NGHIEN ciru KHOA HQC
DjNH Ll/CfNGOONGTHCri ACETAMINOPHEN, DEXTROMETHORPHAN HYDROBROMID VA PHENYLEPHRIN HYDROCLORID TRONG VIEN NANG MEM
RHUMENOL DAY XO
B A N G P H I / O N GPHAP HPLC
LE THI THIEN HlfCfNG, PHAN THI THUY CHI Vien Kiem nghiem thudc Trung trong
l.Datvande
Vien nang mfim RHUMENOL DAY XO do cong ty SOFTGEL HEALTHCARE PVT LIMITED san xuat CO cong thiic bao che nhu sau:
Acetaminophen 325,0 mg Dextromethorphan hydrobromid 10,0 mg Phenylephrin hydroclorid 5,0 mg Ta dugc vira du 1 vien Acetaminophen giam dau tam thai, Dextromethorphan hydrobromid giam ho khan, Phenylephrin hydroclorid dieu tri ngat mui, viem xoang. Dang thuoc ket hop ba hoat chat nay su dung vao ban ngay de dieu tri tam thai cac tri6u chiing cua c^m ciim: ho, nghet mui, chay nu6c mui, hat hai, nhiic dau va cac dau nhiic khac. De dinh lucmg ba hoat chat tren dam bao do chinh xac, tin cay, tien hanh dan gian va nhanh chong, chiing toi giai thieu ket qua nghien ciiu dinh lugmg dugc dong thai Acetaminophen, Dextromethorphan hydrobromid va Phenylephrin hydroclorid trong vien nang mem RHUMENOL DAY XO bang phuang phap HPLC.
2. Thfrc nghiem vd kit qud
2.1. Thiit bi, dung cfi, hod chat, chat chudn vd doi tugfng nghien cum
2.1.1. Thiet hi, dung cu
Da dugc hieu chudn theo ISO/IEC 17025.
May sic ky long hieu nang cao Agilent Technologies vai detector DAD.
- Cgt Purospher Star RP18e (250 x 4 mm; 5 ^m).
- Can phan tich Mettler Toledo AB 204 do chinh xac 0,1 mg.
Bg Igc dung cho sic ky vai mang Igc 0,45 nm.
- Cac dyng cu thuy tinh chinh xac: binh dinh miic, pipet.
2.1.2. Hod chdt vd ehdt chudn
Cac chat chuan cua Vien Kiem nghiem thuoc Trung uang: Acetaminophen, Dextromethorphan hydrobromid va Phenylephrin hydroclorid;
- Acetonitril loai dung cho HPLC;
- Kali dihydrophosphat loai PA;
- Acid phosphoric loai PA.
2.L3. Doi tuang nghien cm
Vien nang rakm RHUMENOL DAY XO (Acetaminophen 325 mg, Dextromethorphan hydrobromid 10 mg va Phenylephrin hydroclorid 5 mg). Cong ty san xuk: SOFTGEL HEALTHCARE PVT LIMITED.
2.2. Phuofngphdp nghien cuu 2.2.1. Diiu kien HPLC
Pha dong: Dung dich kali dihydrophosphat 5,4 g/1 dugc dieu chinh den pH 3,0 bang acid phosphoric - acetonitril (60:40).
- Cot: Purospher Star RP18e (250 x 4 mm; 5 nm).
Detector UV: 280 nm.
Toe do dong: 1,0 ml/phut.
The tich tiem: 20 |al.
2.2.2. Phuang phdp xir ly mdu
Dung moi pha mdu: Nuac - Acetonitril (60:40) Dung dich chudn: Can chinh xac mot lugng Acetaminophen chuan, Dextromethorphan hydrobromid chuan va Phenylephrin hydroclorid chuan hoa tan trong dung moi pha mau; pha loang timg buac neu can de dugc dung dich chuan CO nong do Acetaminophen khoang 160 ^g/ml;
Dextromethorphan hydrobromid khoang 100 |ig/ml va Phenylephrin hydroclorid khoang 50 ng/ml. Loc qua mang loc 0,45 i^m.
Dung dich thu cho dinh lupng Dextromethorphan hydrobromid vd Phenylephrin hydroclorid: Can chfnh xac mot lugng che pham tuong iing vai 1 vien nang
T^p chf KIEM NGHIEM THUOC - S6 2.2011; TSp 9.(32) 1
vao binh dinh miic 100 ml, them khoang 50 ml dung moi pha miu, lac sieu am trong 15 phiit dk hoa tan, pha loang den the tich vai cung dxmg moi, lac deu, loc qua gi§y loc. Loc qua mang loc 0,45 [im (dung dich A).
Dung dich thu cho dinh luang Acetaminophen:
Hiit chinh xac 5,0 ml dung dich A vao binh dinh miic 100 ml, them dung moi pha mau den vach, lac deu, loc qua mang Igc 0,45 ^m.
3. Ket qua va ban luan 3.1. Chon ^eu kien sdc ky
Chiing toi da khao sat va lua chon dugc dieu kien sac ky nhu da neu a phan 2.2.1, sic ky do thu dugc cua dung dich chuan va thu the hien a hinh 1 va 2.
Tren sac ky do chiing toi thay pic ciia Acetaminophen, Dextromethorphan hydrobromid va Phenylephrin hydroclorid tach ro rang, pic doi xiing va ggn. Nhu vay viec lua chon cac dieu kien sac ky la phu hgp.
mAU
400- 300 - 200- 100 -
DM>1 A. Sla=280,4Ret=0«(f<UONGlRHUMENOL S
a
t
i « * •
«
t n
A
's s
* 1 g %
1 % 1 1
§
A
2 4 min
Hinh 1. Sdc ky do dinh luang dong thdi Acetaminophen, Dextromethorphan hydrobromid vd
Phenylephrin hydroclorid cua mdu chudn.
Norm.
350 - 3 0 0 - 250 - 200 - 150 - 100 - 5 0 -
DAD1 A, Sig=280,4Re»=ofl(HUONG\RHUMENOL
i
•s
S' &
g
s>
2.1
K±l
s s
•S 1
= }
& 1 1 i -s
« 8
I K
^1 . . . . . . . , 2 4 min
Hinh 2. Sdc ky do dinh lupng Dextromethorphan hydrobromid vd Phenylephrin hydroclorid cua
mdu thic.
mAU . 400 - 300 - 200 - 100-
DAD1 A,SiB=280.4RB*'Bn(hUONGWHUMENOL
,:•
% s
-S a
L
2 4 min
ITinh 3. Sdc ky do dinh luang Acetaminophen ciia mdu thit.
3.2. Ddnh gid phuang phdp 3.2.1. Khao sdt tinh ddc hieu
Dua vao thai gian luu va pho UV cua cac pic thu dugc: K§t qua cho thay dung dich thu cho cac pic CO thai gian luu va pho UV tuang iing vai cac pic Acetaminophen, Dextromethorphan hydrobromid va Phenylephrin hydroclorid cua dung dich chuan.
3.2.2. Khao sdt tinh thich hap ciia he thong sdc ky Tinh thich hgp cua he thong sac ky dugc xac dinh bang each tien hanh tiem lap lai 6 lan dung dich chuan co nong do Acetaminophen khoang 160 ng/ml;
Dextromethorphan hydrobromid khoang 100 |ag/ml va nong do Phenylephrin hydroclorid khoang 50 ng/ml.
Ghi lai cac gia tri ve thoi gian lim, dien tich pic, do can xiing cua pic va do phan giai gifta cac pic.
Ket qua khao sat tinh thich hgp cua he thong sac ky dugc trinh bay a bang 1.
Bdng 1. Ket qud khao sdt tinh thich hap cua he thong sdc ky
TT
1
2
3
4
5
Thong so
Thoi gian liru (phut) RSD cua thai gian Imi (%) RSD cua dien tich pic (%) He so bat doi Do phan giai voi acetaminophen
Acetaminophen
3,03
0,12
0,25
1,25
Dextromethorphan hydrobromid
4,58
0,11
0,32
1,54
8,42
Phenyleplirm hydroclorid
2,19
0,18
0,29
1,05
5^2
Nhdn xet: Cac s6 lieu thu dugc cho thSy he thong sac ky tren phu hgp cho viec phan tich dinh tinh va djnh lugng dong thai Acetaminophen, Dextromethorphan hydrobromid va Phenylephrin hydroclorid.
Tap chi KIEM NGHIEM THUOC - So 2.2011; TSp 9.(32)
3.2.3. Khao sdt khoang tuyen tinh
Khao sat tren 5 dung dich chuin hon hgp vai 5 ndng do khac nhau, kk qua dugc ghi lai trong bang 2, 3, 4.
Bdng 2. Ket qud khao sdt khoang tuyen tinh ciia acetaminophen
STT 1 2 3 4 5
Phuang trinh hoi qui
Ndng dg Cng/ml) 40 60 80 160 240
Dien tich pic (mAU.s) 705,22 1067,89 1412,26 2842,14 4232,21
y = 17,646 x +5,0486 r = 0,9999
4500 4000 3500 3000 2500 2000 1500 1000 500 0
t 100 200 300
Bang 3. Kit qud khao sdt khoang tuyen tinh cua Dextromethorphan hydrobromid
STT 1 2 3 4 5 Phuffng trinh
hoi qui
Nong dp (Tig/ml)
50 75 100 150 200
Dien tich pic (mAU.s) 382,07 574,20 760,14 1140,91 1522,30 y= 7,5947 x +2,5371
r = 0,9999
1600.00
1400.00 y = 7.5947X + 2.5371 y/^
1200.00 ^ ^ 1000.00 y ^
800.00 j / ^ 600.00 ^ 400.00 ^ 200.00
0.00
0 SO 100 150 200 250
Bang 4. Kit qud khao sdt khoang tuyen tinh ciia Phenylephrin hydroclorid
STT 1 2 3 4 5
Phucmg trinh hoi qui
Ndng dp (fig/ml) 25 37,5
50 75 100
Difn tich pic (mAU.s) 291,49 437,25 581,00 870,90 1169,69
y = 11,69 X-2,1153 r = 0,9999
1400
'^^^ y = 11.69X - 2.1153 ^ R' = 0.9999 ^/^
1000 >^
800 Z * ^
600 j ^
400 y ' 200
0
0 20 40 60 eo 100 120
Nhdn xet: V6i dieu kien sic ky da lya chon, trong khoang nong do da khao sat, cac chat deu c6 su tuang quan tuyln tinh chat che giua nong do voi dien tich pic dap iing.
2.3.4. Khao sdt dp lap lai
Khao sit tren vien nang mIm RHUMENOL DAY XO vai cac dieu kien sSc ky va each xii ly miu nhu a muc 2.2.1 va 2.2.2, iSp lai thi nghi?m 6 lan, k6t qua dugc trinh bay trong bang 5.
T9P chi KIEM NGHI$M THU6C - So 2.2011; T3p 9.(32)
Bdng 5. Kit qud khao sdt dp lap lai cita phuang phdp
STT
1 2 3 4 5 6
Lupng cdn (g)
1,3086 1,3102 1,3086 1,3095 1,3106 1,3084 Trung binh
RSD (%)
Acetaminophen Dien tich pic
2756,39 2758,31 2754,23 2745,63 2756,68 2754,84
Hdm luang (%)' 97,90 97,85 97,82 97,45 97,76 97,86 97,77 0,16
Dextromethorphan hydrobromid Dien tich pic
685,93 686,59 686,82 686,22 684,72 684,33
Hdm lu(mg (%) 95,75 95,73 95,88 95,73 95,44 95,54 95,68 0,16
Phenylephrin hydroclorid Diin tich pic
493,23 493,11 494,03 494,08 493,32 492,60
Hdm luang (%)' 96,29 96,15 96,45 96,39 96,16 96,18 96,30 0,13
Nhdn xet: Kdt qua tren cho thiy phuang phap co do lap lai t6t (RSD = 0,13% - 0,16%), co the ap dung d6 dinh lugng cac mau thanh pham.
2.3.5. Khao sdt dp diing
Do diing dugc xac dmh bang phuang phap them ( da biet truac ham lugng, lugng hoat chat them vao i thu. Tien hanh tren 6 mau thii rieng biet theo muc 2.'.
[ chuan. Them mot lugng chinh xac cac chat chuan vao mau thu I tuong ling khoang 20% lugng cua m6i hoat chat co trong miu 1.2. Ket qua dugc the hien trong bang 6.
Bdng 6. Kit qud khao sdt dp diing cua phuang phdp khi them 20% luang chdt chudn
STT
1 2 3 4 5 6
Luang can(g)
1,3034 1,3062 1,3025 1,3084 1,3073 1,3107 TB (%) RSD (%)
Acetaminophen Luang
them vdo (mg)
65,0 65,0 65,0 65,0 65,0 65,0
Luang tim lai (mg) 64,747 64,493 64,753 64,929 64,454 65,423
%rim lai 99,6 99,2 99,6 99,9 99,2 100,7
99,7 0,54
Dextromethorphan hydrobromid Luang
them vdo (mg)
2,0 2,0 2,0 2,0 2,0 2,0
Luang tim lai
(i»g) 1,998 2,004 2,000 1,993 1,996 1,992
%rim lai 99,9 100,2 100,0 99,6 99,8 99,6 99,8 0,23
Phenylephrin hydroclorid Luang
them vdo (mg)
1,0 1,0 1,0 1,0 1,0 1,0
LufTtg tim lai (mg) 0,993 0,997 1,002 1,001 0,996 0,992
%rini lai 99,3 99,7 100,2 100,1 99,6 99,2 99,7 0,40
Nhdn xet: Ket qua khao sat cho thay phuang phap co do diing cao (Lugng tim lai: 99,7% - 99,8%), dam bao tot cho viec dinh lugng dong thai Acetaminophen, Dextromethorphan hydrobromid va Phenylephrin hydroclorid trong cac che pham vien nang.
2.3.6. Khao sdt gidi han dinh luong (LOQ)
Pha loang din dung dich chu§n va dung dich thu d8n ndng do thdp nhdt va ti8n hanh sSc ky theo muc 2.2.1. Giai han dinh lugng dugc xac dinh tai n6ng do ma 6 do tin hieu cua may cho dap iing gip 10 lin so voi nhiiu duong nhn va co do chinh xac chip nhan dugc. Ket qua, giai han dinh lugng la khoang 3 |ig/ml voi Acetaminophen; 10 ^g/ml vai Dextromethorphan hydrobromid va 5 |ig/ml d6i vai Phenylephrin hydroclorid.
4. Ket luan
Cac ket qua thuc nghiem cho thiy chuong trinh HPLC ma chiing toi xay dung a tren cho phep djnh lucmg dugc ddng thai Acetaminophen, Dextromethorphan hydrobromid va Phenylephrin hydroclorid trong Vien nang mem RHUMENOL DAY XO voi do chinh xac cao, do diing tdt, khoang tuySn tinh rdng vai do tuomg quan tuyen tinh chat che giiia ndng do va dien tich pic trong khoang ndng do da khao sat, tiSn hanh dan gian va nhanh chong. Chiing tdi hi vgng co th8 ap dung phuang phap nay dh dinh lugng ddng thai Acetaminophen, Dextromethorphan hydrobromid va Phenylephrin hydroclorid trong cac ch6 phim tuong tu.
Tap chi KIEM NGHIEM THUOC - So 2.2011; T^p 9.(32)
Tai lieu tham khao 1. Dugc diSn Viet Nam, Lin xuit ban thii tu, nam 2009.
2. BR 2009.
3. USR32.
4. Sandie Lindsay, High Performance Liquid Chromatography, 2nd edition, 1994.
SUMMARY
An HPLC method for simultaneous determination of Acetaminophen, Dextromethorphan hydrobromide and Phenylephrine hydrochloride in RHUMENOL DAYXO soft capsule is introduced:
The chromatographic conditions are as follows:
Mobile phase: 5.4 g/l Potassium dihydrogen phosphate solution (adjusted to pH 3.0) acetonitrile (60:40). "^ ^.,
- Detector UV: 280 nm.
- Column: Purospher Star RP18e (250x4 mm; 5 ^im).
- Flow rate: 1.0 ml/min.
- Injection volume: 20 pi.
NGHIEN curu DINH Ll/GTNG CONESSIN
CHIETXUATTCTCAYMl/C
H O A T R A N G B A N GPHl/ONG
P H A P SACKY LONG HIEU
N A N GCAO
LUC THI VAN, TRINH VAN LAU. TRAN THI HONG ANH Vien Kiem nghiem thuoc Trung uang
l . D a t v a n d l
Cay M\rc hoa tring (Mdc hoa tring, Thung muc la to) CO ten khoa hgc Holarrhena pubescens (Buch. Ham.) Wall, ex G. Don., Syn. Holarrhena antidysenteria (Roxb. ex Flem.) A. DC, hg Tnic dao (Apocynaceae). Vd than va cac bd phan khac cua cay dugc dung tri ly amip. Vo cung dugc dung trj sdt, ia chay, viem gan. Vd va la dung nau nuac tam ghe. Cd the diing vd re gia giap ngam rugu ciing vdi vd re cay Hde dk bdi chua benh ghe [1], [2 ].
Conessin la mdt alkaloid steroid dugc chiet xuat tu cay Muc hoa trang theo de tai nhanh "Nghien ciiu chiet tach va tinh che kaempferol tir Don la dd va conessin tii Muc hoa trang de lam chat chuan phuc vu kiSm nghiem dugc heu" (Ma sd KC. 10.16/06-10), thudc de tai khoa hgc cap nha nudc "Nghien ciiu phat trien va iing dung cdng nghe phuc vu bao ve, cham sdc va nang cao siic khde cdng ddng" (KC. 10/06-10) [1].
Ngudi ta dung conessin hydroclorid hay hydrobromid de dieu tri ly amip, cd tac dung nhu emetin nhung khdng dgc. Theo mdt sd cdng trinh nghien ciiu khoa hgc, conessin cd tac dung chira Ii amip, la chat ddi khdng histamin, cd tac dung chon loc tren thu the Hj [1],[2].
De phye v\i cdng tac nghiem thu de tai va thiet T9P chi KlfeM NGHIf M THU6C - S6 2.2011; TSp 9.(32)
lap chat chuan, chiing tdi da nghien ciiu dinh lugng conessin (chiet xuat tu cay Muc hoa trang) bang phuang phap sac ky ldng hieu nang cao.
2. Thye nghiem va ket qu^
2,1. Thiet bi, hda chat, chat chuan va mau nghien cmi
2.1.1. Thiet bi vd dung cu
May sac ky ldng cao ap Shimadzu LC 20A Series.
Cot sic ky Phenomenex RP 18 (250 x 4,6 mm;
5 \ivn).
- Can phan tich Mettler Toledo AG 245, do chinh xac 0,01 mg.
Dung cu thuy tinh chinh xac, binh dinh miic, pipet, dng dong...
2.1.2. Hoa chdt, chdt chudn
Methanol tinh khiet dung cho sac ky ldng (hang Merck).
- Natri dihydrophosphat tinh khiet phan tich (PA).
Triethylamin tinh khiet phan tich (PA).
- Acid phosphoric tinh khiet phan tich (PA).
Chat chuan gde conessin (Vien Kiem nghiem thudc Trung uong).
