as pe he Uni ed S a es
Pha macopeia (U.S.P.) and
B i ish Pha macopoeia (B.P), Eu opean Pha macopoeia (Eu . Ph.) s anda ds. A e e ence s anda d o
Lincomycin HCl wi h a ce i ied po ency was also employed o ensu e accu a e quan i a i e analysis and se e
as a p ima y benchma k o analy ical alida ion.
Fo excipien s, Magnesium S ea a e was used as a lub ican o educe in e pa icle ic ion and imp o e
powde low du ing capsule illing, while Ae osil-200 (colloidal silicon dioxide) unc ioned as a glidan o
p e en powde agglome a ion and imp o e uni o mi y o capsule ill weigh . The inal dosage o m was
encapsula ed in emp y gela in capsule shells, size #0, which a e widely a ailable and sui able o
accommoda ing he equi ed ill weigh o he de eloped o mula ion.
All excipien s used we e o pha maceu ical g ade and complied wi h pha macopeial s anda ds. Thei selec ion
was based on p io compa ibili y s udies, li e a u e p ecedence, and es ablished use in o al capsule
o mula ions.
2.2 Me hodology
Fo quali a i e and quan i a i e analysis, high-pu i y analy ical-g ade eagen s and sol en s we e employed.
The key chemicals included hyd ochlo ic acid, pu i ied wa e , sodium ace a e, glacial ace ic acid, monobasic
po assium phospha e, sodium hyd oxide, phospho ic acid, 2-me hyl-2-p opanol, di-po assium hyd ogen
o hophospha e, e abu ylammonium hyd ogen phospha e, sodium ede a e, and sodium me abisul i e. Each
eagen was p ocu ed om ce i ied supplie s, and only analy ical-g ade ma e ials we e used o ensu e
accu acy, ep oducibili y, and compliance wi h pha macopeial speci ica ions.
These eagen s we e p ima ily u ilized o p epa ing mobile phases, bu e s, and solu ions equi ed in high-
pe o mance liquid ch oma og aphy (HPLC), UV spec opho ome ic analysis, dissolu ion s udies, and Ka l
Fische i a ion o mois u e con en analysis. The use o ambe -colo ed olume ic lasks minimized ligh -
induced deg ada ion o Lincomycin HCl du ing solu ion p epa a ion, gi en he known pho osensi i i y o
Lincomycin. The o mula ion and e alua ion s udies we e pe o med using well-calib a ed and alida ed
labo a o y ins umen s. The ollowing equipmen played a c ucial ole:
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Re ige a o – Used o he s o age o e e ence s anda ds, wo king s anda ds, and sensi i e eagen s a
con olled condi ions o p e en deg ada ion.
HPLC sys em – Equipped wi h a s a iona y phase (HPLC Column: 4.6 mm × 25 cm, 5 µm packing, L-7/C8
Type) o ch oma og aphic sepa a ion, quan i ica ion, and pu i y analysis o Lincomycin HCl. The selec ion
o a C8 column ensu ed adequa e e en ion and esolu ion o he analy e unde op imized ch oma og aphic
condi ions. UV-Visible Spec opho ome e – Employed o apid de e mina ion o d ug con en and
calib a ion cu e cons uc ion du ing me hod alida ion.
Dissolu ion Appa a us (USP Type I, 12-baske assembly) – U ilized o in i o elease es ing o
e alua e dissolu ion beha io o o mula ed capsules unde simula ed gas oin es inal condi ions
Ka l Fische Appa a us – Used o he de e mina ion o mois u e con en in aw ma e ials and inal
o mula ions o ensu e compliance wi h pha macopeial limi s.
Glasswa e – High-quali y olume ic lasks (ambe -colo ed), beake s, g adua ed pipe es, and o he
s anda d labo a o y glasswa e we e used o p epa e solu ions wi h high accu acy.
Each ins umen was calib a ed acco ding o s anda d ope a ing p ocedu es (SOPs) be o e use.
2.2.1 P e- o mula ion S udies
Be o e he ac ual o mula ion, p e- o mula ion e alua ions we e conduc ed o s udy he physical and chemical
p ope ies o Lincomycin HCl and i s in e ac ion wi h excipien s. These included:
O ganolep ic e alua ion (appea ance, colo , and odo ) o API
Solubili y p o ile in wa e and a ious bu e s o guide dissolu ion me hod design.
FTIR compa ibili y s udies be ween he API and excipien s (Magnesium S ea a e, Ae osil-200) o con i m he
absence o any chemical in e ac ions ha migh a ec d ug s abili y.
Mic ome i ic p ope ies o he blend (bulk densi y, apped densi y, angle o epose, comp essibili y index,
Hausne ’s a io) o ensu e sui abili y o capsule illing and good low beha io
2.2.2 Fo mula ion De elopmen :
The o mula ion o Lincomycin HCl capsules was de eloped by di ec blending and encapsula ion. The
weighed quan i ies o API and excipien s we e geome ically mixed in a clean, d y s ainless-s eel con aine .
Ae osil-200 was i s blended wi h he d ug o enhance uni o mi y and low, ollowed by addi ion o
magnesium s ea a e as a lub ican in he inal mixing s ep.
The inal homogeneous blend was encapsula ed in o ha d gela in capsule shells, size #0, using a semi-
au oma ic capsule illing machine. Filled capsules we e s o ed in ambe -colo ed con aine s o p o ec om
ligh -induced deg ada ion.
Table# 01 Scale up o Te acycline o mula ion De elopmen :
S , #
RAW MATERIALS
Role o Ing edien s
1
Lincomycin HCl
API
2
Colloidal silicon dioxide
Glidan
3
p egela inized s a ch (co n)
Lub ican
4
S ea ic acid
Dissolu ion Enhance
5
Emp y Gela in Capsule Shell#0 (Da k blue
Cap and Ligh blue body)
To enclose medicine
2.2.3 Pos - o mula ion Assessmen
The de eloped capsules unde wen a ious physicochemical e alua ions:
Weigh Va ia ion Tes : Twen y Capsules we e andomly selec ed and weighed indi idually and collec i ely
o check compliance wi h pha macopeial speci ica ions. Disin eg a ion Tes : Conduc ed in pu i ied wa e a
37 ± 0.5 °C o ensu e ha capsules disin eg a ed wi hin he pha macopeial ime limi . Dissolu ion Tes ing:
Pe o med using USP Dissolu ion Appa a us I (baske me hod) wi h app op ia e dissolu ion media. Samples
we e wi hd awn a de ined in e als and analyzed by HPLC o UV spec opho ome e . D ug Con en
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Uni o mi y: Capsules we e assayed indi idually using alida ed HPLC me hods o ensu e uni o m
dis ibu ion o he d ug. Mois u e Con en : De e mined using Ka l Fische i a ion o con i m s abili y and
p e en deg ada ion due o excessi e wa e con en .
2.2.4 Analy ical Me hod Valida ion:
Quan i a i e es ima ion o Lincomycin HCl in he o mula ed capsules was ca ied ou by HPLC me hod,
op imized wi h a C8 column and a sui able mobile phase p epa ed om The me hod was alida ed in
acco dance wi h ICH guidelines as ollows:
3. Linea i y ac oss a speci ied concen a ion ange
4. Accu acy and P ecision h ough eco e y s udies and epea abili y es ing
5. Speci ici y agains excipien s and po en ial deg ada ion p oduc s
6. Robus ness and Ruggedness by delibe a e a ia ions in me hod pa ame e s
7. Complemen a y UV spec opho ome ic analysis was employed o apid
es ima ion and c oss- e i ica ion o esul s’
3. RESULTS AND DISCUSSION
Quan i y o API was subjec ed o po ency adjus men o in acco dance wi h he API Con en s and some
adjus men o di e ence in quan i y o API a e po ency adjus men we e be deduc ed using Colloidal silicon
dioxide. The esul s gi en as in he able as:
Table 02 Fo mula ion De elopmen
S , #
RAW MATERIALS
Role o Ing edien s
Pe cen age o
Ing edien s/cap.
Scale
mg/capsule
1
Lincomycin HCl
API
98.33%
590.0*
2
Colloidal silicon dioxide
Glidan
1.0%
6.0**
3
p egela inized s a ch (co n)
Lub ican
0.33%
2.000
4
S ea ic acid
Dissolu ion Enhance
0.33%
2.000
Emp y Gela in Capsule
Shell#0 (Da k blue Cap and
Ligh blue body)
To enclose medicine
----
100
To al weigh o Con en s/ capsule
600mg
To al weigh o capsule wi h shell
700mg
The pic o ial iew o he wo king is elabo a ed as:
S ep-1
Figu e-02. Lincomycin (Final Mix)
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S ep-2
Figu e-03 Lincomycin o mula ion de elopmen (Ac i e Raw Ma e ial/ API)
S ep-3
Figu e-04 Lincomycin o mula ion de elopmen (a In e media e/ A e Encapsula ion)
S ep-4
Figu e-05 Lincomycin Finished Pha maceu ical P oduc (A e Packaging) Packed in Alu-P c Foil
3,1 Adjus men and hei ou comes in Analy ical me hod De elopmen (Con en s By HPLC)
In 10.0g o he Sodium hyd oxide pelle s in 90mL olume ic lasks and dilu e up o he ma k wi h wa e .
As a mobile phase, add 8.1 mL o Phospho ic Acid in 600 mL o wa e and adjus wi h ammonium hyd oxide
o a pH o 6.0
Bu e solu ion……………………………….…….505.5 mL
Ace oni ile……………………………….……..….97.2 mL
Me hanol… ………………………………………...97.2 mL
Mix, shake well and degas by sonica ion and il e i h ough 0.45 µm memb ane il e .
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Ch oma og aphic Condi ions:
Column : L7/ C8 (4.6mm × 25cm)
De ec o : 210 nm
Flow a e : 1.0 mL / min
Injec ion olume : 20 µl
Column Temp. : 46 ºC
Sepa a ely injec (p o ided 0.45µ disc il e ) equal olumes o s anda d and sample p epa a ions in o he
ch oma og aph and eco d he esponses o he majo peaks. The ela i e s anda d de ia ion o eplica e
injec ions o s anda d p epa a ion should no be mo e han 2.0%, ailing ac o NMT: 1.3. Calcula e he
pe cen age o Lincomycin by compa ison o majo peak esponses o s anda d and sample solu ions
In 2.2mL Hyd ochlo ic Acid (usually 37%) in 50mL wa e , now dilu e up o 500mL wi h wa e and
mix well. A e e ence solu ion o a comme cial b and was also used o his adjus men s.
Table # 04 P oduc Desc ip ion o in- i o s udy bo h e e ence and es p oduc
Desc ip ion
Re e ence / Compa a o P oduc
Tes P oduc
P oduc Name
No elink 500mg Capsule
Lecin 500mg Capsule
Composi ion
Each ha d capsule con ains:
Lincomycin (U.S.P):…… 500 mg
Each ha d Capsule Con ains:
Lincomycin (U.S.P):…… 500 mg
Lo #
NL-3542
T-001
Figu e-08 Re e ence P oduc used o in-Vi o S udy/Bioequi alence
Resul s in ch oma og am a e sequen ially elabo a ed as u he :
Figu e-06 HPLC Repo / Ch oma og am o Lincomycin HCl Re e ence
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Figu e-07 HPLC Repo / Ch oma og am o Lincomycin Tes Sample
The eco e y o Lincomycin be ween 95.0 o 105.0%; he RSD is o Lincomycin 2.0 %. and ailing ac o is
less he 0.7 The p ecision as well as he ep oducibili y o his me hod was sa is ac o y.
Resul s o Compa a i e Dissolu ion P o ile a e as unde :
Figu e-9 G aphically ep esen a ion o CDP o e e ence and es p oduc (pH 1.2)
Figu e-10 G aphically ep esen a ion o CDP o e e ence and es p oduc (pH 4.5)
30.00
80.00
45 60 75
D ug Release (%)
Time Poin s (minu es)
Acidic Medium (pH 1.2)
(Lincomycin)
40.00
50.00
60.00
70.00
45 60 75
D ug Release (%)
Time Poin s (minu es)
Ace a e Bu e (pH 4.5)
(Lincomycin)
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Figu e-11 G aphically ep esen a ion o CDP o e e ence and es p oduc (pH 6.8)
A compa a i e dissolu ion s udy was conduc ed be ween he es p oduc , Lecin 500 mg Capsules, and he
e e ence p oduc , No elink 500 mg Capsules. The di e ence ac o ( 1) was calcula ed o be 2.87%, and he
simila i y ac o ( 2) was 79.27%. These alues all wi hin he accep able egula o y limi s ( 1 < 15 and 2 >
50), indica ing a high deg ee o simila i y be ween he dissolu ion p o iles o he wo p oduc s. Bo h he es
and e e ence o mula ions exhibi ed mo e han 75% d ug elease wi hin 75 minu es in an acidic medium,
which is a key equi emen o immedia e- elease o mula ions. Addi ionally, he dissolu ion beha io was
ound o be compa able in ace a e bu e (pH 4.5) and phospha e bu e (pH 6.8), u he suppo ing he
consis ency o d ug elease ac oss di e en physiological pH condi ions. These indings con i m ha he es
p oduc mee s he egula o y c i e ia o dissolu ion p o ile simila i y, p o iding s ong e idence o in i o
bioequi alence. As such, he da a suppo he conclusion ha Lecin 500 mg Capsules pe o m simila ly o
No elink 500 mg Capsules, and can be conside ed pha maceu ically equi alen in e ms o dissolu ion
beha io .
3.2 Excipien compa ibili y s udy
A Desc ip ion o he D ug-Excipien Samples is as unde :
Table-05 Desc ip ion o Te acycline o mula ion unde he compa ibili y s udy:
S , #
RAW MATERIALS
Role o Ing edien s
Pe cen age o
Ing edien s/cap.
Scale
mg/capsule
1
Lincomycin HCl
API
98.33%
590.0*
2
Colloidal silicon dioxide
Glidan
1.0%
6.0**
3
p egela inized s a ch (co n)
Lub ican
0.33%
2.000
4
s ea ic acid
Dissolu ion Enhance
0.33%
2.000
Emp y Gela in Capsule
shell#0 (Da k blue Cap and
Ligh blue body)
To enclose medicine
----
100
To al weigh o Con en s/ capsule
600mg
To al weigh o capsule wi h shell
700mg
D ug Subs ance-Excipien Compa ibili y S udy:
Samples a e p epa ed o d ug subs ance and o he ing edien s as pe Table#1 a e one mon h a accele a ed
condi ion ( empe a u e: 40°C±2°C, ela i e humidi y 75%±5%).
D ug subs ance compa ibili y is o be assessed h ough HPLC analysis o ing edien s and d ug subs ance in
30.00
35.00
40.00
45.00
45 60 75
D ug Release (%)
Time Poin s (minu es)
Phospha e Bu e (pH 6.8)
(Lincomycin)
