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These two parts were subsequently supplemented by further guidelines which are integral parts of these GMP for pharmaceutical products. All these texts are available on the Medicines web page (http.www.who.int/medicines/organization/qsm/activities/qualityassurance/gmp/gmpc over.html). 这两部分随后通过药品GMP指南不可缺少的一部分内容进一步补充，这些内容可在药品网站上查到： (http.www.who.int/medicines/organization/qsm/activities/qualityassurance/gmp/gmpc
9. Personnel 人员 General 概述 Key personnel 关键人员 10. Training 培训 11. Personal hygiene 人员卫生 12. Premises 厂房 General 概述 Ancillary areas 辅助区域 Storage areas 仓储区域 Weighing areas 称量区域 Production areas 生产区域 Quality control areas QC区域 13. Equipment 设备 14. Materials 物料 General 概述 Starting materials 起始物料 Packaging materials 包装材料 Intermediate and bulk products 中间体和成品 Finished products 制剂 Rejected, recovered, reprocessed and reworked materials 不合格、回收、返工和重加工物料 Recalled products 召回产品 Returned goods 退货 Reagents and culture media 试剂和培养基 Reference standards 标准品 Waste materials 废弃物料 Miscellaneous 其他 15. Documentation 文件 General 概述 Documents required 需要的文件 16. Good practices in production 生产良好规范 General 概述 Prevention of cross-contamination and bacterial contamination during production生产过程中交叉污染和额细菌污染的预防

Antioxidative and ACE inhibitory activities of protein hydrolysates from the muscle of brownstripe r

Process Biochemistry 46(2011)318–327Contents lists available at ScienceDirectProcessBiochemistryj o u r n a l h o m e p a g e :w w w.e l s e v i e r.c o m /l o c a t e /p r o c b ioAntioxidative and ACE inhibitory activities of protein hydrolysates from themuscle of brownstripe red snapper prepared using pyloric caeca and commercial proteasesSutheera Khantaphant a ,Soottawat Benjakul a ,∗,Hideki Kishimura ba Department of Food Technology,Faculty of Agro-Industry,Prince of Songkla University,Hat Yai,Songkhla,90112,ThailandbLaboratory of Marine Products and Food Science,Research Faculty of Fisheries Science,Hokkaido University,Hakodate 041-8611,Japana r t i c l e i n f o Article history:Received 4June 2010Received in revised form 10September 2010Accepted 10September 2010Keywords:Protein hydrolysate Antioxidative activityGastrointestinal model system Oxidation model system Two-step hydrolysisa b s t r a c tProtein hydrolysates from the muscle of brownstripe red snapper (Lutjanus vitta )prepared using Alcalase or Flavourzyme as the ﬁrst step with 40%degree of hydrolysis (DH),followed by hydrolysis with pyloric caeca protease (PCP)as the second step for 2(HAP)and 1h (HFP),respectively,were prepared and determined for their antioxidative and angiotensin I-converting enzyme (ACE)inhibitory activities.HAP had the higher DPPH and ABTS radical scavenging activity and ferric reducing antioxidant power (FRAP),while HFP showed the higher ferrous chelating activity and ACE inhibitory activity (p <0.05).Both HAP and HFP were able to retard lipid oxidations in lecithin–liposome and ␤-carotene–linoleic acid oxidation model systems in dose-dependent manner.HAP and HFP contained 87.36and 86.55%protein (wet basis),respectively with glutamic acid/glutamine as the major amino acids,followed by aspartic acid/asparagine,lysine,alanine and leucine,respectively.HFP showed a slightly greater efﬁciency in prevention of lipid oxidation in all systems tested.Antioxidative activities,except DPPH radical scavenging activity,of both HAP and HFP after being subjected to gastrointestinal tract model system (GIMs)increased,suggesting the enhancement of antioxidative activities of both hydrolysates after ingestion.©2010Elsevier Ltd.All rights reserved.1.IntroductionLipid oxidation is one of the major deteriorative processes in many types of foods,leading to the changes in food quality and nutritional value.Additionally,potentially toxic reaction products can be produced.Lipid oxidations have been known to be the major causes of many serious human diseases,such as cardiovascular dis-ease,cancer,and neurological disorders as well as the aging process [1,2].To prevent oxidative deterioration of foods and to provide protection against serious diseases,such as cancer and atheroscle-rosis [3,4],it is important to inhibit the oxidation of lipids and the formation of free radicals occurring in the foodstuff and living body.To tackle the problem,antioxidants,both natural and synthetic ones,have been used widely.Nevertheless,synthetic antioxidants have been suspected of being responsible for toxicity in the long term and their use in foodstuffs is restricted or prohibited in some countries [5–7].Therefore,there is a growing interest on natu-ral antioxidants,especially peptides derived from hydrolyzed food proteins.Protein hydrolysates from several ﬁsh species includ-ing round scad (Decapterus maruadsi )[8],yellow stripe trevally∗Corresponding author.Tel.:+6674286334;fax:+6674558866.E-mail address:soottawat.b@psu.ac.th (S.Benjakul).(Selaroides leptolepis )[9],Paciﬁc hake (Merluccius productus )[10],tilapia (Oreochromis niloticus )[11],silver carp (Hypophthalmichthys molitrix )[12]and smooth hound (Mustelus mustelus )[13]have been reported to possess antioxidative activities.Hypertension has been considered as the most common seri-ous chronic health problem [14].Since angiotensin I-converting enzyme (ACE)(EC 3.4.15.1)is physiologically important in raising blood pressure,the inhibition of ACE activity can lead to an over-all antihypertensive effect.The synthetic ACE inhibitors are now widely used as pro-drugs but these synthetic ACE inhibitors can cause many signiﬁcant undesirable side effects [14,15].Therefore,the natural safe compounds are desirable for prevention of hyper-tension instead of the synthetic counterpart.Among those,food protein derived peptides are promising natural products exhibiting ACE inhibitory activities.Protein hydrolysates with antihyperten-sive activity have also produced from sardinelle (Sardina pilchardus )by-products [16],tuna cooking juice [17],salmon protein [18]and tilapia [19].In Thailand,brownstripe red snapper (Lutjanus vitta )is one of the raw materials for surimi production [20].Besides being pro-duced into surimi,this species and its viscera,especially pyloric caeca,can be used as raw material for production of protein hydrolysate and as the source of proteases,respectively.Produc-tion of ﬁsh protein hydrolysates with bioactivity can pave the way1359-5113/$–see front matter ©2010Elsevier Ltd.All rights reserved.doi:10.1016/j.procbio.2010.09.005S.Khantaphant et al./Process Biochemistry46(2011)318–327319for full utilization of these species.Many factors affect the bioac-tivity of protein hydrolysates,e.g.type of proteases[9],steps of hydrolysis[21],etc.This work aims to study antioxidative and ACE inhibitory activ-ities of protein hydrolysate from brownstripe red snapper muscle prepared using its pyloric caeca protease in combination with commercial proteases via2-step hydrolysis and to investigate the bioactivities of selected hydrolysate after digestion in gastrointesti-nal tract model system.2.Materials and methods2.1.Enzymes and chemicalsAlcalase2.4L(E.C.3.4.21.62)(2.4AU/g)and Flavourzyme500L(E.C.3.4.21.77) (500LAPU/g)were provided by Novozyme(Bagsvaerd,Denmark).2,2 -Azinobis (3-ethylbenzothiazoline-6-sulfonic acid)(ABTS),1,1-diphenyl-2-picrylhydrazyl (DPPH),2,4,6-trinitrobenzenesulfonic acid(TNBS),1,1,3,3-tetramethoxypropane, 3-(2-pyridyl)-5,6-diphenyl-1,2,4-triazine-4 ,4 -disulfonic acid(ferrozine),2,4,6-tripyridyl-triazine(TPTZ),l-␣-phosphatidylcholine(lecithin)and linoleic acid were purchased from Sigma(St.Louis,MO,USA).Thiobarbituric acid(TBA),potassium persulfate,␤-carotene and Tween40were obtained from Fluka(Buchs,Switzer-land).Tris(hydroxymethyl)aminomethane(Tris–HCl)was procured from Merck (Darmstadt,Germany)and sodium sulﬁte was obtained from Riedel-deHaën(Seelze, Germany).All chemicals were of analytical grade.2.2.Preparation of pretreatedﬁsh minceBrownstripe red snapper,stored in ice and off-loaded approximately24–36h after capture,were purchased from a dock in Songkhla province,Thailand.Fish were transported in ice with theﬁsh/ice ratio of1:2(w/w)to the Department of Food Technology,Prince of Songkla University within1h.Upon arrival,wholeﬁsh were washed and onlyﬂesh was separated manually.Flesh was minced to uniformity using Moulinex AY46blender(Group SEB,Lyon,France)and phospholipid mem-brane was removed by further homogenizing with nine volumes of cold8mmol L−1 CaCl2solution containing5mmol L−1citric acid[22]using an IKA Labortechnik homogenizer(Selangor,Malaysia)at a speed of11000rpm for2min.After contin-uous stirring for60min at4◦C,the sample was centrifuged at4000×g for15min at4◦C using a Beckman Coulter centrifuge Model Avant J-E(Beckman Coulter,Inc., Fullerton,CA,USA).Thereafter,the pellet was washed by homogenizing withﬁve volumes of cold distilled water using a homogenizer at a speed of11000rpm for 2min,followed by stirring at4◦C for15min prior to centrifuging at9600×g for 10min at4◦C.The washing process was repeated twice.Pretreated mince obtained was kept in polyethylene bags and placed in ice until use,but not longer than2h.2.3.Preparation of proteases from pyloric caecaPyloric caeca from brownstripe red snapper was collected and powdered in liq-uid nitrogen.Thereafter,the pyloric caeca extract was prepared according to the method of Khantaphant and Benjakul[20].Pyloric caeca powder was suspended in ten volumes of extraction buffer(50mmol L−1Tris–HCl buffer,pH8.0,contain-ing10mmol L−1CaCl2.The mixture was homogenized at11000rpm for2min.The homogenate was continuously stirred for30min at4◦C and centrifuged at8000×g for30min at4◦C.The supernatant wasﬁltered through a Whatmanﬁlter paper No.1(Schleicher&Schuell,Maidstone,England).Theﬁltrate obtained was further subjected to40–60%saturation ammonium sulfate precipitation.After stirring at 4◦C for30min,the mixture was centrifuged at8000×g for30min at4◦C and the pellet obtained was dissolved in50mmol L−1Tris–HCl buffer,pH8.0followed by dialysis against20volumes of the extraction buffer overnight at4◦C with three changes of dialysis buffer.The dialysate was kept in ice and referred to as‘pyloric caeca protease;PCP’.The proteolytic activity of PCP was determined using casein as a substrate under optimal condition(60◦C and pH8.5)2.4.Preparation of protein hydrolysate from brownstripe red snapper2.4.1.One-step hydrolysis using different single proteasesPretreated brownstripe red snapper mince with a protein content of95.42%pro-tein(dry basis)determined by Kjeldahl method[23]was mixed with50mmol L−1 Tris–HCl buffer with pH of7.0,8.0and8.5for hydrolysis by Flavourzyme,Alcalase and PCP,respectively,to obtain aﬁnal protein concentration of20g protein L−1.The mixtures were homogenized at a speed of11,000rpm for1min and the pH was rechecked and readjusted using1mol L−1NaOH or1mol L−1HCl.The homogenates were pre-incubated for10min at50◦C for Alcalase and Flavourzyme and at60◦C for PCP[20].To start the hydrolysis,the different levels of enzymes were added into the mixture to obtain the desirable degree of hydrolysis(DH)of20,30and 40%following the method of Benjakul and Morrissey[24].After2h of hydrolysis, the reactions were inactivated by placing the mixture in boiling water for10min. Thereafter,the mixture was centrifuged at2000×g at4◦C for10min.The super-natant was collected and referred to as protein hydrolysate prepared using Alcalase (HA),Flavourzyme(HF)or PCP(HP).All protein hydrolysates were determined for antioxidative activities.2.4.2.Two-step hydrolysis using different proteasesAfter2h of theﬁrst hydrolysis,the mixtures with DH of40%,which had the high-est antioxidative activities,were heated for10min in boiling water and adjusted to the desirable pH using1mol L−1NaOH or1mol L−1HCl for proteases used for the second step.Those proteases included Alcalase,Flavourzyme and PCP.The same amount of proteases used in theﬁrst step was added into the pre-incubated mix-ture with optimal temperature of each protease.Reaction was conducted for1,2, 3and5h.At the time designated,the reaction was terminated by submerging the mixture in boiling water for10min.The mixture was then centrifuged at2000×g at4◦C for10min and the supernatant was collected and adjusted to neutral pH. The neutral solution was referred to as‘protein hydrolysates’and lyophilized to obtain hydrolysate powder.The hydrolysates obtained from HP with further hydrol-ysis using Alcalase(HPA)or Flavourzyme(HPF),hydrolysate from HA using PCP or Flavourzyme for the second step(HAP and HAF)and hydrolysate from HF using PCP or Alcalase for the second step(HFP and HFA)were determined for their antioxida-tive and ACE inhibitory activities.2.5.Determination of antioxidative activities2.5.1.DPPH radical scavenging activityDPPH radical scavenging activity was determined according to the method of Blois[25]with a slight modiﬁcation.Sample solution(1.5mL)was added with1.5mL of0.1mmol L−1DPPH in950mL L−1ethanol.The mixture was allowed to stand for 30min in dark at room temperature.The resulting solution was measured at517nm. The control was prepared in the same manner except that distilled water was used instead of the sample.The DPPH radical scavenging activity was calculated from Trolox standard curve(0–60␮mol L−1)and expressed as␮mol Trolox equivalents (TE)g−1protein.2.5.2.ABTS radical scavenging activityABTS radical scavenging activity was determined as described by Re et al.[26]. ABTS radical(ABTS•+)was produced by reacting ABTS stock solution(7.4mmol L−1 ABTS)with2.6mmol L−1potassium persulfate at the ratio of1:1(v/v).The mixture was allowed to react in the dark for12h at room temperature.Prior to assay,the ABTS•+solution was diluted with methanol to obtain an absorbance of1.1(±0.02) at734nm.To initiate the reaction,150␮L of sample was mixed with2.85mL of ABTS•+solution.The mixture was incubated at room temperature for2h in dark. The absorbance was then read at734nm.A Trolox standard curve(0–200␮mol L−1) was prepared.Distilled water was used instead of the sample and prepared in the same manner to obtain the control.ABTS radical scavenging activity was expressed as␮mol TE g−1protein.2.5.3.Ferric reducing antioxidant power(FRAP)The ability of samples to reduce ferric ion(Fe3+)was evaluated by the method of Benzie and Strain[27].FRAP reagent(a freshly prepared mixture of 10mmol L−1TPTZ solution in40mmol L−1HCl,20mmol L−1FeCl3.6H2O solution and300mmol L−1acetate buffer,pH3.6(1:1:10,v/v/v))(2.85mL)was incubated at 37◦C for30min prior to mixing with150␮L of sample.The reaction mixture was allowed to stand in dark for30min at room temperature.Absorbance at593nm was read and FRAP was calculated from the Trolox standard curve(0–60␮mol L−1)and expressed as␮mol TE g−1protein.The control was prepared in the same manner except that distilled water was used instead of the sample.2.5.4.Ferrous chelating activityChelating activity of samples towards ferrous ion(Fe2+)was measured by the method of Benjakul et al.[28]with a slight modiﬁcation.Sample(200␮L)was mixed with800␮L of distilled water.Thereafter,0.1mL of2mmol L−1FeCl2and0.2mL of 5mmol L−1ferrozine were added.The mixture was allowed to react for20min at room temperature.The absorbance was then read at562nm.The standard curve of EDTA(0–1.0mmol L−1)was prepared.The control was prepared in the same manner except that distilled water was used instead of the sample.Ferrous chelating activity was expressed as␮mol EDTA equivalents g−1protein.2.6.Determination of ACE inhibitory activityThe angiotensin I-converting enzyme inhibitory activity was determined as described by Hayakari et al.[29]with slight modiﬁcations.Sample(0.3mL)was incubated with725unit ACE L−1(50␮L)at37◦C for5min.Thereafter,the incubated mixture was added into the assay mixture(0.2mL)which was100mmol L−1K2HPO4 buffer(pH8.3)containing600mmol L−1NaCl and3mmol L−1hippuryl-l-histidyl-l-leucine(HHL).The mixture was incubated at37◦C for15min.To terminate the reaction,1.5mL of30g L−12,4,6-trichloro-1,3,5-triazine(dissolved in dioxane)and 3mL of0.2mol L−1K2HPO4buffer(pH8.3)were added and mixed thoroughly.The mixture was left for10min until the solution was clear and the absorbance at382nm was determined.Sample blank was prepared in the same manner except that HHL was added after the reaction mixture was terminated.The control was prepared320S.Khantaphant et al./Process Biochemistry46(2011)318–327by using distilled water instead of the sample,whereas the control blank was donein the same manner with the control but HHL was added after the termination.%inhibition of ACE was determined using the following equation:%Inhibition=A C−A SA C×100where A C=A control−A control blank and A S=A sample−A sample blank.2.7.Determination of antioxidative activity in different model systems2.7.1.ˇ-Carotene linoleic acid emulsion model systemThe antioxidative activity of the sample in the␤-carotene linoleic acid emulsion model system was determined as described by Binsan et al.[30].␤-Carotene(1mg) was dissolved in10mL of chloroform.Thereafter,the solution(3mL)was added to 20mg linoleic acid and200mg Tween40.Chloroform was then removed by purg-ing with nitrogen.Fifty milliliters of oxygenated distilled water were added to the ␤-carotene emulsion and mixed well.Hydrolysate(200␮L)was then mixed with 3mL of oxygenated␤-carotene emulsion to obtain theﬁnal concentrations of500 and1000ppm.The oxidation of␤-carotene emulsion was monitored spectrophoto-metrically at470nm after0,10,20,3040,60,90and120min of incubation at50◦C in dark.BHT at levels of100ppm was also used.The control was prepared by using distilled water instead of hydrolysate in the assay system.2.7.2.Lecithin liposome model systemThe antioxidative activity of protein hydrolysates in a lecithin liposome system was determined according to the method of Frankel et al.[31]slightly modiﬁed by Thiansilakul et al.[8].Lecithin liposome system was prepared by suspending lecithin in deionized water at a concentration of8g L−1.The mixture was stirred with a glass rod followed by soniﬁcation for30min in a sonicating bath(Elma Model S30H, Singen,Germany).Protein hydrolysate(3mL)was added to the lecithin liposome system(15mL)to obtain aﬁnal concentration of1000ppm.The mixture was son-icated for2min.To initiate the reaction,20mL of0.15mol L−1cupric acetate were added.The mixture was shaken in the dark at120rpm using a shaker(Heidolph Model Unimax1010,Schwabach,Germany)at37◦C.The systems containing25or 100ppm BHT were also prepared.The control was prepared in the same manner, except that distilled water was used instead of sample.Oxidation in lecithin lipo-some systems was monitored at6h intervals by determining the formation of TBARS and conjugated dienes.2.8.Determination of thiobarbituric acid reactive substances(TBARS)Thiobarbituric acid reactive substances(TBARS)were determined as described by Buege and Aust[32]with a slight modiﬁcation.Sample was homogenized with TBARS solution(3.75g L−1TBA,150g L−1TCA and0.25mol L−1HCl)with a ratio of 1:4(w/v).The mixture was heated in boiling water for10min to develop the pink color.Then the mixture was cooled with running water and centrifuged at5000×g for10min at room temperature using Hettich centrifuge(Hettich Model MIKRO-20,Tuttlingen,Germany).The supernatant was collected and measured at532nm using a UV-1800Spectrophotometer(Shimadzu,Kyoto,Japan).TBARS was calcu-lated from a standard curve of malonaldehyde(MDA)(0–10mg L−1)and expressed as mg MDA kg−1sample.2.9.Determination of conjugated dieneConjugated diene formed in the sample was measured according to the method of Frankel et al.[31].Sample(0.1mL)was dissolved in methanol(5.0mL)and con-jugated dienes were measured as the increase in absorbance at234nm.2.10.Preparation of gastrointestinal tract model system(GIMs)Gastrointestinal tract model system was prepared according to the method of Lo et al.[33]with slight modiﬁcation.Hydrolysate powder was dissolved in distilled water to obtain a concentration of50g protein L−1.The solution was adjusted to pH 2.0with1mol L−1HCl and pepsin dissolved in0.1mol L−1HCL was added to obtain theﬁnal concentration of40g pepsin kg−1protein.The mixture was incubated at 37◦C for1h with continuous shaking(Memmert Model SV1422,Schwabach,Ger-many).Thereafter,the pH of reaction mixture was raised to5.3with1mol L−1NaOH before adding20g pancreatin kg−1protein and the pH of mixture was adjusted to 7.5with1mol L−1NaOH.The mixture was incubated at37◦C for3h with continuous shaking.The digestion was terminated by submerging the mixture in boiling water for10min.During digestion,the mixture was randomly taken and determined for antioxidative activities at0,20,40,60,80,100,120,150,180,210and240min.2.11.Proximate analysisHAP and HFP were determined for protein,fat,ash and moisture contents according to the methods of AOAC[23]with the analytical number of992.15,991.36, 942.05and950.46,respectively.2.12.Amino acid analysisHAP and HFP were hydrolyzed under reduced pressure in4.0M methanesul-fonic acid containing0.2%(v/v)3-2(2-aminoethyl)indole at115◦C for24h.The hydrolysates were neutralized with3.5M NaOH and diluted with0.2M citrate buffer (pH2.2).An aliquot of0.4mL was applied to an amino acid analyzer(MLC-703;Atto Co.,Tokyo,Japan).2.13.Determination of protein concentrationProtein concentration was measured by the method of Lowry et al.[34]using bovine serum albumin as a standard.2.14.Statistical analysisExperiments were run in triplicate.All data were subjected to analysis of vari-ance(ANOVA)and differences between means were evaluated by Duncan’s Multiple Range Test.For pair comparison,T-test was used[35].SPSS Statistic Program(Ver-sion10.0)(SPSS Inc,Chicago,IL,USA)was used for data analysis.3.Results and discussion3.1.Antioxidative activities of brownstripe red snapper protein hydrolysates prepared with single step of hydrolysis usingdifferent proteasesProtein hydrolysates from brownstripe red snapper muscle prepared using protease from pyloric caeca of brownstripe red snapper,Alcalase and Flavourzyme referred to as HP,HA and HF, respectively,with different DH(20,30and40%DH)showed varying antioxidative activities(Fig.1).3.1.1.DPPH radical scavenging activityThe DPPH radical scavenging activity of HA increased with increasing DH(p<0.05)(Fig.1(a)).HF showed the similar activity at all DH used(p>0.05),whereas HP showed no further increase in activity when DH was higher than30%(p>0.05).At all designated DH,HA and HF showed the higher activity than did HP(p<0.05). The result suggested that the peptides in different hydrolysates might be different in term of chain length and amino acid sequence, which contributed to varying capabilities of scavenging DPPH rad-icals.The increase in DPPH radical scavenging activity of HA was in agreement with Thiansilakul et al.[8]who reported the increases in DPPH radical scavenging activity as the DH of the hydrolysate from round scad muscle protein prepared using Flavourzyme and Alcalase increased.On the other hand,Klompong et al.[9]found that DPPH radical scavenging activity of protein hydrolysate pre-pared from the muscle of yellow stripe trevally using Flavourzyme and Alcalase decreased when DH increased.Invert correlation between DH and DPPH radical scavenging activity was obtained for protein hydrolysates prepared from alkaline-aided channel catﬁsh protein isolates using Protamex[36].You et al.[37]reported that loach protein hydrolysate showed the greater DPPH radical scav-enging activity when DH increased.DPPH is a stable free radical and can be scavenged with a proton-donating substance,such as an antioxidant[25].Therefore,protein hydrolysates from brown-stripe red snapper muscle more likely contained peptides acting as hydrogen donors,thereby scavenging free radicals by converting them into more stable products.3.1.2.ABTS radical scavenging activityIn general,ABTS radical scavenging activities of protein hydrolysates increased as DH increased(p<0.05)(Fig.1(b)).The highest activity was observed in HP and HA with40%DH(p<0.05). However,no difference in activity was found in HF with all DH used(p>0.05).Among all hydrolysates,HA had the lowest activity for all DH tested(p<0.05).At DH of20%,HF showed higher ABTS radical scavenging activity than did HP(p<0.05).Conversely,HPS.Khantaphant et al./Process Biochemistry46(2011)318–327321Fig.1.Antioxidative activities of protein hydrolysate from brownstripe red snapper muscle prepared using pyloric caeca protease(PCP)from brownstripe red snapper (HP),Alcalase(HA)and Flavourzyme(HF)with different DHs.Bars represent the standard deviation(n=3).Different capital letters within the same enzyme used indicate signiﬁcant differences(p<0.05).Different letters within the same DH indicate signiﬁcant differences(p<0.05).had the highest activity when DH of40%was used(p<0.05).Pro-tein hydrolysate from alkali-solubilized tilapia protein prepared using various proteases showed a sharp increase in ABTS radical scavenging activity when DH increased from18to23%[11].Loach protein hydrolysate showed a similar result,in which ABTS radical scavenging activity increased with increasing DH[37].For both DPPH and ABTS assays,HF showed no differences in activities with all DH tested(p>0.05).ABTS radical assay is used for determining the antioxidative activity,in which the rad-ical is quenched to form ABTS-radical complex[26].Generally, all hydrolysates contained peptides,which were able to scav-enge ABTS radicals,leading to the termination of radical chain reaction.3.1.3.Ferric reducing antioxidant power(FRAP)Ferric reducing antioxidant power(FRAP)measures the reduc-ing ability against ferric ion(Fe3+),indicating the ability of hydrolysates to transfer an electron to the free radical[27].FRAP of different hydrolysates with varying DH is depicted in Fig.1(c). An increase in FRAP was observed in all hydrolysates when DH increased(p<0.05).FRAP of HA was generally higher than those of HP and HF at all DHs tested(p<0.05).The result suggested that HA had the greater reducing power than did others,leading to the greater efﬁcacy in prevention and retardation of propagation in lipid oxidation.However,protein hydrolysates prepared from alkaline-aided channel catﬁsh protein isolates showed the decrease in reducing power with increasing DH[36].Raghavan et al.[11] reported that alkaline-solubilized tilapia protein hydrolysate pre-pared using Flavourzyme showed the increase in reducing ferric ion when DH increased.The hydrolysis most likely increased the reducing power,especially when the cleavage of peptides increased as indicated by the increase in DH.The result indicated that pep-tides generated from the hydrolysis by different proteases had the different capacities of providing electron to the radicals.3.1.4.Ferrous chelating activityFerrous chelating activities of hydrolysates prepared using dif-ferent proteases with different DHs are shown in Fig.1(d).Chelating activity against Fe2+of HP slightly increased when DH increased up to30%(p<0.05).For HA and HF,the decreases in ferrous chelating activities were found as DH increased(p<0.05).Ferrous ion(Fe2+) is the most powerful pro-oxidant among metal ions[38],leading to the initiation and acceleration of lipid oxidation by interaction with hydrogen peroxide in a Fenton reaction to produce the reactive oxy-gen species,hydroxyl free radical(OH•)[39].Therefore chelation of metal ions by peptides in hydrolysates could retard the oxida-tive reaction.The result indicated that a higher DH rendered HA and HF with lower metal-chelating activities.The shorter chain of peptides might lose their ability to form the complex with Fe2+. At DH of20%,HF showed the highest chelating activity(p<0.05), followed by HA and HP,respectively.Peptides in HF could effec-tively chelate the Fe2+,leading to the retardation of initiation stage. The result indicated that the limited hydrolysis of muscle protein resulted in the enhanced ferrous chelating activity,compared with the excessive hydrolysis.The higher chelating activity of HP was coincidental with the higher DPPH and ABTS radical scavenging activity and FRAP,as the DH increased.Fe2+chelating activity of round scad protein hydrolysate prepared using Alcalase showed the increase in chelating activity with increasing DH,but those treated with Flavourzyme showed no difference in activity at all DH tested[8].With the same enzymes used,chelating activity of pro-tein hydrolysate prepared from the muscle of yellow stripe trevally322S.Khantaphant et al./Process Biochemistry46(2011)318–327using Flavourzyme and Alcalase increased with increasing DH[9]. Higher ferrous chelating activity was reported for hydrolysate of sil-ver carp using Alcalase and Flavourzyme when DH increased[12]. Apart from Fe,other transition metals,such as Cu and Co can affect the rate of lipid oxidation and decomposition of hydroperoxide. Theodore et al.[36]reported that Cu2+chelating activity of catﬁsh protein hydrolysate increased with increasing DH.Some proteins and peptides can chelate metal ions like Fe2+due to the presence of carboxyl and amino groups in the side chains of acidic and basic amino acids[10,40].Alcalase is endopeptidase capable of hydrolyzing proteins with broad speciﬁcity for peptide bonds and prefers for the uncharged residue,whereas Flavourzyme is a mixture of endo-and exopeptidase enzyme,which can produce both amino acids and peptides[41].Hydrolysates showing differ-ent antioxidative activities might be attributed to the differences in the exposed side chains of peptides as governed by the speciﬁcity of different proteases towards peptide bonds in the proteins[42]. DH also greatly inﬂuenced the peptide chain length.The higher DH,the more cleavage of peptide chains took place.Peptides with various sizes and compositions had different capacities of scaveng-ing or quenching free radicals[8,9].PCP,Alcalase and Flavourzyme more likely cleaved the peptide bonds in brownstripe red snap-per muscle at different positions,resulting in the different products with varying antioxidative activities.With40%DH,all hydrolysates functioned more effectively as primary antioxidant,whereas the secondary antioxidative activity,chelating ability,was lowered.To enhance the antioxidative activity of peptides,especially as the pri-mary antioxidant,the hydrolysate with40%DH was prepared for theﬁrst step of hydrolysis.3.2.Antioxidative and ACE inhibitory activities of brownstripered snapper protein hydrolysate prepared with two-stephydrolysis using different proteases3.2.1.DPPH radical scavenging activityDPPH radical scavenging activity of protein hydrolysates with various hydrolysis times in the second step of hydrolysis using another protease is shown in Fig.2(a).For HAF,HAP,HFA and HFP, the activities increased with increasing time up to2h(p<0.05). Thereafter,the gradual decrease was observed when hydrolysis times were3and5h(p<0.05).No changes in DPPH radical scav-enging activity were observed for HPF,while HPA showed the maximal activity at3h of hydrolysis(p<0.05).The results sug-gested that scavenging activity against DPPH radical was enhanced by additional hydrolysis using another enzyme with an appropriate time.Generally,hydrolysates prepared by further hydrolyzing the hydrolysate obtained from theﬁrst step of hydrolysis with another protease for2h had a greater scavenging ability against DPPH rad-ical.3.2.2.ABTS radical scavenging activityABTS radical scavenging activity of protein hydrolysates pre-pared by two-step hydrolysis is shown in Fig.2(b).Only HAF and HAP showed the increases in ABTS radical scavenging activity,com-pared with their parent counterparts,HA.HAP with the hydrolysis time of2h for the second step had the highest activity(p<0.05). This suggested the enhancement of ABTS radical scavenging activ-ity when PCP was used for further hydrolysis of HA.However,the further hydrolysis of HF with another protease had no impact on the increases in activity(p>0.05).For HP,the decrease in activities was obtained when the second step of hydrolysis was performed (p<0.05).The results suggested that the second step of hydrolysis could slightly increase ABTS radical scavenging activity,depending on theﬁrst hydrolysate as well as the types of protease used for the second step.It was noted that the abilities of scavenging ABTS and DPPH radicals by hydrolysates were different.This might be due to the differences in ability of scavenging the different radicals,ABTS and DPPH,by the same peptide.3.2.3.Ferric reducing antioxidant power(FRAP)Fig.2(c)shows FRAP of protein hydrolysates prepared using two-step hydrolysis.The increases in FRAP were observed for all hydrolysates,when the second step of hydrolysis was applied.For HF and HP,when Alcalase was used for the second step of hydroly-sis,the continuous increases in FRAP were observed(p<0.05).For other hydrolysates,the hydrolysis time was the factor affecting the FRAP of resulting hydrolysate,depending on theﬁrst hydrolysate and the types of protease used for the second step of hydrolysis.The increase in FRAP with increasing hydrolysis time was coincidental with the increase in DPPH radical scavenging activity(Fig.2(a)). For HAF,HAP and HPF,the activities were increased with increas-ing hydrolysis time up to2h(p<0.05).For HFP,the hydrolysis time more than1h had the negative effect on FRAP(p<0.05).Gener-ally,further hydrolysis with another protease led to the increase in FRAP,but the activities of resulting hydrolysate were governed by hydrolysis time.3.2.4.Ferrous chelating activityFerrous chelating activities of hydrolysates prepared by two-step hydrolysis are shown in Fig.2(d).HFA and HPA showed the marked decrease in chelating activities when the second hydroly-sis was conducted(p<0.05).Alcalase used in the second step might generate peptides with the lower ability in Fe2+chelating.The abil-ities of HAF,HAP,HFP and HPF in chelating Fe2+ion were more pronounced,compared with their parent hydrolysate counterparts. The increases were observed when a certain time of the second hydrolysis was used.Among all protein hydrolysates,HFP showed the highest ferrous chelating activity,especially when the second step hydrolysis time of1h was used(p<0.05).3.3.ACE inhibitory activityThe inhibitory effect of all hydrolysates prepared using two-step hydrolysis against angiotensin I-convering enzyme(ACE)was determined as shown in Fig.2(e).For theﬁrst hydrolysate prepared using the different single proteases(0h),HF showed the highest ACE inhibition(11.44%)(p<0.05).Flavourzyme might produce the peptides with ACE inhibitory activity.Raghavan and Kristinsson [19]used Cryotin and Flavourzyme for hydrolysis of tilapia protein and found the higher ACE inhibition by hydrolysate prepared using Flavourzyme.When the second step of hydrolysis using another protease was performed,HFA and HFP showed the higher ACE inhi-bition when hydrolysis time of2and3h was used,respectively.HAP also showed high ACE inhibitory activity.Wu et al.[43]reported that shark meat treated with enzyme showed the higher ACE inhi-bition than that of untreated one.ACE inhibition by tilapia protein hydrolysate was reported,especially with increasing DH[19].Fur-thermore,smaller peptides are the better ACE inhibitors than the larger counterpart[19].The hydrolysis might release ACE inhibitory peptides in hydrolysate[43].Hydrolysates from muscle of differ-entﬁsh have been reported to possess ACE inhibitory activity,e.g. hydrolysates from tilapia protein[19],freshwater clam muscle[44], shark meat[43],Atlantic salmon,Coho salmon,Alaska pollack and southern blue whiting muscle[45].From the results,the use of Alcalase for theﬁrst hydrolysis and the use of PCP in the second hydrolysis for2h(HAP)yielded the resulting hydrolysate with the higher DPPH and ABTS radical scavenging activities and FRAP,compared to other hydrolysates (p<0.05).However HF hydrolyzed with PCP in the second hydrol-ysis step for1h(HFP)showed much higher chelating activity, compared with others(p<0.05).Therefore,HFP and HAP prepared with1and2h for the second step of hydrolysis,respectively,were。

药剂师英文简介

Most community pharmacists are salaried employees, but some are self-employed owners. About 24 percent of salaried pharmacists work in hospitals. Others work in clinics, mail-order pharmacies, pharmaceutical wholesalers, home health care agencies, or the Federal Government.

Many community and hospital pharmacies are open for extended hours or around the clock, so pharmacists may work nights, weekends, and holidays. Consultant pharmacists may travel to nursing homes or other facilities to monitor patients' drug therapy.

About 21 percent of pharmacists worked part time in 2004. Most full-time salaried pharmacists worked approximately 40 hours a week. Some, including many self-employed pharmacists, worked more than 50 hours a week.

Most pharmacists work in a community setting, such as a retail drugstore, or in a health care facility, such as a hospital, nursing home, mental health institution, or neighborhood health clinic.

甲磺酸伊马替尼晶型相关专利综述

广东化工2019年第13期·114·第46卷总第399期甲磺酸伊马替尼晶型相关专利综述王茜，夏宇祥(国家知识产权局专利局专利审查协作江苏中心，江苏苏州215163)Patent Review of Imatinib Mesilate’s Crystal FormWang Qian,Xia Yuxiang(Patent Examination Cooperation Jiangsu Center of The Patent Office,CNIPA,Suzhou215163,China)Abstract:Imatinib mesylate,the representative of the most excellent anti-leukemia drugs,is a kind of polymorphous substance.In order to break through the patent blockade of the original manufacturers,other applicants tend to focus on the preparation and product of new crystalline forms when submitting patents.In this paper,the patent applications related to imatinib mesylate crystalline forms are introduced,and the patent application proposals in related fields are given.Keywords:imatinib mesylate；crystalline forms；patent通常所说的伊马替尼是甲磺酸伊马替尼(imatinib mesilate)，商品名为“格列卫”(gleevec)，化学名为4-[(4-甲基-1-哌嗪)甲基]-N-[4-甲基-3-[[4-(3-吡啶)-2-嘧啶]氨基]苯基]-苯胺甲磺酸盐。

Risk-Based Qualification for 21 Century

9 March 2005 1 Rev 2 A White Paper on Risk-Based Qualification for the 21st CenturyForwardThe pharmaceutical industry is experiencing change at an incredible pace. Recent and significant product recalls, coupled with extreme pressure to reduce costs to the consumer while maintaining product quality, have brought great scrutiny to the industry. Once “economic proof” suppliers, manufacturers are now forced to compete on a quality and cost basis like never before. An area within our industry that is ripe for change is the facility and equipment qualification process. The current process is document intensive and does little to add value and provide assurance that the product manufactured is of the highest quality. The current process also does not follow a clear path of patient risk mitigation and clear product and process understanding. At the same time the present practices in most companies are very cost ineffective. There is a potential benefit in streamlining these practices by establishing industry standards and mechanisms, which ensure the quality and feasibility of a facility or equipment project from the initial user requirements to the final performance qualification.This whitepaper defines the principles upon which such practices should be based. It gives the directions for how ISPE, in cooperation with industry and regulators, aims to establish a risk-based approach to qualification. This is in accordance with the risk-based thinking that both industry and regulators are striving to attain.A Qualification Task Team, convened at the request of ISPE's International Leadership Forum in response to challenges from FDA, has drafted the attached White Paper on "Risk-Based Qualification for the 21st Century." The task team has received input from over three-dozen representatives of industry, equipment vendors, validation consultants, and regulators. Several white papers on this subject have been drafted and reviewed by staffs within pharmaceutical companies from August 2004 through January 2005. The attached white paper represents the evolution of ideas from the previous white papers, which culminated in an intensive workshop on the subject that was held at ISPE’s Tampa Conference in February 2005.Both the FDA and industry recognize that for most companies, qualification (IQ/OQ/PQ) has become an expensive, time-consuming process that adds little value in terms of ensuring equipment is fit for use in pharmaceutical manufacturing. This task team was challenged to recommend far-reaching changes to how qualification is structured and executed, in order to improve the industry's ability to deliver manufacturing facilities that meet product and process quality requirements in a timely and cost-effective manner. This whitepaper proposes that the equipment and facility commissioning and qualification activities should follow a risk-based approach based on the concept of risk mitigation for patients. For simple standard manufacturing equipment that are extensively used in the industry, the required effort should be far less than for complex custom-built equipment. For very quality-competent suppliers, the C&Q activities need not overlap much with the supplier's good engineering practices, to include supplier's own inspections and testing.Furthermore, the user's C&Q activities should depend on documented quality activities, including testing conducted by the supplier or integrator of an equipment or facility. This reliance could be justified based on an independent vendor or equipment certification scheme, or it could be supported by a risk assessment/ quality audit assessment by the customer in a manner similar to GAMP practices.This guidance should be in the form of internationally accepted standards through ASTM, possibly supplemented by ISPE technical documents. Such standards and guidance need to strip away the non-value added aspects that currently plague qualification practices, and focus instead on demonstrating, through PQ, that user requirements have been met. As a proposal to accomplish this, the team identified a number of principles upon which improvement can be based. It is the team's belief that if both industry and regulators can reach agreement in these areas, and provide clear guidance that all will adhere to, significant cost and time savings can be achieved, while at the same time improving the quality of systems and equipment for the manufacture of pharmaceutical products.Statement of the Current SituationIn March, 2001, ISPE published Volume 5 of the Pharmaceutical Engineering Guides for New and Renovated Facilities, “Commissioning and Qualification.” This guide was developed by an international team from pharmaceutical manufacturers and suppliers, and was subject to FDA review and endorsement. Many organizations within the industry have implemented some or all of the principles found within the Commissioning and Qualification Baseline Guide. Implementation has ranged from simple use of system impact assessments to eliminate qualification of indirect and no-impact systems, to full use of all guide principles, including component impact assessments focused on only those aspects that can affect product quality, and maximizing use of documented good engineering practices. In such cases, IQ/OQ protocols have been reduced to a few pages, identifying the 10% or so of components and functions necessary to meet user requirements and support quality manufacturing, referencing GEP documentation to avoid repeated inspections and testing.However, much improvement remains to be realized. As this paper is being written, a number of major pharmaceutical manufacturing capital projects, either planned or in progress, are struggling with issues such as:•The ISPE guide is not a regulatory document nor even regulatory guidance – fear that field inspectors won’t respect guide principles.•Projection of OQ protocols running thousands of pages per unit operation.•Unwillingness (due to fear of regulatory action and entrenched practices) to narrow focus of IQ/OQ/PQ to just that which can affect product quality.•Designs are not perfect – writing IQ against the detailed design (as per the V-model) leads to unnecessary and excessive deviations. Project teams areprocessing hundreds of such deviations, few or none of which results in fieldchanges, but instead are simply a paper exercise.•Pre-mature implementation of regulatory change control.•How to qualify PAT systems.The current situation needs to be improved in several major ways:1.The C&Q Guide provided the impact assessment process to reduce systemssubject to qualification (i.e., avoid the qualification “bullet”). The entire way inwhich we perform qualification needs to be revised so that it no longer representsa bullet to be avoided.2.Guidance or standards are needed that not only define a minimum standard, butalso state what is excessive.3.Guidance or standards are needed that are recognized as official, e.g., that carrythe same weight as an FDA or ICH guidance document.4.The FDA and international community have shifted to a risk-based approach tocompliance. This represents opportunity for further evolution of C&Q practices.Shifting the Qualification ParadigmA risk-based approach to qualification means that we must first understand our processes, and be able to identify and assess the risks to product quality inherent in a particular manufacturing process. We must then ensure that adequate risk-control mechanisms have been incorporated into the design; these risk-control mechanisms should be the focus of our qualification efforts. Items that pose little or no risk to product quality should not receive regulatory attention; the assumption of some risk is inevitable.The current qualification paradigm is based on a comprehensive verification of installation and operation against detailed functional requirements and detailed design specifications. This needs to shift to a focus on just those aspects that can directly affect product quality. Those aspects need to be determined using principles of risk management as espoused by FDA and international regulators. The project team and vendors, exercising good engineering practices and judgment, should confirm that the installation and operation is acceptable from an engineering perspective; IQ/OQ should audit the GEP/ commissioning work to confirm that those quality-impacting aspects have been achieved. The PQ is the true test of acceptability as defined by a process-based user requirements specification. More focus should be applied to design qualification, to ensure Quality by Design objectives have been met.Process Analytical Technology systems can be very sophisticated – these systems often use a custom process model and control scheme based on detailed process understanding. If we were to apply today’s “brute force” or “shotgun” method of qualification, excessive time and money will be required to validate these systems, which will discourage their implementation. New, innovative approaches to testing these systems, which include consideration of new knowledge gained during such testing, must be defined and deployed to the industry.Principles for 21st Century QualificationA group of over 20 representatives, primarily senior managers from industry, plus consultants and regulators, met in Tampa on 14 February 2005 to complete work on a set of guiding principles for this effort. The group agreed on 10 principles of good Commissioning and Qualification Practices:1.Focus on that which affects product quality. Qualifying equipment to putapproved protocols on the shelf is not the end goal; qualifying processes shouldbe the primary focus. To achieve this, definition and control of a process-basedUser Requirement Specification is an important function with quality impact. The primary quality and regulatory focus should be to ensure critical processparameters, critical functions, and critical design features that could affect productquality are defined and controlled.2.Requirements. User requirements, based on the process (and not on equipment orsystems), are the key to acceptability. The PQ is generally where userrequirements are confirmed as being satisfied. Hence, IQ/OQ are subordinate in importance to the PQ.3.Risk assessments, process development and experimental design are used toidentify critical features, functions, and critical process parameters. Thesebecome the basis for qualification (IQ/OQ/PQ). Having a solid processunderstanding will foster regulatory expectations for Quality by Design.4.Only critical process parameters will be used as the basis on which to define theformal “qualification information.” This should also include any physical design features or control functions that could impact the ability to clean, sterilize,sanitize, or properly manufacture the product, to the extent these activities impact product quality and safety.5.All activities must contribute value to the start-up and delivery of manufacturingcapacity. We won’t do anything just for the sake of regulatory compliance.Activities that are simply a paperwork exercise, resulting in no impact toinstallation, operation, or performance of systems, should be reduced oreliminated. Engineering judgment should be used to determine how to inspect or test specific features and functions of equipment and systems.6.Risk-based asset delivery. Different types of equipment and systems (custom,off-the-shelf, simple, complex, etc.) require different levels of attention to ensure quality. An approach to defining how much “good engineering practice” should be applied to a given item, based on risk of problems, should be applied rather than “cook book” lists of activities and documents. The GAMP categories use this approach for automation systems; similar approaches for equipment andsystems should be defined and described.7.Value-added documents. Documents serve a useful purpose of controllingactivities, they ensure completeness, and they serve as a record of what occurred.Only data which serves a useful purpose should be collected. Acceptability of documents should be based on technical merit; documents should not be “dressed up” to meet some imagined regulatory expectation. The operations andmaintenance groups should determine the acceptability of turnover packages, for it is they who will ultimately use them for on-going operations and maintenance.e of supplier documentation. Supplier’s standard inspection and testdocumentation may be used and no other documents be produced that duplicate this information, provided that documentation clearly shows the items of interest have been verified or tested in an appropriate manner. This is subject to thesupplier being of adequate quality.9.Test planning. Defined tests should only be carried out once (at an agreedlocation and by agreed parties, i.e. either by the supplier or by the manufacturingcompany, with accountabilities agreed upfront), unless there is a clear justification for undertaking further tests at a later stage of commissioning. Commissioningshould be a comprehensive activity, with IQ/OQ as an audit that commissioningverified the quality-impacting items. PQ may involve additional testing. Sometests may occur at different stages of development/ implementation and therefore appear to be repeats.10.Fostering innovation. Any program must remain flexible enough to apply soundand qualified scientific and engineering judgment based on the situation at hand.We must not be too prescriptive as to stifle innovation.Action PlanISPE’s Qualification Task Team has studied this subject for the past six months. We believe the above principles are sound. Some of them are already contained in the current Baseline Commissioning and Qualification Guide. However, it is our assessment that additional development of the Guide is needed to further streamline how industry performs these tasks. It is time to update the Guide and move further down the path to rationalize qualification practices in our industry.It is our assessment that consensus based standards would benefit the industry, by providing a clear set of expectations (as well as what is not expected) that both industry and regulators would adhere to. It is our understanding that consensus standards such as from ASTM are required by Executive Order to be used by US federal agencies. The development of these standards would, by their nature, ensure input and consensus from all interested parties. We believe that concise standards, backed up by an updated, complimentary Baseline Guide, would be valuable tools for industry and could cause a major improvement in how we deliver GMP manufacturing capacity.The Qualification Task Team also identified a number of concerns and pitfalls that we expect to encounter as we first debate the specifics of change, and then attempt to implement such change throughout the industry:1.Where does responsibility and accountability lie when equipment later provesfaulty?2.What is an appropriate level of oversight to ensure vendors and project teams aredoing what they are supposed to do?3.How will we deploy consensus standards to regulators world-wide, and what sortof transition acceptance phase will be needed?4.The focus on user requirements is good, but more clarification on what userrequirements are is needed.5. A process is needed to deal with legacy systems. How should be take advantageof knowledge of existing products and processes?6.How is “engineering judgment” gauged or applied? It is a sound concept butfurther definition will be needed.7.We will need to define required documentation. We should look to how otherworld-class industries achieve quality in their manufacturing facilities – what do they find adds value/ is necessary?8.We mustn’t be too prescriptive such that we stifle innovation and creativity.9.We mustn’t let standards be a barrier to entry of new vendors with noveltechnology or a better mousetrap.10.Significant organizational change will be necessary, and this may be the biggestbarrier to success. The QA organization will need to refocus away from thedetails of individual installation and operational test cases and protocols, andinstead focus on critical features and risks to product quality, based on processunderstanding and risk assessments and confirmed by a comprehensive PQ. ISPE’s Qualification Task Team requests endorsement from ILF as to the general basis for this initiative, endorsement for the 10 guiding principles, and endorsement regarding the recommended path forward – creation of qualification standards and concurrent updating of the Commissioning and Qualification Baseline Guide.Respectfully submitted, Qualification Task Team Steering CommitteeBrian Lange, Merck & Co., Inc.Bruce Davis, Astra ZenecaPaul D’Eramo, Johnson & JohnsonTim Howard, Eli Lilly and CompanyGert Moelgaard, NNE A/SRobert Chew, Commissioning Agents, Inc.ILF SponsorsRon Branning, GenentechUlrich Rudow, Johnson & Johnson。

新ISBP讲解

7.关于语言—language---有何变化现： it is expected that documents issued by the beneficiary will be in the language of the credit.
新：（1）Where a credit is silent with respect to the language of the documents, they may be issued in any language.
新ISBP讲稿
PRELIMINARY CONSIDERATIONS 与 GENERAL PRINCIPLES
1现在的一样，规定“申请人承担开证、修改申请不清晰的风险”。增加：开证行应确保信用证清晰、不矛盾。 The issuing bank should ensure that the credit or amendment it issues is not ambiguous or conflicting in its terms and conditions.
2. 关于“/” 新增加：if, nevertheless, a virgule is used and no context is apparent, this will allow one or any of the options. 比如：“Red/Black/Blue” with no further clarification will mean: only Red, or only Black, or only Blue, or any combination of them.（等于and）
5.增加了FROM的定义 600 第3条：（1）用于确定装运日时包括所述日期： “from” when used to determine a period of shipment include the date or dates mentioned. 比如：Shipment must be effected 20 days from 5.1.

欧盟GMP附录15：确认与验证(修订版英文+中文)培训课件

EUROPEAN COMMISSION ENTERPRISE DIRECTORATE-GENERALSingle market, regulatory environment, industries under vertical legislationPharmaceuticals and cosmeticsBrussels,30 March 2015EudraLex欧盟药品管理法Volume 4EU Guidelines forGood Manufacturing Practice forMedicinal Products for Human and Veterinary Use第四卷欧盟人用和兽用药品GMP指南Annex 15: Qualification and Validation附录15:确认和验证Legal basis for publishing the detailed guidelines:Article 47 of Directive 2001/83/EC on the Community code relating to medicinal products for human use and Article 51 of Directive 2001/82/EC on the Community code relating to veterinary medicinal products. This document provides guidance for the interpretation of the principles and guidelines of good manufacturing practice (GMP) for medicinal products as laid down in Directive 2003/94/EC for medicinal products for human use and Directive 91/412/EEC for veterinary use.发布该细化指南的法律依据:人用药物欧共体法案指令2001/83/EC第47章和兽用药物欧共体法案指令2001/82/EC第51章。

欧洲药典CEP证书修订更新规定指南中英对照版

欧洲药典CEP证书修订更新规定指南中英对照版Date of implementation: 1 March 2010Introduction:The holder of a Certificate of suitability shall inform the EDQM of any change to the information in the certification dossier by sending an application form and all necessary documents demonstrating that the conditions laid down in the present guideline are met.Classification of changesThe changes have been classified in three categories (notification/minor/major) depending on the potential impact of the change on the quality of the final substance. These three categories are based on those (IA-IAIN/IB/II) of the Commission Regulation (EC) No 1234/2008 concerning the examination of variations to the terms of marketing authorisation for medicinal products for human use and veterinary medicinal products.Any change not classified as a notification or a major change should be classified as a minor change except in the following cases where a new application should be submitted:- addition of a new route of synthesis and/or a new manufacturing site where the specifications of the final substance are different from the one already approved- transfer to a new holder that is not the same legal entity as the approved one, where the transfer does not occur because of a merger or because the company is sold, and where the manufacturer does not take out the Certificate of suitability in their own name.The changes related to Ph. Eur. monograph revisions or any other regulatory requirements are treated separately andgenerally initiated by the EDQM.执行日期：2010年3月1日介绍：欧洲药典适用性证书持有人必须向EDQM报告所有与申报文件有关的变更，申报时应填写申请表格和所有必要的资料，证明变更符合现行指南的规定。

Danaher-harvard business review 下(中文)

下半部分：英文原文8-17页Danaher Business System (DBS)At the core of Danaher’s operating model and acquisition strategy was the Danaher Business System (DBS). The firm’s investor presentations described DBS as “defining our high-performance culture. DBS is who we are and how we do what we do.” Outsiders noted that DBS “is a set of management tools borrowed liberally from the famed Toyota Production System. In essence it requires every employee, from the janitor to the president, to find ways every day to improve the way works get done.”34 While such programs were “de rigeur for manufacturers for years, the difference at Danaher (was) the company started lean in 1987, one of the earliest U.S. companies to do so, and it has maintained a cultish devotion to making it pay off.”35 The lean approach replaced a traditional “batch-and-queue” manufacturing system with a “single-piece flow” that minimized inprocess time and so reduced inventory and other overhead costs. “In a typical Danaher factory, floors are covered with strips of tape indicating where everything should be, from the biggest machine to the humblest trash can. Managers determine the most efficient place for everything, so a worker won’th ave to walk an extra few yards to pick up a tool, for instance.”36Danaher业务管理系统（DBS）Danaher的操作模型和并购计划的核心便是Danaher业务管理系统（DBS）。

欧盟GMP附录15确认和验证中英文新版

欧盟GMP附录15确认和验证欧盟GMP附录15确认和验证ANNEX 15 附件15Qualification and Validation确认和验证Table of Contents 目录1. Qualification and Validation 确认和验证2. Planning for Validation 验证计划3. Documentation 文件4. Qualification 确认5. Process Validation 工艺验证6. Cleaning Validation 清洁验证7. Change Control 变更控制8. Revalidation 再验证9. Glossary 术语表Qualification and Validation 确认和验证Principle 原理1.This Annex describes the principles of qualification and validation which are applicable to the manufacture of medicinal products. It is a requirement of GMP that manufacturers identify what validation work is needed to prove control of the critical aspects of their particular operations. Significant changes to the facilities, the equipment and the processes, which may affect the quality of the product, should be validated. A risk assessment approach should be used to determine the scope and extent of validation.1.本附件描述了确认和验证的原理，适用于医药产品的生产者。

WHO号技术报告附件药物生产技术转移指南中英文

WHO第961号技术报告附件7 药物生产技术转移指南（中英文1/4）2013-09-29 14:16:27|分类：WHO|字号订阅World Health OrganizationWHO Technical Report Series, No. 961, 2011WHO第961号技术报告附件7 药物生产技术转移指南Annex 7附件7WHO guidelines on transfer of technology in pharmaceutical manufacturingWHO药物生产技术转移指南1. Introduction介绍2. Scope范围3. Glossary术语4. Organization and management组织和管理5. Production: transfer (processing, packaging and cleaning)生产：转移（工艺、包装和清洁）6. Quality control: analytical method transfer质量控制：分析方法转移7. Premises and equipment厂房设施和设备8. Documentation文件9. Qualification and validation确认和验证References参考文献1.Introduction介绍These guiding principles on transfer of technology are intended to serve as a framework which can be applied in a flexible manner rather than as strict rigid guidance. Focus has been placed on the quality aspects, in line with WHO’s mandate.本指南中关于技术转移的原则意在作为一个框架，以不同方式应用，而不是一个需要严格遵守的指南。

欧洲药典4.5的执行

需要进一步的支持性文件来支持那些药典专论杂质列表中未规定的杂质的地方，在所附的列表中都被标上了‘3’。在这种情况下，标上了 ‘2’的文档也是需要提供的，并要有图谱，和内控方法进行交差验证，要有杂质论述和药典专论杂质列表的对比。
Translated and prepared by HUKAI
本程序是不收费的，除非在递交上述文件的同时递交了你的文件的行政和/或技术更改(主要的或次要的)资料，或同时递交了证书的更新。
Translated and prepared by HUKAI
Yours faithfully, C.POUGET Head of Certification Unit Enclose: list of revised texts of European Pharmacopoeia Supplement 4.5 for implementation 1 July 2003.
含有一些修订过的专论的欧洲药典增补 4.5将于2003年7月开始执行。作为证书持有者，你有责任及时地就相关的原料药更新质量规格和测试程序以执行2003年7月的新版或专论修订过的内容。
Translated and prepared by HUKAI
You hold a certificate for a substance for which a technically revised monograph has been published (section ‘REVISED TEXTS’, see list enclosed), you should proceed as follows:
In the cases ‘2’ and ‘3’ you are expected to reply to this letter and provide the supporting documents as soon as possible and in any case no later than the implementation date of the revised monograph.

IPEC全文译稿中英文对照

FOREWORD 前言The quality of excipients is critical to assure the safety, quality and efficacy of medicines. Excipients have a wide range of applications and are essential components of the drug product formulation. Characteristics that excipients impart to formulated drug products include cosmetic appearance, stability and delivery of the active ingredient. Therefore, applying appropriate Good Manufacturing Practice (GMP) principles to excipients is essential. 辅料的质量对保证药品的安全、质量和功效是至关重要的。

辅料的应用范围广泛，是药品生产配方的基本成分。

辅料的特性直接影响药品的配方，包括化妆品的外观、稳定性和活性成分的输送。

因此，应用适当的GMP 规则是辅料的基础。

In contrast to finished dosage forms and Active Pharmaceutical Ingredients (APIs), there are no specific GMP regulations for excipients. In addition, there are a large number of applications of this diverse range of materials which makes the development of excipient GMP guidelines challenging. However, there is a general expectation that excipients are manufactured to recognised GMP principles.与制剂和原料药相比，没有明确的针对辅料的GMP的规则。

衍生品新披露要求【外文翻译】

外文翻译外文题目Derivatives: New Disclosures Required 外文出处 CPA JOURNAL外文作者Barbara Apostolou and Nicholas G.Apostolou原文：Derivatives: New Disclosures RequiredIn March 2008,FASB issued SFAS161,Disclosures about Derivative Instruments and Hedging Activities,which expands disclosures about derivative instruments and hedging activities. This new statement amends SFAS 133,Accounting for Derivative instruments and Hedging Activities, which established the basic rules governing derivatives. The new requirements imposed by SFAS 161 must be applied prospectively for interim periods and fiscal years beginning after November 15,2008.SFAS 133 was issued in 1998 and has been substantially amended three times in an attempt to keep pace with the increasing complexity of derivative transactions and the evolving nature of hedging activities. SFAS 133 was criticized for inadequately disclosing information about how a company's derivative and hedging activities affect its financial position, income, and cash flows. Consequently, SFAS 161 provides enhanced disclosures about the reasons for using derivatives, the accounting for derivatives and hedging activities, and their effect on the company's financial position, financial performance,and cash flows.The ensuing discussion will describe the objectives of holding and issuing derivatives, the risk exposure assumed by their issuance, the accounting for these instruments, and their effect on the issuer's financial statements. In addition, the changes made by SFAS 161 to the existing disclosure requirement mandated by SFAS 133 are described. FASB was dissatisfied with SFAS 133's disclosure requirements regarding an entity's derivative and hedging activities and their impact on financial position. financial performance.,and cash flows. The intent of SFAS 161 is to enhance these disclosures and thereby improve the transparency of financial reporting.BackgroundDerivatives are financial instruments with a value derived from fluctuations in the share of an underlying asset, liability, interest rate，exchange rate, or index. SFAS 133 describes the essential characteristics of a derivative, which are summarized in Exhibit 1.A derivative's value stems from changes in the value of a related asset or liability. The rate or price that relates to the asset or liability included in the derivative is referred to as an underlying. An underlying can assume a variety of forms, including an interest rate, seeurity price,commodity price, foreign exchange rate,index of prices or rates, or other variable.For example, the underlying in an option to buy a stock at $40 per share is the $40 fixed price of the slock and not the stock itself. Changes in the underlying price cause the value of the derivative to change.The value of a derivative also is dependent upon the number of units specified in the derivative contract. For example, the value of an option to buy stock at $40 per share depends not only upon the stock price, but also upon how many shares can be purchased under the terms of the option. The total value of a derivative at any given time is determined by both the underlying price and the notional amount.Derivatives typically require little or no initial investment because the investment is not in the actual asset or liability underlying the derivative, but rather in the change in value attributable to an underlying. For example, if the price of a stock increases from $40 to $42 and an investor holds an option to buy at $40, the value of the option has increased by at least $2. If the investor owned the underlying stock. its total value would also obviously increase. The key difference is that the option holder needs to make little or no initial investment, while the owner of the stock must make a significant investment.Net settlement refers to the ability to settle a derivative contract by exchanging cash or entering into an offsetting contract without actually buying or selling the related asset or liability. For example, a futures contract is an agreement between two parties that commits one party to sell a commodity or security to the other at a given price and on a specified future date. The sale of a contract that was previously purchased liquidates a futures contract in the same way that the sale of 100 shares of GM stock would liquidate an earlier purchase of 100 shares of GM stock. Very few speculators have the desire lo take delivery of 5,000 bushels of Corn,60,000 pounds of soybean oil, or even $ 100,000 in Treasury notes. As a result, gains and losses are generally realized by buying or selling offsetting futures contracts prior to the deliverydate.Derivatives can be based upon currencies,commodities, government or corporate debt, home mortgages, stocks, interest rates, or any number of indices (including an index of weather conditions).Although there are hundreds of different types of derivatives, they can be classified into three main classes:• Futures/Forwards—contracts to buy or sell an asset at a specified future date at a price specified today;• Options—contracts giving the holder the right, but not the obligation, to buy or sell an asset at a specified price on or before a given date: and•Swaps—where two parties agree to exchange one stream of cash flows against another stream.The use of derivatives can be divided into three main areas: hedging, dealing,and speculating. Traditionally, hedging was undertaken by businesses to limit their risk exposure. For example, a wheat farmer and bread maker could enter into a futures contract to exchange cash for wheat in the future. Both parties have reduced their risk because the farmer knows the price to be received and the bread maker knows that wheat will be available. More recently,speculators have come to dominate the market for derivatives. In most financial derivatives markets, the value of speculative trading far exceeds the value of hedge trading. Dealers, such as banks, are the ones creating the derivatives contracts and making money from the difference between the buying (bid) and selling (offer) price. Although most dealers also trade derivatives for their own account, this proprietary trading is usually kept separate from their market-making function.Derivatives can be traded over-the-counter(OTC) or on an exchange. OTC derivatives are traded directly between two parties, without using an exchange to facilitate the trade. The OTC derivatives market is staggeringly large and until recently continued to grow at a dynamic pace. According to the Bank for International Settlements, the total outstanding notional amount as of June 2007 was a mind-boggling $516 trillion (see Exhibit 2). Notional amount is defined by FASB in SFAS 133 (para. 540) as: "A number of current units, shares, bushels, pounds, or otherunits specified in a derivative instrument." In other words, the notional amount serves as the basis for calculating cash flows under the contract. Although the notional amount gives an indication of the size of the derivatives market, it is not a fair measure of the risk associated with derivatives. For example,a derivative contract might involve a 10-year Treasury rate on $10 million in exchange for a floating rate. Although the notional face value of the contract is for $10 million, this amount does not change hands. For each payment period, the 10-year Treasury rate is multiplied by $10 million in order to calculate the required interest payment.Exchange-traded derivatives are those derivatives with standardized terms that are traded on organized derivatives exchanges. Prominent examples are the options and futures contracts traded on the largest U.S. derivatives exchange. CME Group. The exchange acts as an intermediary between two parties and assures that there is a counterparty that will honor the trade, regardless of the movement of the market.Although the accounting for derivatives used in hedging activities can be complex, the rules governing stand-alone derivatives can be easily summarized: • All derivatives must be carried on the balan ce sheet at fair value. FASB defines fair value as the amount at which an asset (liability) could be bought (incurred) or sold (settled) in a current transaction between willing parties. Typically, the best evidence of fair value is quoted market prices.• The amount shown for the derivative has to be adjusted to fair value on each balance sheet date.• The change in the fair value of the derivative from the previous balance sheet date is reflected as a holding gain or loss on the income statement. The only exception to the recognition of a holding gain or loss is for derivatives that qualify as hedges.FASB has specified exacting criteria for classifying a derivative as a hedge: Management must provide sufficient documentation at the beginning of the hedge term to identify the objective and strategy of the hedge, the hedged instrument and the hedged item, and how effective Ihe hedge is in eliminating a specific market risk for a specific hedged item. Effectiveness is the ability of the hedged instrument to offset any change in the fair value or cash flows of the hedged item. Effectiveness is testedwhen the hedge is initiated, every three months thereafter, and each time financial statements are prepared. To be characterized as effective. the derivative instrument must have the ability to offset between 80% and 125% of the changes in the fair value or cash flows of the hedged item.Derivatives that qualify as hedges can be classified as follows:• Fair value hedges are designated to hedge the exposure to chang es in the fair market value of an existing asset or liability, or a firm commitment (e.g., buying or selling inventory). The gains and losses on the hedged asset or liability and the hedging instrument are currently recognized in the income statement.• Ca sh flow hedges are designated to hedge the exposure to changes in cash flows of an existing asset or liability (e.g.,future interest payments on variable-interest debt), or an anticipated transaction (a futures contract that a corn miller will pay in two months for corn). The distinction between highly effective and ineffective hedges determines when gains and losses on cash flow hedges are recognized in income. The gain or loss on the ineffective portion of the hedge is reported in current income. The gain or loss on the effective portion of the hedge flows should be reported in other comprehensive income (i.e., not included in net income).• Foreign currency hedges are designated to hedge the exposure to changes in currency exchange rates of an existing asset or liability, a firm commitment, a forecasted transaction, or a multinational company's net investment in a foreign operation. In general. FASB applies the fair value and cash flow hedge accounting rules to foreign currency exchange exposure. However, a derivative designated as hedging a net investment in a foreign operation has its gain or loss reported in other comprehensive income (i.e., not included in net income).Derivative RiskAs noted above, the derivatives market has exploded in recent years, with financial institutions reaping huge profits by convincing clients of the value of derivatives as a way to off-load or manage market risks. Warren Buffett warned in Berkshire Hathaway. Inc.'s 2002 annual report that such highly complex financial instruments are time bombs and "financial weapons of mass destruction" that couldharm not only their buyers and sellers, but the whole economic system. Some derivative contracts. Buffett wrote, appear to have been devised by "madmen." SFAS 133 requires derivatives to be marked-to-market and the gain or loss to be reflected in income. The deterioration of credit markets over the past year has made it very difficult to determine the market value of some derivatives.The ongoing worldwide credit crunch began with the bursting of the U.S. housing bubble in 2007. Rising housing prices in recent years led to lenders offering borrowers mortgages with minimal down payments and adjustable-rate mortgages. The incentives given to borrowers by financial institutions resulted in a redhot housing market that was eventually overbuilt. The surplus inventory of homes resulted in a decline in new home construction and in housing prices. Default rates on subprime and adjustable-rate mortgages made to higher-risk borrowers skyrocketed, creating a ripple effect that impacted mortgage-based securities and related derivatives, precipitating a liquidity crisis in the banking system.Derivatives involve risks so great that even a single rogue trader can imperil a major financial institution. The danger was highlighted by the SociétéGénérale incident,reported in January 2008, which resulted in the largest trading fraud—$7.2 billion — ever carried out by a single person .Société Générale is a major French bank, operating in 77 countries with 120,000 employees. A trader, Jerome Kerviel,previously worked as a back-office employee for the bank, where he gained an intimate knowledge of SociétéGénérale's processing and control procedures. Kerviel was promoted to a front-office position, where he engaged in arbitrage transactions intended to profit from small differences between a futures index and the equity futures upon which the index is based. Instead of properly hedging his positions, however, Kerviel allegedly used his knowledge of the lapses in SociétéGénérale's control environment to take massive unhedged positions in futures. In a short period of time, he had committed Stxiété Genénile to more than $70 billion in notional positions, according to the bank, which exceeded the net worth of StxriétéGénérale. A suspicious email from a large bank confirming trades precipitated an investigation into Kerviel's activities.The event that initially shocked the financial world was the collapse of Bear Stearns, one of ihe largest U.S. investment banks. In a deal reached on Sunday, March 16. 2008. J.P. Morgan Chase agreed to pay $2 per share (later raised lo $10 per share) to buy all of Bear Steams. The market price per share of Bear Steams was over $30 the previous Thursday, and over $170 in January 2007. The Federal Reserve expedited the bailout by issuing a nonrecourse loan of $29 billion to cover losses in Bear Steams' investments in mortgage-backed securities and other derivatives.Federal Reserve Chairman Ben Bernanke testified before the Joint Economic Committee that the default of Bear Steams could have led to a worldwide systemic financial crisis.Source:CPA JOURNAL,2008:28-30译文：衍生品:新披露要求2008年3月，美国财务会计准则委员会发布财务会计准则公告161号，揭露了衍生工具和套期保值活动，扩大了衍生工具和套期保值活动的披露要求。

ProbioticsandAnt...

Probiotics and Antimicrobial Peptides:The Creatures’and Substances’Future in the Twenty-First Century:An Opinion LetterMichael L.ChikindasPublished online:16May 2014ÓSpringer Science+Business Media New York 2014While the number of scientiﬁc publications related to health-promoting bacteria and antimicrobials of proteina-ceous nature is growing,there is an obvious need for a better comprehension of what these studies actually pro-vide the community and which directions these studies should be taking.With no intention of neglecting funda-mental science (in which true knowledge helps separate fact from ﬁction,and eventually ﬁnds its practicality),I would still like to step on a dangerous path and speculate on the near-future progress in the aforementioned ﬁeld of science with emphasis on possible beneﬁcial outcomes.The number of marketed food products and dietary supplements labeled as ‘‘probiotics’’or ‘‘containing probi-otic bacteria’’is virtually incalculable.In 2010,the global probiotic market was estimated at $22.6billion and is projected to reach $28.8billion by 2015[1].Very few microorganisms have been subjected to thorough in vitro studies conﬁrming their speciﬁc health-promoting activity,and even fewer were subsequently subjected to and passed the appropriate human trials.A few studies of interest include those which investigated the strains Lactobacillus plantarum 299v,Lactobacillus casei Shirota,Lactobacillus rhamnosus GG should probably be mentioned—yet even the latter was reported on several occasions as associated with bacterial sepsis (for review see:[2]).Additionally,probiotics can be truly dangerous,posing as a wolf in the sheep’s clothing:They have been linked to an increase in mortality rate if administered to severely immunocompro-mised patients [3].Those interested in ‘‘legal’’probiotics may consider looking at the list of so-called ‘‘CommercialStrains Sold As Probiotics’’[4]where the phrase ‘‘sold as’’is perhaps the most telling in indicating the less that ﬁrm certainty about these strains being large-scale human trial-proven as delivering measurable beneﬁts.Still,there is WHO guidance on probiotics [5]and a brief overview of the claims discussed as required and sufﬁcient for placing a microorganism into a group of ‘‘good fellows’’[6].Finally,NIH Grant 5R01HG005171-02resulted in a document entitled ‘‘Federal Regulation of Probiotics:An Analysis of the Existing Regulatory Framework and Recommendations for Alternative Frameworks’’[7]which only conﬁrms the murky reality of less than stringent and frequently discon-nected regulations currently being observed.As of December 14,2012,the European Food Safety Authority (EFSA)posed serious and soundly justiﬁed [8]restrictions on labeling food products containing probio-tics—the matter was strongly opposed by the major world manufacturers of microorganism-fermented/containing food products and by the large-scale manufacturers of microbial cultures [9].Currently,there are two (European and North American)different statuses resulting in two different approaches addressing the real problem:how to separate the truly beneﬁcial microorganisms from the crowd of overall harmless but ‘‘indifferent’’bugs broadly used to make some products attractive to the consumers [10].All of these leave a few questions and hopefully some answers.The diversity and multiplicity of the human microbiota cannot be ignored.While broad generalizations can be made concerning the frequency of certain species in our body’s ecological niches,they are genetically variable [11].Certainly,the common strains associated with healthy human subjects should do no harm when ingested with a food product or administered otherwise.The additional health beneﬁts are somewhat doubtful;why should they deliver anything in addition to what they already provide?M.L.Chikindas (&)School of Environmental and Biological Sciences,Rutgers State University,New Brunswick,NJ 08901,USA e-mail:************************.edu Probiotics &Antimicro.Prot.(2014)6:69–72DOI 10.1007/s12602-014-9161-7Also,if a product claims to contain live,active and health-promoting bacteria but does not deliver the expected effect to some(few?many?)consumers,should this claim be withdrawn or carefully revised(e.g.,[12])?Clearly,pro-biotics showing statistically signiﬁcant and reproducible positive effects in clinical[13]or other large-scale trials [14,15]should keep their well-deserved and earned name. Furthermore,while the society’s primary concern is focused at the use of probiotics for various human beneﬁts, the issue is similarly important when it concerns animals of agricultural importance[16],aquacultures[17],and poultry [18].Therefore,it is only logical to expect an increase in the number of studies which focus on understanding the of mechanisms of action of health-promoting bacteria which show positive and reproducible results in clinical and other, speciﬁc large-scale environments when used under the supervision of medical or other appropriate for the study/ application professionals.In addition,with already numerous statistically sound and highly reproducible results,the science of probiotics for the improvement in animal husbandry will continue blooming.As for the spe-cies Homo sapiens,we still have toﬁnd how diverse our individual and group(ethnic,social,age,gender,etc.) microbiomes are(e.g.,see:[19]).While probiotics deliver health-promoting assistance to eukaryotic organisms in many ways(e.g.,immunomodu-lation,cancer prevention,cholesterol reduction,etc.),the ability to produce and utilize a variety of‘‘weapons’’active against pathogens and other unwelcomed ecological niche-intruders is well documented.Healthy human lactic acid bacteria produce several potent antimicrobials such as hydrogen peroxide,bacteriocins,and of course,lactic acid. In fact,approximately60%of healthy vaginal lactobacilli are reported as having all three defense mechanisms[20]. Therefore,natural antimicrobial proteins go hand-in-hand with probiotics in their defense against numerous pathogens.Natural antimicrobials of proteinaceous nature and particularly those ribosomally produced by bacteria and called bacteriocins is a youngﬁeld of science.However, two of the most studied bacteriocins,namely nisin and pediocin PA-1/AcH,have already established theirﬁrm presence in line with other natural food preservatives, either claimed as an active substance in formulation or being present in a fermented active extract[21].Bacte-riocins are often inseparable from probiotics,being one of the weapons used by these healthy bacteria in their combat against pathogens[22].The last few years have seen a trend in broadening exploration of possible use of bacteriocins in personal care and medical applications [23].Bacteriocins from Gram-positive bacteria are increasingly reported as active against Gram-negative microorganisms when used in combination with syner-gistically acting substances[24],or even alone[25]. Numerous studies published indicating increased activity and/or broader spectrum of engineered bacteriocins[26]. Recent reports demonstrate antiviral activity of some bacteriocins(e.g.,[27]).All of these and particularly growing number of reports on synergy of bacteriocins with other stressors have catalyzed the exploration of bacteriocins as‘‘helpers’’with conventional antibiotics, wherein antimicrobial proteins can improve the activity of the drug and decrease the rate of the targeted microor-ganisms’resistance to combinational therapy(e.g.,[28, 29]).The availability and convenience of using database on antimicrobial proteins is of a great assistance in these studies[30].Since bacteriocins are biodegradable and most of them are overall positively charged,hydrophobic substances,to assure their efﬁciency in challenging environment of eukaryotic organisms,proper delivery systems should be considered[31].And,while bacterio-cins will probably not replace conventional antibiotics [32],they may serve as a valuable asset to already mar-keted drugs,especially in considering safety features of bacteriocins such as biodegradability and lack of immu-nogenicity[33].Executive SummaryNota bene:To keep this opinion letter concise,in the previous pages,some of the bullets were not elaborated on.•Probiotics should be distinguished between food fer-menting and otherwise safe,naturally occurring microorganisms.•There is an urgent need for revisiting the deﬁnition of probiotics and for their clear classiﬁcation based on functions and deliverables as conﬁrmed in large-scale statistically sound trials.•It is logical to expect more research focused on probiotics for following applications:•medicinal(cancer prevention,treatment of dysbact-eriosis,immunomodulation,anti-viral prophylaxis,etc.)•this will include bioengineered functional probiotics;•personal care;•animals of agricultural importance;•poultry;•aquacultures.•Novel approaches for formulation and delivery of probiotics will continue to emerge.•Bacteriocin-producing probiotics will be gaining more attention as well as probiotics with well-characterized and fully understood functions.•In addition to already discovered,novel bacteriocins will continue to be explored for following applications:•food preservations;•antimicrobials in personal care formulations;•medicinal use in combination with conventional antibiotics and/or other nature-derived synergisti-cally acting antimicrobials;•anti-viral formulations;•spermicidal.•To use bacteriocins in the most efﬁcient manner in various applications,there is a growing demand for understanding of kinetics of bacteriocin–microbe inter-action.This will continue being addressed at several levels:•conditions simulating natural environment(low number of cells and low concentration ofbacteriocin);•bacteriocins as stressors for bioﬁlm prevention and their activity against formed bioﬁlms.•A signiﬁcant effort is expected to be dedicated to design and study of functional vehicles for targeted and controlled delivery of bacteriocins and synergistically acting stressors.Acknowledgments The author is grateful to Dr.Montville, Dr.Matthews,and Ms.Cavera for their critical reading and discussion.Conﬂict of interest The author declares that he has no conﬂict of interest.References1.Anonymous(2010)Global probiotic market to grow:analyst.FLEXNEWS.27Sept2010htinen SJ,Boyle RJ,Margolles A et al(2009)Safety assess-ment of probiotics.Prebiotics and probiotics science and tech-nology.Springer Science&Business Media,LLC,New York, pp1193–12353.Besselink MG,van Santvoort HC,Buskens E et al(2008)Probioticprophylaxis in predicted severe acute pancreatitis:a randomised, double-blind,placebo-controlled ncet371(9613):651–659 4.California Dairy Research mercial strains soldas probiotics./home/checkoff-investments/uspro biotics/products-with-probiotics/#commercial.Accessed24April 20145.Guidelines for the Evaluation of Probiotics in Food:Joint FAO/WHO Working Group meeting,London Ontario,Canada,30 April-1May2002.http://www.who.int/foodsafety/fs_manage 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期货交易管理暂行条例法律英语

期货交易管理暂行条例法律英语〔1999年5月25日国务院第18次常务会议通过，自1999年9月1日起施行，1999年6月2日中华人民共和国主席令第267号公布，自1999年9月1日起施行〕(Adopted at the 18th Executive Meeting of the State Council on May 25, 1999, promulgated by Decree No. 267 of the State Council of the People's Republic of China on June 2, 1999, and effective as of September 1, 1999 )第一章总那么Chapter I General Provisions第一条为了规范期货交易行为，加强对期货交易的监督治理，爱护期货市场秩序，防范风险，爱护期货交易各方的合法权益和社会公共利益，制定本条例。

Article 1 These Regulations are formulated in order to regulate the conduct of trading in futures, enhance the supervision and administration of futures trading, maintain order in the futures market, prevent and minimize risks, and protect the legitimate rights and interests of the parties trading in fixtures and the public interests of society.第二条从事期货交易及其相关活动的，必须遵守本条例。

Article 2 The conduct of any trading in futures and its related activities shall comply with these Regulations.第三条从事期货交易活动，应当遵循公布、公平、公平和诚实信用的原那么。

欧盟GMP附录1计算机系统中英文对照

.EUROPEAN COMMISSION欧盟委员会HEALTH AND CONSUMERS DIRECTORATE-GENERAL卫生与消费者协会Public Health and Risk Assessment公共卫生与风险评估Pharmaceuticals药品Brussels,SANCO/C8/AM/sl/ares(2010)1064599EudraLexThe Rules Governing Medicinal Products in the European Union 欧盟药品生产规范Volume 4卷4Good Manufacturing PracticeMedicinal Products for Human and Veterinary Use人用与兽用药品良好生产管理规范Annex 11: Computerised Systems附件11：计算机系统Legal basis for publishing the detailed guidelines: Article 47 of Directive2001/83/EC on the Community code relating to medicinal products for human use and Article 51 of Directive 2001/82/EC on the Community code relating to veterinary medicinal products. This document provides guidance for the interpretation of the principles and guidelines of good manufacturing practice (GMP) for medicinal products as laid down in Directive 2003/94/EC for medicinal products for human use and Directive 91/412/EEC for veterinary use.依法发布的具体指导方针：2001/83/EC第47条人用药品规范和2001/82/EC第51条兽用药品规范。

药典的协调

药典的协调Pharmacopoeial harmonisation…an industry perspectiveDr Stephen W. CookEC-China High Level Regulatory DialogueWorkshop on GMP/Pharmacopoeia/APIs11-12th July 2012药典的协调…工业界展望中国-欧盟药品注册高层对话GMP/药典/APIs研讨会11-12th July 2012 / 2012年7月11 -12日Introduction ?Harmonised pharmacopoeia help to ensure patient access to high quality medicines and vaccines worldwide As part of the 2015 ChP development, Industry recognises;–Increased opportunity to provide comments for 2015 ChP revisions–Greater clarity on the drafting procedures–Engagement by CPC with other pharmacopoeial bodies–Reference to other pharmacopoeias during drafting of monographs ?Industry encourages this development and would like to raise several points for discussionIntroduction引言?协调药典有助于保证患者使用世界上的高质量药品和疫苗。

作为2015版中国药典发展的一部分，企业认识到：–提供2015版中国药典修订说明的机会增加；–起草规程更清晰；–CPC与其它药典机构的参与；–专论起草过程中参考其它药典。