Nghien CLFU - Ky thuat
qua trinh sinh acid lactic cua vi khuan, dong thdi lupng acid lactic sinh ra lam giam pH mdi trudng lgi dupe phan ung vdi MgCOj mdi bo sung vao dd tao s y on djnh pH trong qua trinh nudi d y .
K e t l u a n
Nghien eiru nay n h i m myc dieh tham dd kha nang sinh t i n g hap magnesi lactat bdng each nudi d y Lactobacillus acidophilus ATCC 4356 trong mdi trudng thich hpp cd bo sung MgCOg Da t i l n hdnh dinh tinh magnesi laetal thu d u p c b l n g SKLM vdi hp s l Rf = 0,67 theo phuang phap cua Dupe d i l n Viet Nam IV vd chung minh d u true san pham bdng pho hong ngoai, pho k h l i , pho cpng hudng tir hat nhdn proton; san p h i m magnesi lactat tinh e h l dat lieu chuan k i l m nghiem theo Duac dien Anh - 2010. Oa tim dupe pH = 6,0 la thich hpp d l nudi d y vi khuan vd bo sung dan magnesi carbonat vao cde thai diem sau m i l 24 gid Idn men; hieu s u i t lao magnesi lactat vdi mdi Irudng MRS bo sung 2,6 % sua la 67,9 %.
Tai lieu tham khao
1 p. J. M. Baets (2013), "Process for the fermentative production of tactic acid from a plant extract the presence of a caustic magnesium sair, WIPO Patent Application. EP2604696 A l .
2. Bg Y te (2012), Duoc dien Vidt Nam IV, tr. 375 3. British Pharmacopoeia Commission (BPC) (2010), Bnstish Pharmacopedia, pp. 1325.
4. A. M. Eyal, P. R. Gruber,P Mc, Williams and D. R. Witkze (1998), "A process for the recovery of lactic acid", PCT int., App/..(Cargill incorporated USA), pp. 26.
5. J. V. Krneken, R Otto., G. N. Nanninga, J. J.
De Vnes (2005), "Process for the preparation of lactic acid or lactate from a magnesium lactate compnsing medium". United States Patent Office, US 7705m B2.
6. DSm Thanh Xuan, Le Ngpc Khanh, Le Thj Ttiu Hien (2012), "Nghien cuu su dyng te bao vi khuSn L acidophilus c l dinh tren chat mang alginat trong len men san xult caici lactat". Tap chi Hoa hgc, 50 (50A), pp.117-121.
7 F. Widdel (2010), "Theory and measurement of bacterial growth", Grundpraktikum Mikrobiologie, 4 Sem. (BSc). Universitat Bremen, pp. 4-11 {Ngay nhan bdi: 27/04/2015 - Ngay duyet ddng: 02/07/2015)
^ Danh gia tac dung chong viem thirc nghiem cua cao dae bao che tir dich chiet ethanol
bai thuoc EZ
Nguyen Manh Tuyin'*, Phung Hoa Binh', Le Thi Minh^
Nguyen Duy ThuSn^ Pham Thi Van Anh^ Vu Duy Hong*
' Truang Dgi hoc Dugc Hd Noi -Hoc vien Y Dugrc co truyen J^etNam
^Truang Dgi hpc YHd Ngi
^Vien Huyet hgc vd truyin mdu Trung uang 'E- mail: [email protected]
Summary
EZ-ethanol extracts were evaluated the local anti-inflammatory effects by the croton oil ear fest, acute anti-inflammatory effects by canageenin-induced edema in hind paw of mice and chronic antl-inflammatory effects by granuloma test of mice, at doses of 1.92 g and 5.76 g/kg bw. of rrtice (correspond to 1 time and 3 times of the clinical dose). The EZ-ethanol extracts did not provoked anti- inflammatory effects in the croton oil ear tests, but exhibited acute and chronic anti-inflammatory effects
T.APCHI DL P C H p C - 7/2015 (SO 471 N.4M 55)
• Nghien cu'u - Ky thuat
through inhibiting carrageenin-induced edema in hind paw after 4 and 6 hours, decreasing significantly granuloma weight, histological indices as compared with the control group (p<0.05).
Keywords: EZ-ethanol extraction, anti-inflammatory, experimental.
D a t v a n d e
Phuang thuoc EZ d u p e xay d y n g d y a tren ea s d ly ludn y hpc c6 t r u y i n k i t hpp vdi cac k l l qud nghien c u u t h y c nghipm. Phuang thuoc EZ da d u p e dung d l d i l u tn trdn ngudi b^nh (Phdng c h i n tn PhOng Gia Dudng) the hipn tdc dyng va hipu lye trong dieu trj viem da ca dja (eczema), o l tham djnh gia trj dieh thye cua bdi t h u l c , chOng tdi da t i l n hdnh bao e h l cao d | e ICr djch c h i l t ethanol ciia bdi t h u l c va l i l n hanh nghien c d u nhdm myc lidu:
- Ddnh gid tdc di^ng chdng vidm cap tai chd trdn chudt nhdt trdng.
- Danh gia tdc dijng chdng vidm cap vd m§n tlnh todn thdn trdn chugt nhdt tring.
N g u y e n l i e u , phu'O'ng t i e n v a phu'O'ng p h a p n g h i e n CLPU
Nguyen lieu nghien cu'u
Bai thu6c EZ gdm: kim ngdn d i n g (Caulis cum folium Lonicerae), nuc nac {Cortex Oroxylf), hod hoa (Flos Styphnolobii japonici imaturf), t h u a n g nhT ttr {Fructus Xanthii strumarii), hodng bd {Cortex Phellodendri) dpt lieu ehuan DOVN IV.
Cao d$c duw: bdo che tie djch chiet ethanol bai thuoc EZ (ggi tit id cao EZ-EtOH):
edn rieng tCrng d u p e lieu da d u p c xir I;? dat tieu c h u i n theo k h l i lupng tuang ung vdi 5 thang (300 g d u p c lidu). C h i l t hdl luu rieng lirng vj bdng ethanol 70% 3 l l n , m l i lan 2 gid vdi lupng ethanol g i p 5 lan d u p c lieu. P h l i hpp dieh chiet, d t thu hoi dung mdi, ly tdm lay djch. Cd dae tren cdch thuy thu d u p e 40 g cao dpc, hay 1 g cao d | c l u a n g duang 7,5 g d u p c lieu.
Wda chit, thuoc thir: Aspirin - Aspegic (DL- lysine aeelylsalicylat) gdi bpl 100 mg (Sanofi Aventis). Methyl prednisolon (Medrol) vien ndn 4 mg (Pfizer). Dung djch carragenin 1 % . Dau croton (Sigma - Ode). Cac hda ehat khdc dat lidu chuan phdn tich.
Phirong ti$n: Mdy do viem Plethysmometer No7250 cua Hang Ugo-Basile (Italy). Thudc do
dd day (dp ehinh xde 0,02 mm). T h i l l bj sinh t h i l t (biopsy punch) dudng kinh 7 mm. Can dipn l u (dd chinh xac 0,001 g).
Opng v|it nghien c i r u
Chupt nhdt trang chung Swiss, ca 2 giong, khoe mgnh, trpng lupng 18- 22 g do Vien Vp smh djch t l Trung u a n g eung e l p .
P h u a n g phap nghien ciru
Danh gia tac dung chong vidm dp tai cho Chudt nhdt trdng d u a c ehia ngau nhien thanh 4 Id, moi Id 10 eon nhu sau:
Ld 1 (md hinh): Gdy md hinh + eho u l n g nude d t 0,2 m l / I O g chupt.
Ld 2 (ehirng duang): Gdy md hinh + cho u l n g melhylprednisolon 20 mg/kg chudt.
Ld 3 (cao EZ-EtOH lieu thip): Gay md hinh + cho u l n g cao EZ-EtOH lieu 1,92 g/kg ehudt (tuang ung 14,4 g d u p c lipu (DL)/kg).
Ld 4 (cao EZ-EIOH l i l u cao): Gdy md hinh + eho uong eao EZ-EtOH l i l u 5,76 g/kg ehudt (tuang irng 43,2 g DL/kg).
Chupt dupc cho uong thuoc thu 2 ngay trude khi t h u nghiem. Ngay t h u 3, trudc khi gay md hinh 1 gid vdi l i l u luang nhu trdn, chudt d u a c gdy md hinh bang cdch bdi 20 pi dung djch dau croton (40 mg ddu pha trong 2 ml aceton) len mpt ngodi tai phai. Tai trdi khdng bdi de doi ehilu. Trudc khi gay md hinh, chudt d u p c do chieu day tai tai vi tri sat dinh cua lai each xa chdp syn vanh tai.
Sau 6 gid, chupt d u p c gay me bang thiopental, do lai ehilu day lai chudt. Cat d phan trung tdm vdi dudng kinh 7 mm bang dung cy sinh thilt, edn xac dinh khoi lupng '^'^i.
Ddnh gia tdc dung chong viem cip toan than
Chudt nhal trang d u p c ehia ngdu nhien thdnh 4 Id, mdi Id 10 con:
Ld 1 ( d l i chirng): cho u l n g nude d l 0,2 m l / I O g chudt.
Ld 2 (ehirng duang): cho u l n g aspirin l i l u 200 mg/kg ehudt.
TAP CHI DUpc H p c - 7/2015 (SO 471 NAM 55)
Nghien CLPU - Ky thuat
Ld 3 (eao EZ-EIOH l i l u thdp): cho uong eao EZ-EtOH l i l u 1,92 g/kg ehudt
Ld 4 (eao EZ-EtOH lieu cao): cho udng cao EZ-EIOH lieu 5,76 g/kg chudt.
Chupt dupe udng t h u l c 3 ngdy lien tyc tarde khi gay viem. Ngay thu 3, sau khi u l n g thude thir 1 gid, liem earrageenin 1 % (tnang nude mudi sinh ly) 0,05 ml/ehudl vdo gan ban ehdn sau, ben phdi. Do t h l tich ehdn chudt ( d i n Ididp d ehan) bdng may Plethysmometer d cac thdi d i l m : trudc khi gay viem (VO); sau khi gay viem 2 gid (V2), 4 gid (V4), 6 gid (V6) ^-^i.
Danh gia tac dung chong viem man tinh toan than
Chupt n h i t trdng dupe ehia ngau nhien thanh 4 Id, moi Id 10 con:
Ld 1 (doi chung): eho udng nudc d t 0,2 ml/10 gchuOl.
Ld 2 (chirng duang): eho uong methyl prednisolon lieu 10 mg/kg chudL
Ld 3 (cao EZ-EtOH lieu thap): cho ulng cao EZ-EtOH l i l u 1,92 g/kg chupt.
Ld 4 (eao EZ-EtOH lieu cao): cho uong cao EZ-EtOH l i l u 5,76 g/kg chuot.
Cdy amianl trpng lupng 6 mg (spi amiant c6 dudng kinh khoang 2 mm d u a c ve trdn vd say tiet khuan trong 1 gid d 120 °C, t i m earrageenin 1 % vao da gay chudt. Sau khi d y amiant, chu^t d u p c cho uong n u d e d t hope thuoc thu lidn ^ c trong 10 ngdy. Ngdy t h u 11, g i l l chupt, bdc tdch k h l i u hat, dem say khd d nhidt dp 56 "C trong 18 gid. Can trpng lupng u hat sau khi dd dupc sdy khd '''•^.
XiP iy so iieu
So lieu d u p c b i l u d i l n d u d i dgng x ± SD, k i l m djnh bdng test t - Student S y khdc bi$t c6 y nghla thong ke khi p < 0,05.
K§t qua nghien cu'u
Tdc d u n g c h o n g v i d m cap tai cho tren tai c h u p t nhdt bdng dau c r o t o n
Bang 1: Chieu ddy tai chudt truoc vd sau khi gay viem 6 gid' (n =10}
1 Lo f (mo hinh) 2 Lo 2 (melhylprednisolon) 3 L6 3 (cao EZ-EIOH lieu (hap) 4 L6 < (cao EZ-EtOH 1 ^ cao)
0,02 1,92 5,76
Chieu day tai Truvc 21,20 ±3,29 21,00 ±4,32 21,80 ±3,71 19,8012,39
ichugt, (x± SD, 10,im) Sau 6 giu 27,00 + 3,68*
23,40 ±3,12' 25,40 ±3,86- 26,00 ±4,7r (Ghi chu: so sdnh truoc sau: +, p<0,05; so vol nhdm chtmg: *, p<0,05)
Khdng ed s y khdc biel ve chieu day tai ben phai d thdi ^ I m t n j d c nghien cuu giua cac Id.
Cy Id md hinh, chieu day eua tai phai lang len ro rpt so vdi t n j d c khi gdy viem (p < 0,05). Ld uong melhylprednisolon, c h i l u day lai ed tang nhung khdng ed s y khac biet so vdi trudc khi gay viem (p > 0,05). Nhung s y khdc biet cd y nghTa thong
ke so vdi Id md hinh (p < 0,05), t h l hipn tdc dyng chdng viem rd ret cua thuoc doi chirng duang,
Cao EZ-EtOH lieu 1,92 g/kg vd lieu 5,76 g/kg mdi ehl t h l hidn ed xu hudng giam dp ddy tai chupt so vdi Id md hinh, s y khac bipt khdng cd y nghTa thdng ke (p > 0,05).
Bang 2: HJidi luong tai chuot vd m u c dp uc che viem (n =10) Lothu
Lo 1 (mo hinh) Lo 2 (melhylpredneolon) Lo 3 (cao EZ-EtOH lieu thap]
Lo 4 (cao EZ-EIOH leu cao) Ueu(g/ffg}
0,02 1.92 5,76
Khoi tuang tai citugt, (X± SD, mg) Taitiai Tai ptiai 14,20 + 2,66
12,90±1,45 13,90 ±1,45 14,70 ±3,20
33,80 ±6,37 17,60±3,31' 30,10±4,36*
30,01 ±2,96-
Mi>c dg IK clK vi6m
(%)
44,08 10.06 12,43 {Ghi chu: so sdnh truoc sau:+, p<0,05; so vol nhom chung: *, p<0,05)
TAP CHi Dirpc H p c - 7/2015 (SO 471 NAM 55)
• Nghien ciPu - Ky thuat
Cf 16 md hinh, khdi luang tai phdi tang rd ret so vdi tai ben trai. Ld uong melhylprednisolon, khoi lupng tai gidm ro ret so vdi Id md hinh (p < 0,05), mirc dp ii-c c h l viem cua mettiylprednisolon la 44,08%.
Cao EZ-EIOH lieu 1,92 g/kg va 5,76 g/kg deu
Bang 3: Anh huong cua cao EZ-EtOH den do phO chdn chuot (n =10)
cd xu h u d n g giam khoi lupng tai chupt so vdi Id md hinh vdi muc dp uc c h l vidm lan lupt Id 10,06 % va 12,43 %. S y khac bidt khdng cd y nghTa t h i n g ke (p > 0,05).
Tdc d u n g c h l n g v i e m cap t o d n thdn
Uiu - Saulgi&C/^ $au4giiy(V) Sau6gia(V^
% giim phu Sl % giam phu so vdi diung
% giam phu so Lo 1 (doi diLPng)
L£ 2 (aspinn)
p,.j<0,05
57,47 ±19,02 p,^<0,05 t a 3 (cao EZ-EtOH lieu thap) 1,92 84,55 ±22,93
p,j>0,05
60,06 + 14,56 p,j<0,05 44,65114,30
Lo 4 (cao EZ-EIOH lilu cao) 5.76 p,^> 0,05 p„<0,05
53.76118,95 43.04±15,86 p,^<0,001 41,30 P,^<0,01 p„<0,01 p„<0,05 Aspirin 200 mg/kg cd tdc dung ehong viem
cap lai tat d cdc thdi d i l m nghien cuu (p < 0,05).
Cao EZ-EtOH l i l u 1,92 g/kg cd lac dung ldm gidm phii chdn ehudt d thdi diem 6 gid (p < 0,05), lai thdi diem 4 gid sau gay viem, mdi chi ed xu hudng lam giam phil ehan chudt (p > 0,05).
Cao EZ-EtOH lieu 5,76 g/kg cd tdc dung Idm gidm phii chdn chupt d cac thai d i l m sau gay viem 4 gid (p < 0,001) va 6 gid (p < 0,01). Sau
gay viem 2 gid, ed xu hudng lam giam phil ehan chudt (p > 0,05).
Cao EZ-EtOH l i l u 5,76 g/kg cd tdc dung ehong vidm tot han d lieu 1,92 g/kg. S y khac biet ed y nghTa thong ke d cdc thai diem danh gid sau gay vidm 2 gid (p < 0,05), 4 gid (p < 0,01) va 6 gid (p < 0,05)
Tac d u n g c h o n g viem man todn than
Bang 4: Anh huung cua cao El-EtOH ten Idto Ld
Lfi 1 (doi cht>ng) Lfi 2 (methylprednisofon) Lfi 3 (cao EZ-EtOH lieu thap) Lfi 4 (cao EZ-EtOH Ii4u cao)
Liiu (g/ligj
0,01 1,92 5,76
KImi iugng u (mgi 66,23*18,67
43,36 + 7,58 38,77±11,73 63,90 ±18,11
lu^ng u hat p so voi to 1
<o,os
<0,01
>0,05 (n =10)
p so voi 16 2
>0,05
>0,05 Melhylprednisolon lieu 10 mg/kg/ngdy ed tde
dyng lam giam trpng lupng khoi u hat so vdi Id ehirng (p < 0,05), the hipn tac dung ehong viem rd rpt.
Cao EZ-EtOH lieu 1,92 g/kg lam giam trpng lupng k h l i u hgt so vdi Id chung (p < 0,01), tde dyng nay t u a n g duang vdi methylprednisolon lieu 10 mg/kg (p > 0,05). Lieu 5,76 g/kg chupt.
cao EZ-EIOH giam trpng lupng khoi u hat khdng dang k l so vdi id chirng (p > 0,05).
Ban luan
Ve tac d u n g c h o n g viem cap tai c h o K i t qua nghidn cuu eho thay, ed s y khdc bidt rd rpt v l chieu day va k h l i lupng tai chudt d id md hinh trudc va sau gay viem 6 gid, chung to TAP CHI D U p c H p c - 7/2015 (SO 471 NAM 55)
Nghien CLFU - Ky thuat
md hinh dupe thye hipn thanh edng. Thuoc ddi ehCrng duang methylprednisolon t h l hien hieu lyc chong viem ro rpt tren md hinh nay, phu hop vdi ede nghien euu da cdng bo ^.
S y khae bipt v l dp day vd khoi lupng lai ehudt d eae Id t h u eao EZ-EIOH, lieu 1,92 g/kg va 5,76 g/kg trudc va sau gdy viem d p b l n g tac nhan gay viem la dau croton khdng cd y nghta t h i n g ke (p > 0,05), ehirng td d l i l u thir vd trdn mo hinh t h u nghidm nay, eao EZ-EtOH khdng ed tdc dung ehong viem d p tpi cho.
Ve tac d u n g chong viem cap toan than Mdt ^ong nhung b i l u hien cua qua trinh viem Id gian mach, bach d u xuyen mach, lang t i l t eac chat trung gian hod hpc nhu prostaglandin, histamin... gdy phii '^i. Do vdy, danh gia lae dung ue che phii chdn chudt id mdt Irong nhung thu nghipm thudng quy de thi> tac dung c h l n g viem cua cac t h u l c t u thao dupe.
K i t qua nghien euu eho thay, cao EZ-EtOH d cac lieu thu deu cd lac dyng giam dd phu ban ehdn ehupt. C/ lieu 1,92 g/kg ed tde dyng giam phO sau gdy viem 6 gid (p < 0,05); d l i l u thir 5,76 g/kg, cd tdc dyng giam phii sau gay viem 4 gid vd 6 gid (p < 0,01). Lieu t h u cao han, lac dyng gidm phii d i n sdm han va manh han, ehung td tde dyng ehong viem d p toan than cua eao EZ-EtOH cd xu hudng phy thudc vao lieu.
Ve tdc dMng chong viem man tinh toan thdn
Cao EZ-EtOH, lieu 1,92 g/kg chupt Idm giam trpng lupng u hpt so vdi Id chung (p < 0,05).
M i u nghien euu cd tac dung chdng viem man tfnh tuang duang thuoc doi ehirng duang methyl prednisolon l i l u 10 mg/kg chudt. Tdc dung ndy rat quan trpng trong hd trp dieu tn benh eczema, mpt benh viem da man linh ea dia rdt khd d i l u tn tren lam sdng va phu hpp vdi tac dung chong viem cua nhieu vj thuoc trong bai t h u l c da dupc cdng bo nhu: dan la do, kim ngdn dang, nuc nac... "'•I
Cao EZ-EtOH l i l u 5.76 g/kg ehudt lai khong the hien ed lac dyng u c c h l u hgt thyc nghiem (p > 0,05). O i l u nay ed t h l la do d l i l u cao, hoat chat nhieu han nhung tap ehat eung nhieu hon nen anh hudng d i n kha nang hap thu hopt chat, gay gidm hieu lye eua bai thude? Tuy nhien gia thuyit ndy d n ed cdc nghien ciru t i l p theo de ehirng minh.
Ket luan
- Cao EZ- EtOH khdng cd lae dung chong viem tai ehd d l i l u 1,92 g/kg va 5,76 g/kg.
- Cao EZ- EtOH cd lac dung ehong viem cap toan than d lieu 1,92 g/kg va 5,76 g/kg, tdc dyng ed xu hudng phy thupc vao lieu.
- Cao EZ-EtOH ed tde dyng c h l n g viem man tinh loan than d l i l u 1,92 g/kg.
Tai lieu tham khao
1. A. Tubaro, P. Dri, G. Delbello, et aL(1985). The croton oil ear test revisited. Agents and Actions. 17:
(3-4) 347 - 349
2. McCoil S. R., Hurst N P, Cleland L G.
(1986), "Modulation by phorbol myristate acetate of araehidonie acid release and leukotrlene syn-thesis by human polymorphonuclear leukocytes stimulated with A23187'', Biochemical and Biophysical Research Communications.14^. 399-404.
3. Orley Vanesa Celeste, lldenize Barbosa da Silva, et al. (2004),''lnve5tigation of the anti-inflammatory and analgesic activities of a sample of Brazilian propolis'.
Acta Farm. Bonaerense. 23 (3), 285-91.
4 A. C. B. Paula, L. S, S Hayash, J. C. Freitas (2003), "Anti-inflammatory and antispasmodic acfivity of Ipomoea imperati (Vahi) Griseb (Convolvulaceae), phannacological effects of Ipomoea imperati', Brazilian Joumal of Medical and Biological Research, 36, 105- 112
5 Ezequiel Paulo Virlato, Erica Silva Bianchetti, Kelem Costa dos Santos et al. (2009), "Study of high dilutions of Copaiba oil on inflammatory process", Int J. High Dilution Res., 8(26), 9-14.
6 Vien Dupc lipu (2004), Cay thuoc va ddng vat lam thuoc & Vidt Nam tap 1, Khoa hoc va Ky thugt. Hi Noi, tr 814-819, 930-934, 971-976, 1044-1047.
7. Vien Duoc lieu (2004), Cay thuoc vd dgng v$t lam thuoc a Viet Nam t§p 2, Khoa hpc va Ky thuSt, Ha Noi, tr. 106-110,480-484
(Ngay nhan bdi: 03/05/2015 - Ngay duyet dang: 02/07/2015)
10 T.\P C H i D t r p c H p c - 7/2015 (S6 471 NAM 55)