Bacterial communities and enzyme activities of
PAHs polluted soils
V.Andreoni a,*,L.Cavalca a ,M.A.Rao b ,G.Nocerino b ,S.Bernasconi c ,E.Dell ?Amico a ,M.Colombo a ,L.Gianfreda
b
a
Dipartimento di Scienze e Tecnologie Alimentari e Microbiologiche,Universita
`degli Studi,Via Celoria 2,20133Milano,Italy
b
Dipartimento di Scienze del Suolo,della Pianta e dell’Ambiente,Universita
`di Napoli Federico II,Via Universita
`100,80055Portici,Napoli,Italy c
Dipartimento di Chimica Organica e Industriale,Universita
`degli Studi,Via Venezian 21,20133Milano,Italy Received 11July 2003;received in revised form 1June 2004;accepted 10June 2004
Abstract
Three soils (i.e.a Belgian soil,B-BT,a German soil,G,and an Italian agricultural soil,I-BT)with di?erent prop-erties and hydrocarbon-pollution history with regard to their potential to degrade phenanthrene were investigated.A
chemical and microbiological evaluation of soils was done using measurements of routine chemical properties,bacterial counts and several enzyme activities.The three soils showed di?erent levels of polycyclic aromatic hydrocarbons (PAHs),being their contamination strictly associated to their pollution history.High values of enzyme activities and culturable heterotrophic bacteria were detected in the soil with no or negligible presence of organic pollutants.Genetic diversity of soil samples and enrichment cultures was measured as bands on denaturing gradient gel electrophoresis (DGGE)of ampli?ed 16S rDNA sequences from the soil and enrichment community DNAs.When analysed by Shan-non index (H 0),the highest genetic biodiversity (H 0=2.87)was found in the Belgian soil B-BT with a medium-term exposition to PAHs and the poorest biodiversity (H 0=0.85)in the German soil with a long-term exposition to alkanes and PAHs and where absence,or lower levels of enzyme activities were measured.For the Italian agricultural soil I-BT,containing negligible amounts of organic pollutants but the highest Cu content,a Shannon index =2.13was found.The enrichment of four mixed cultures capable of degrading solid phenanthrene in batch liquid systems was also studied.Phenanthrene degradation rates in batch systems were culture-dependent,and simple (one-slope)and complex (two-slope)kinetic behaviours were observed.The presence of common bands of microbial species in the cultures and in the native soil DNA indicated that those strains could be potential in situ phenanthrene degraders.Consistent with this assumption are the decrease of PAH and phenanthrene contents of Belgian soil B-BT and the isolation of phenanth-rene-degrading bacteria.
From the fastest phenanthrene-degrading culture C B-BT ,representative strains were identi?ed as Achromobacter xylosoxidans (100%),Methylobacterium sp.(99%),Rhizobium galegae (99%),Rhodococcus aetherovorans (100%),Ste-notrophomonas acidaminiphila (100%),Alcaligenes sp.(99%)and Aquamicrobium de?uvium (100%).DGGE-pro?les of culture C B-BT showed bands attributable to Rhodococcus ,Achromobacter ,Methylobacterium rhizobium ,Alcaligenes and Aquamicrobium .
0045-6535/$-see front matter ó2004Elsevier Ltd.All rights reserved.doi:10.1016/j.chemosphere.2004.06.013
*
Corresponding author.Tel.:+39250316724;fax:+39250316694.E-mail address:vincenza.andreoni@unimi.it (V.Andreoni).
Chemosphere 57(2004)
401–412
https://www.doczj.com/doc/075067500.html,/locate/chemosphere
The isolation of Rhodococcus aetherovorans and Methylobacterium sp.can be consistent with the hypothesis that dif-ferent phenanthrene-degrading strategies,cell surface properties,or the presence of xenobiotic-speci?c membrane car-riers could play a role in the uptake/degradation of solid phenanthrene.
ó2004Elsevier Ltd.All rights reserved.
Keywords:Soil chemical/enzymatic characteristics;DGGE;Bacterial diversity;Phenanthrene consumption;Batch liquid systems
1.Introduction
Polycyclic aromatic hydrocarbons(PAHs)are wide-spread in nature(i.e.soil,water and sediments)because of several polluting anthropogenic activities(Samanta et al.,2002).They have been recognised as a potential health risk due to their intrinsic chemical stability,high recalcitrance to di?erent types of degradation and high toxicity to living organisms(Alexander,1999).
PAHs present in soil may exhibit a toxic activity to-wards di?erent plants,microorganisms and inverte-brates.Microorganisms,being in intimate contact with the soil environment,are considered to be the best indi-cators of soil pollution.In general,they are very sensi-tive to low concentrations of contaminants and rapidly response to soil perturbation.An alteration of their activity and diversity may result,and in turn it will re-?ect in a reduced soil quality(Schloter et al.,2003).Soil enzyme activities are the driving force behind all the bio-chemical transformations occurring in soil.Their evalu-ation may provide useful information on soil microbial activity and be helpful to establish e?ects of soil speci?c environmental conditions(Dick et al.,1996).
Numerous research e?orts are being dedicated to the search of proper remediation technologies to remove as much as possible contaminants from the environment or to transform them into less toxic compounds.Bioreme-diation appears to be an appealing technology to ap-proach the recovery of PAH-polluted sites(Harayama, 1997).Several microorganisms are capable to mineralise a large variety of PAHs and/or to break down them to their less-toxic metabolites(Cerniglia,1992).The very low water-solubility of PAHs and the slow mass-transfer rates from solid phase may limit their availability to microorganisms,thus hindering natural attenuation microbial processes.However,some bacteria degrade sorbed PHAs at di?erent rates,indicating organism-spe-ci?c bioavailability(Grosser et al.,2000).
Bioremediation of PAH contaminated sites rely either on the presence of autochthonous degrading bac-teria which capabilities might be stimulated in situ (Margesin and Schinner,1997),or on the inoculation of selected microorganisms with desired catabolic traits in bioaugmentation techniques(Straube et al.,1999). When microorganisms are added to speed up degrada-tion in contaminated environments,the duration assess-ment and biological process e?ciency depend on the evolution of bacterial communities in terms of composi-tion and catabolic activity.Denaturing gradient gel electrophoresis(DGGE)analysis of16S rRNA genes represents a powerful tool to study the bacterial commu-nity structures in complex environments as well as in enrichment cultures(Muyzer and Smalla,1998).How-ever,the combination of both culture-independent and culture-dependent techniques might provide useful and complementary information on the structure of micro-bial communities.
Soils with di?erent pollution history were preliminary characterized in terms of their chemical properties,enzy-matic activity and culturable heterotrophic bacteria.Site characterization is a pre-requisite when dealing with any remediation approach of a polluted site(Smith and Mason,1999).Indeed,chemical and biochemical proper-ties may assist in the analysis of the ability for the soil to be recovered(Margesin et al.,2000).Moreover,the enrichment and selection of bacterial phenanthrene-degrading cultures,capable of degrading solid phenanth-rene in batch liquid systems were performed.The kinetics of phenanthrene disappearance by enriched cultures,the comparison of their degradation rates and their species composition were also investigated,as assessed by DGGE analysis of PCR-ampli?ed16S rDNA gene fragments. The enrichment of such cultures is a necessary step to obtain microorganisms with the desired catabolic traits, usable in the bioaugmentation of polluted soils.
2.Materials and methods
2.1.Chemicals
Phenanthrene was at>96%purity(Sigma Aldrich, Germany).Solvents at99.9%purity and all the other chemicals,reagent grade were supplied by Analar, BDH Ltd.,(Germany),unless otherwise stated.
2.2.Soil description and sampling
Three soils having a di?erent pollution history were https://www.doczj.com/doc/075067500.html,ly:
(1)A German soil,G,polluted by a long-term exposi-
tion(>50years)to alkanes and PAHs,leading to the formation of a typical light non-aqueous phase
402V.Andreoni et al./Chemosphere57(2004)401–412
liquid(LNAPL)contamination(Saccomandi and Gianfreda,2001).The soil is from Turingia(Ger-many)and its pollution is dated back to II World War.The site is still heavily contaminated because no remediation actions were implemented on it. (2)An Italian agricultural soil,I-BT,from the North
of Italy,with no or negligible presence of pol-lutants.
(3)A Belgian soil,B-BT,from a?uvial canal of Bruxe-
lles(Belgium),characterised by a medium-term(<3 years)exposition to PAHs.The soil was subjected to an accidental pollution event that caused a spread distribution of PAHs on its surface.The soil was sampled after3years from the pollution event.
Italian and Belgian soil samples were taken random by ram-drilling at a depth of5–15cm.German soil was drawn from within the LNAPL phase,immediately above the water table(at a depth ranging from5.5to7.6m below soil surface).Soil samples were packed on-site into sealed polythene bags,and transported to the labo-ratory,stored dark and cooled(4°C).Samples were homogenised,sieved to<0.2mm and stored at4°C until used.
Investigations were performed also on Italian(I-AT) and Belgian(B-AT)soils after bioremediation pilot experiments.Soils were treated aerobically in a bioreac-tor for5months;the experimental procedure adopted and the obtained results are under a patent.Unfortu-nately,no further information was provided by the site?s owner.German soil was not treated because previous laboratory investigations demonstrated that any e?ort to bioremediate it was unsuccessful(Saccomandi and Gianfreda,2001).
2.3.Determination of chemical and microbiological properties
The soils were characterized with respect to both phys-ical and chemical as well as microbiological properties.In particular,a set of enzyme activities(e.g.dehydrogenase,?uorescein diacetate hydrolase,arylsulphatase,phospha-tase and urease)and culturable heterotrophic bacterial cell number were determined.Molecular biodiversity of total bacterial populations was also analysed,according to methods described below(Section2.6).
Chemical and physical analyses were performed on air-dried and sieved(<2mm)samples according to standard techniques(Methods of Soil Chemical Analy-sis,1996).Soil organic C was determined by the method of dichromate oxidation,pH was measured by glass electrode in1:2.5H2O suspensions,total N was meas-ured by the standard Kjeldahl method.Particle size dis-tribution was assessed by the pipette-method.Overall content of PAHs and alkanes of German soil was deter-mined according to Saccomandi and Gianfreda(2001).Heavy metals were determined by atomic adsorption spectroscopy(AAS)after acid digestion with HF/HNO3.
Enzyme activities were determined on fresh moist soils sieved<2mm.The arylsulphatase(ARYL)and phosphatase(PHO)activities were determined according to Tabatabai and Bremner(1970)and Sannino and Gianfreda(2001),respectively.Speci?c substrates(p-nitrophenyl derivatives)and bu?ers were used for each enzyme.Urease(UR)activity was measured as described by Kandeler and Gerber(1988).Dehydrogenase(DH) assays were performed using soluble tetrazolium salt (TTC)as an arti?cial acceptor(Trevors,1984).The activ-ity of?uorescein diacetate hydrolase(FDAH)was as-sessed as described by Adam and Duncan(2001).A unit(U)of ARYL,DH and PHO enzyme activity was de?ned as the micromoles of substrate transformed at 30°Chà1by1g of dried soil.The FDAH and UR activ-ities were expressed as micrograms of substrate hydro-lysed at30°Chà1by1g of dried soil.Control tests with autoclaved soils were carried out to evaluate the spontaneous or abiotic transformation of substrates.
To enumerate culturable heterotrophic bacteria,10g of each soil sample were suspended in45ml sterilised Na4P2O7(0.2glà1in bidistilled water)in300ml glass bottles for1h on a shaker,in order to separate bacteria from soil particles.One millilitre of supernatant ob-tained after10min sedimentation was then10-fold serial diluted in NaCl9glà1solution.Appropriate dilutions were plated onto10%strength Tryptic Soy Agar med-ium for a total heterotrophic bacterial count;100l lmlà1 cycloheximide were added to the medium to inhibit the growth of eukaryotes.The plates were incubated at 28°C for8days and then counted.
Unless otherwise speci?ed,all results reported are averages of triplicate determinations.
2.4.Enrichment and isolation of phenanthrene-degrading cultures
Freshly prepared-phenanthrene stock solution in ace-tone(20mgmlà1)was added to500ml glass bottles.The acetone was allowed to evaporate before adding100ml of autoclaved M9mineral salt medium(Kunz and Chapman,1981)to have a?nal concentration of 200mg là1phenanthrene.Then10g of soil samples were added to a series of bottles.The bottles were te?on-stop-pered and incubated in the dark at25°C with agitation on a reciprocal shaker at96rpm for3weeks.Periodi-cally(3weeks)10ml aliquots of grown cultures were transferred into fresh medium under the same condi-tions.
Di?erent bacteria were isolated from the enrichment cultures.The isolates were grown on M9liquid medium containing100mglà1phenanthrene.Pure cultures were identi?ed by16S rDNA gene nucleotide sequence ana-lysis according to the method below described.
V.Andreoni et al./Chemosphere57(2004)401–412403
2.5.Measurements of phenanthrene utilisation rates
The mixed cultures were grown at25°C with shaking in500ml bottles containing100ml M9mineral medium supplemented with200mglà1phenanthrene.Four bot-tles for each culture were prepared.At each sampling time the concentration of phenanthrene was determined on duplicate sacri?cial bottles and the other two bottles were utilised to perform protein content analysis(Brad-ford,1976)and to extract total DNA(see below).Two bottle-controls(without bacteria)were run in parallel to account for the abiotic loss of phenanthrene.
The extraction and quanti?cation of phenanthrene was determined as follows.Culture broths were ex-tracted three times with50ml CH2Cl2;the organic layers were collected,dried with Na2SO4,?ltered and the sol-vent was removed under reduced pressure.The residue was solved in2ml of ethyl acetate and4ml of a solution of dodecanol in ethyl acetate(5mgmlà1)were added as internal standard for gas chromatographic analyses.The aqueous phase was acidi?ed by conc.HCl(pH2)and ex-tracted three times with50ml ethyl acetate;the organic layers were collected and processed as before described.
Gas-chromatographic analyses were carried out using a DANI1000Gas-chromatograph,equipped with a FID detector(hydrogen0.9bar,air1.0bar and nitro-gen1.0bar)and a fused silica capillary column WCOT-CP-SIL8CB Chrompack(25m·0.32mm ID),carrier helium(0.8bar),and injection temperature300°C, detection300°C,initial oven temperature140°C (3min),temperature increase10°C minà1,?nal iso-therm250°C,injection volume2l l.The dodecanol R t was6.9min and the phenanthrene R t was11.3min. Detector signal output was monitored by computer and all chromatograms and data were generated and processed by Dani Data Station version1.7software.
2.6.Molecular methods
DNA was extracted from soil samples,enrichment cultures and isolated strains.Soil DNA and enrichment culture DNA were extracted by a bead-beating method (MOBIO,USA)and by BIO101method(Resnova, Italy),respectively,according to the manufacturer instructions.According to Cavalca et al.(2002),protein-ase K(1mgmlà1)was used to extract DNA from strains.
PCR ampli?cation of the16S rDNA was performed on the extracted DNA,by using eubacterial universal primers P27f and P1495r referred to E.coli nucleotide se-quence of16S rDNA gene(Cavalca et al.,2002).Nested PCR reaction for V3ampli?cation was carried out according to Muyzer and Smalla(1998).V3PCR prod-ucts from soil,enrichment culture and bacterial isolates DNAs were characterized by a DGGE run on a vertical acrylamide gel in a DCODE Universal Mutation Detec-tion System(Biorad).DGGE was performed with8% (wt/vol)polyacrylamide gels in TAE bu?er(20mM Tris acetate pH7.5,10mM sodium acetate,0.5mM Na2-EDTA)with a linear chemical gradient ranging from 35%to65%.Denaturant solutions were prepared by mix-ing the appropriate volumes of two0–100%denaturant stock solutions(7M urea,and40%vol/vol formamide (Amersham Biosciences,Swedan).Gels were run at a constant voltage of70V for16h at55°C.Gels were stained in a0.5mg là1ethidium bromide solution and documented with GelDoc System(Biorad).Bands of interest were excised from DGGE using an UV transillu-minator.The excised bands were suspended into200l of PCR water,reampli?ed and sequenced.The nucleotide sequences of16S rDNA of the resulting amplicons and of isolates were determined according to the Perkin El-mer ABI Prism protocol(Applied Biosystems,USA). Primers used in the PCR reaction for sequencing prod-ucts were the same of those in normal16S rDNA PCR reactions.The forward and reverse samples were run on an Applied Biosystems310A sequence analyser.The sequences were compared with similar sequences of refer-ence organisms deposited in public domain databases.
DGGE analyses were performed to compare the bac-terial community structures of soils and enrichment cul-tures.Although the technique could be associated with a variety of PCR biases(Wintzingerode et al.,1997;Fro-min et al.,2002),it provides comprehensive information on the global patterns of microbial diversity(Torsvik and Overas,2002).However,to minimize biases,DGGE analyses were performed on samples treated using iden-tical methods in which DNA extraction and ampli?ca-tion biases are supposed to occur homogeneously.
Shannon index(H0)(Magurran,1988)was used to evaluate the biodiversity of both soils and enrichment cultures,and Sorensen index(S)(Magurran,1988)to evaluate the similarity within soils(native vs.treated soil)and within the deriving cultures.
The Shannon index of soils was calculated on the ba-sis of the number and intensity of bands present on DGGE samples,run on the same gel,as follows: H0?à
P
P i log P i,where P i is the importance probabil-ity of the bands in a gel lane.P i was calculated as fol-lows:P i=n i/N,where n i is the band intensity for each individual band and N is the sum of intensities of bands in a lane.Statistical comparison of di?erent DGGE pro-?les was done with the GelDoc software package.This latter assumes that the population size is proportional to the thickness of bands.Gel analysis included conver-sion of the scanned gel image and normalization in order to correct shift within or between gels,so that bands or peaks of the same molecular size have the same physical position relative to a standard.Once all banding pro?les were in a standardized analysis format,each band could be described by its position on the gel and by its relative intensity.
404V.Andreoni et al./Chemosphere57(2004)401–412
3.Results and discussion
3.1.Physico-chemical and microbiological properties of
soils
The chemical and physical properties of a soil as well as the evaluation of its pollution degree may help to esti-mate the impact of pollutants on the quality of soil un-der investigation,if they are complemented with the measurement of biological properties (Margesin et al.,2000).
Tables 1and 2summarise the physical and chemical properties of investigated soils and the amounts of both organic and inorganic pollutants.
The moderate-high amounts of carbonate and the pH values (measured in H 2O),ranging from 2.68to 5.36and from 6.73to 8.19,respectively,indicate a sub-to moder-ate-alkaline character of soils (Table 1).At the measured pH range soil microbial growth and its activity are usu-ally favoured.As discussed by Smith and Doran (1996),soil pH can provide valuable information on the availa-bility and toxicity of several elements,including Fe,Al,Mn,Cu,Cd and others to plants and microorganisms.German and Italian soils showed comparable amounts of clay,silt and sand fractions (Table 1)whereas Belgian soil had a very low amount of both clay ($7%)and silt ($8%)and a predominant presence of sand (>80%as total of coarse and ?ne fractions).According to USDA classi?cation (Soil Survey Sta?,1993),German and Italian soils can be classi?ed sandy clay loam soils while Belgian is a typically loamy sand soil.
In Belgian and mainly in German soil before treat-ment (B-BT and G)total organic C values,and conse-quently organic matter contents,were very high,being in?uenced by organic pollutant contamination.Thus,their values did not represent natural,endogenous soil organic matter levels,possibly present in the soil in the absence of any contamination.Considering the low amounts of N measured in both soils,the C/N ratios (11.5and 26.3for B-BT and Gsoils,respectively)were higher than those normally found in unpolluted soils.When hydrocarbon-polluted soils are considered,much higher C/N ratios,ranging from a minimum value of 9:1to a maximum of 200:1,are,however,needed to ob-tain a consistent microbial growth and resulting hydro-carbon degradation (Bewley,1996).
The physical and chemical properties of Belgian and Italian soils were also measured after the bioremediation treatment (Table 1).As expected,no signi?cant varia-tions of clay,silt and sand fractions were noted.The 2-fold higher amounts of both N and available K meas-ured in B-AT are likely the result of nutrient supply dur-ing the biological treatment.
According to the current European Union regulation (Commission of the European Communities,1986)
T a b l e 1P h y s i c a l –c h e m i c a l p r o p e r t i e s o f s t u d y s o i l s
S o i l
p H (H 2O )
C a C O 3
(%)
M o i s t u r e (%)
C l a y (%)
S i l t (%)
C o a r s e s a n d (%)F i n e s a n d (%)
O .C .(g k g à1)O .M .(g k g à1)
T o t a l N (g k g à1)
C /N
P -O l s e n (m g k g à1)
A v a i l a b l e K (m g k g à1)
B e f o r e t r e a t m e n t G e r m a n 6.73a *
3.07a
13.0a 24.7a 15.0a 22.0a 38.3a 11.1a 19.1a 0.422a 26.3a T r a c e
n d
(G )(±0.23)a
(±0.15)(±0.90)(±1.5)(±0.97)(±0.99)(±1.0)(±1.50)(±2.20)(±0.09)(±3.45)I t a l i a n 7.67b 3.93a 14.5a 22.5a 24.5b 19.5b 33.4b 7.70b 13.3b 2.20b 3.5b 33.7a 337b (I -B T )(±0.56)(±0.21)(±1.10)(±2.0)(±2.4)(±1.30)(±2.10)(±1.20)(±1.90)(±0.50)(±0.45)(±4.50)(±17.8)B e l g i a n 8.19b 2.93a 11.0a 6.94b 7.75c 40.0c 45.3c 8.2b 14.1b 0.71c 11.5c 15.0b 224c (B -B T )(±1.10)
(±0.09)
(±0.85)(±0.54)
(±0.65)
(±3.20)(±3.60)
(±1.60)
(±2.20)
(±0.09)
(±1.20)
(±1.50)
(±13.4)
A f t e r t r e a t m e n t I t a l i a n 7.73b 5.36b 18.5b 21.6a 23.9b 18.9a 35.6b 7.9b 13.6b 2.85d 2.8b 30.5a 574d (I -A T )(±0.61)(±0.35)(±1.20)(±2.0)(±2.70)(±1.40)(±3.10)(±1.50)(±2.10)(+0.60)(±0.38)(±3.90)(±20.7)
B e l g i a n 8.17b 2.68a 11.0a 6.90b 8.58c 38.6c 45.9c 8.20b 14.1b 1.40e 5.8d 20.2c 495e (B -A T )(±0.60)
(±0.10)(±0.91)(±0.54)(±0.74)(±1.93)(±2.30)
(±0.90)(±1.50)
(±0.80)(±0.40)
(±0.80)
(±18.5)*
F o r e a c h v a r i a b l e d i ?e r e n t l e t t e r s a l o n g s i d e c o l u m n s r e f e r t o s i g n i ?c a n t d i ?e r e n c e s (P (0.05).a V a l u e s i n p a r e n t h e s e s r e p r e s e n t s t a n d a r d d e v i a t i o n .
V.Andreoni et al./Chemosphere 57(2004)401–412405
referring to agricultural soils,investigated soils showed levels of heavy metals all below the maximum permitted concentrations,except for copper in Italian soil that was about twice the safe limit(150mgkgà1soil).
A di?erent situation holds when organic pollutants are considered.German soil resulted heavily polluted by high concentrations of alkanes and PAHs.BTEX and phenols were also detected(data not shown),thus con?rming the presence of a LNAPL widespread pollu-tion(Saccomandi and Gianfreda,2001).In contrast, these pollutants were not detected in Italian soils. Belgian soil presented a detectable amount of PAHs (30.8mgkgà1),being phenanthrene relatively the most abundant(Table2).
The activities of?ve enzymes and the heterotrophic bacteria of the investigated soils are reported in Table3. Arylsulphatase and phosphatase release sulfate and phosphate,the main plant and microbial available S and P forms,from various organic sulfate and phos-phate esters(Nannipieri et al.,2002).Urease catalyses the hydrolysis of urea to carbon dioxide and ammo-nium,and it is widely distributed in microorganisms, plants and animals(Nannipieri et al.,2002).Dehydro-genase activity typically occurs in all intact,viable
Table2
Amounts of inorganic and organic pollutants of study soils
Inorganic(mgkgà1)Organic(mgkgà1)
Soil Cu Zn Cr Ni Fe Alkanes PAH Phenanthrene Before treatment
German145a*88.0a14.0a39.0a 6.1a29094a14a
(G)(±8.7)a(±9.4)(±2.7)(±8.2)(±3.6)(±10.1)(±6.4)(±2.6)
Italian301b121b72.4b75.5b40.3b nd nd nd
(I-BT)(±21.5)(±9.6)(±6.5)(±8.5)(±5.4)
Belgian50.2c124b83.9c55.4c39.0b nd30.8b 4.7b
(B-BT)(±5.3)(±7.5)(±5.3)(±6.5)(±5.6)(±3.2)(±0.7)
After treatment
Italian290b265d70.8b85.7b25.9c nd nd nd
(I-AT)(±19.3)(±12.1)(±5.8)(±9.1)(±3.2)
Belgian52.9c329e67.4b65.6d33.4d nd8.9c0.7c
(B-AT)(±8.5)(±17.5)(±6.4)(±7.6)(±2.4)(±0.87)(±0.6)
*For each variable di?erent letters alongside columns refer to signi?cant di?erences(P(0.05).
a Values in parentheses represent standard deviation.
Table3
Enzyme activities and microbial counts of study soils
Soil ARYL
(l molgà1hà1)PHO
(l molgà1hà1)
UR
(l ggà1hà1)
DH
(l ggà1hà1)
FDAH
(l ggà1hà1)
Total
heterotrophs
CFU(gà1)
Before treatment
German nd 4.10a nd18.9a nd 3.9·105a (G)(±0.045)(±2.1)(±4.0·104) Italian0.388a* 2.20b18.4a0.748b186a 4.9·107b
(I-BT)(±0.07)a(±0.31)(±1.7)(±0.03)(±6.45)(±4.0·106) Belgian0.014b0.35c nd nd8.52b 2.3·107c
(B-BT)(±0.003)(±0.21)(±0.91)(±2.0·106) After treatment
Italian0.555c 3.84d18.8a 1.27c197c 3.9·108d
(I-AT)(±0.09)(±0.40)(±1.6)(±0.08)(±6.51)(±5.0·107) Belgian0.265d 2.90b nd0.049d162d 5.8·108e
(B-AT)(±0.02)(±0.1)(±0.01)(±5.56)(±6.0·107) nd=not detected.ARYL=arylsulphatase,PHO=phosphatase,UR=urease,DH=dehydrogenase,FDAH=?uorescein diacetate hydrolase.
*For each variable di?erent letters alongside columns refer to signi?cant di?erences(P(0.05).
a Values in parentheses represent standard deviation.
406V.Andreoni et al./Chemosphere57(2004)401–412
microbial cells.Thus,its measurement is usually related to the presence of viable microorganisms and their oxi-dative capability(Trevors,1984).Fluorescein diacetate hydrolase(FDAH)has been often used as a sensor and functional indicator of soil health(Adam and Dun-can,2001).Being the?uorogenic substrate uptaken by active cells and then transformed by a large arrays of hydrolytic enzymes,the enzyme has been considered a measure of the soil microorganism activity(Killham and Staddon,2002).
Enzyme activities and total heterotrophs,mainly for Belgian and German soils,are in agreement with the re-sults obtained with soils contaminated by similar pollut-ants(Kiss et al.,1998;Margesin et al.,2000).The German soil was the most contaminated compared to Belgian and Italian soils,having the lowest number of heterotrophs(Table3).
After the biological treatment an increase in CFU of only one order of magnitude was measured in both Bel-gian and Italian soils(Table3).As reported by Margesin et al.(2000)total number of heterotrophs of PAHs pol-luted soils did not greatly increase after biological reme-diation actions,whereas the relative amounts of speci?c pollutant-degrading bacteria increased to a detectable extent.
Enzyme activities also con?rmed that the Italian soil showed the highest microbiological activity.All the measured enzymes were present at moderate to high range levels,usually found in agricultural soils(Nannipi-eri et al.,2002).The relatively low dehydrogenase activ-ity measured in this soil(which seems to contradict the high values of both FDAH activity and total microor-ganisms)could be explained by the possible interference exerted by the high Cu content(Table2)on the analytic assay used.Indeed,Cu may reacts abiotically with the triphenylformazan,the end product of DH catalysis, thus resulting in a underestimation of the soil dehydro-genase activity(Chander and Brookes,1991).
Although the in?uence of other factors deriving from natural and anthropogenic events cannot be ruled out (Gianfreda and Bollag,1996),the complete absence and/or the very low enzymatic activities of both German and Belgian soils could be also partly due to the presence of PAHs in soils.As extensively reviewed by Kiss et al. (1998),even moderate levels of hydrocarbon contamina-tion may cause a signi?cant decline of several soil en-zyme activities,showing each enzyme a di?erent sensitivity to the presence of pollutants.Although the interpretation of enzyme activities of soil is complex be-cause both extracellular and intracellular enzyme activi-ties contribute to the overall soil enzyme activity,some hypotheses might be advanced.In soil,non-polar organ-ic compounds,such as hydrocarbons,may likely exert di?erent e?ects on microbiological properties.Hydro-carbons may be toxic to soil microorganisms which may re?ect in a consistent reduced enzymatic activity;and/or they my cover both organic-mineral and cell sur-faces,thus hindering the interaction between enzyme ac-tive sites and soluble substrates with adverse e?ect on enzyme activity expression(Kiss et al.,1998).Moreover, a synergistic negative e?ect on soil enzyme activities ex-erted by the simultaneous presence of heavy metals can-not be ruled out.
After bioremediation,enzyme activities of Italian and Belgian soils increased to a moderate and a more detect-able extent,respectively.
3.2.Biodiversity of soils
In our analysis,the number of DGGE bands was taken as an indication of species in each sample.The rel-ative surface intensity of each DGGE band and the sum of all the surfaces for all bands in a sample were used to estimate species abundance(Fromin et al.,2002;Sekig-uchi et al.,2002).DGGE pro?les of soils are shown in Fig.1.Many DGGE bands were observed in the pro-?les,thus indicating the presence of di?erent bacterial populations and di?erent relative abundance species in soils.As indicated by the values of Shannon indices, contamination of soils appeared to a?ect their genetic diversity:German soil and native Belgian soil B-BT showed the pooresteH0
G
?0:85Tand the highest
eH0
BàBT
?2:87Tbiodiversity,respectively.For the Italian agricultural soil I-BT,containing negligible amounts of organic pollutants but the highest Cu content,a Shan-non index=2.13was found.
After treatment,a loss of bacterial species diversity
occurred in Belgian soil with a H0
BàBT
equal to1.13.Fur-thermore,the bacterial community of the native soil B-BT showed a marked di?erent pattern when compared with its treated B-AT counterpart.Indeed,the S index of similarity was equal to0.18.Only few bands(‘‘a’’and‘‘b’’in Fig.1)were in common between the two soils,indicating the survival of some predominant species.
On the contrary,for Italian soils only negligible dif-ferences in DNA patterns(S=0.56)were evidenced be-tween the native I-BT and its treated I-AT counterpart
eH0
IàAT
?2:14T,indicating that the bioremediation did not substantially change the community structure of the native one.
3.3.Enrichment of phenanthrene-degrading mixed cul-tures and determination of degradation kinetics
The diversity encountered in the bacterial communi-ties of the study soils prompted us to perform enrich-ments on phenanthrene from all soil samples in order to obtain cultures with di?erent potential strategies to degrade phenanthrene.
Attempts to enrich phenanthrene degrading bacteria from the German soil were unsuccessful(Saccomandi
V.Andreoni et al./Chemosphere57(2004)401–412407
and Gianfreda,2001).The presence of highly bound res-idues in the old-contaminated German soil could have represented a constraint in phenanthrene bioavailability to bacteria thus impairing the possibility to isolate degrading microorganisms.
Four mixed bacterial cultures,named C B-BT and C B-AT,and C I-BT and C I-AT were instead selected from the Belgian and Italian soils before and after the biolog-ical treatment,respectively.
All cultures enriched from Belgian and Italian soils grew on phenanthrene when added as sole C and energy source and turbidity of culture broths increased during incubation.
Fig.2shows the disappearance of200mglà1crystal-line phenanthrene and the corresponding protein con-tents within21-d incubation of the selected cultures.A time course analysis of phenanthrene may provide an estimate of?rst order uptake/degradation rate constant according to the following expression:X t=X0eàkt, where X t is the concentration of phenanthrene in mglà1, k is the uptake/degradation constant and t is the time.
When phenanthrene degradation data of Fig.2were reported in a semilog plot,a one-slope behaviour was observed for C B-BT and C I-AT cultures,while a typical two-slope occurred for C B-AT and C I-BT,suggesting a more complex kinetics of phenanthrene degradation by these cultures(data not shown).This could imply that for culture C B-BT and C I-AT the whole phenanthrene degradation process is dominated by a single,straight-forward key step,whereas for cultures C B-AT and C I-BT a complex mechanism,involving a slower interme-diate step,occurred.
Table4reports the degradation constants calculated by means of a non-linear regression routine applied to phenanthrene degradation data of Fig.2.The?rst step-kinetics occurring for C B-AT and C I-AT,character-ized by low degradation constants,could suggest a slower utilisation of phenanthrene within the?rst8 days.In particular,the very low k1value(0.020dà1)cal-
Fig.1.DGGE analysis of PCR-ampli?ed16S rDNA gene V3fragments from soil samples and from enrichment cultures after six transplants on fresh phenanthrene.Bands were designated as described in the text.G,German soil;B-BT,Belgian soil before treatment;B-AT,Belgian soil after treatment;I-BT,Italian soil before treatment;I-AT,Italian soil after treatment;C B-BT,C B-AT, C I-BT,C I-AT,enrichment cultures from the corresponding soil samples.
408V.Andreoni et al./Chemosphere57(2004)401–412
culated for C B-AT could indicate the presence of a slow phenanthrene mass transfer resulting in a hampered PAH utilisation.By contrast,the mixed culture C B-BT al-most completely utilized200mglà1phenanthrene(more than90%)within10days.Longer times were required for complete degradation by C I-AT and C B-AT(Fig.2 and Table4).All cultures degraded phenanthrene without the appearance of any metabolites in culture broths.
The protein content patterns of culture broths con-?rmed the ability of strains to utilise phenanthrene as the sole C source.The pro?les of protein contents vs. phenanthrene disappearance of cultures C I-AT and C I-BT were the same as C B-BT(data not shown).
C B-AT protein content seems to con?rm that the con-sumption rate by this culture was limited by dissolution dynamics.Indeed,the growth rate of C B-AT,evaluated as protein content(2.33l g mlà1dà1)in the exponential (0–21d)growth phase was lower than that measured for C B-BT(7.78l gmlà1dà1)in the exponential(0–5d) growth phase.The di?erent behaviour of C B-BT com-pared to C B-AT,enriched from the same soil after the biotreatment,could be referred to a di?erent species composition of the cultures(Fig.1).The former con-tained probably bacteria with di?erent PAH-degrading strategies or with di?erent cell surface properties.A bac-terial adhesion to solid phenanthrene and subsequent solubilisation at the level of the cell wall could be hypothesised.Similar mechanisms have been suggested for degradation of solid hydrophobic chemicals(pal-mitic acid)by Pseudomonas strains(Thomas and Alex-ander,1987).
Culture C I-BT,obtained from the Italian agricultural soil I-BT,was for the?rst8days metabolically less ac-tive than culture C B-BT obtained from the Belgian con-taminated soil B-BT.A faster degradation occurred, however,in the last incubation period(k2value for C I-BT higher than k1value for C B-BT,Table4).A dif-ferent phenanthrene-degrading culture was selected from the Italian biotreated soil I-AT and its degrada-tion rate was slower than that of I-BT(Fig.2and Table4).3.4.Biodiversity of enrichment cultures
The DGGE pro?les of the mixed cultures analysed after six21-d-incubation transplants on phenanthrene, when cultures were supposed to be stable and used also for degradation kinetic experiments,are shown in Fig.1. DGGE pro?les of enrichment cultures were less complex than soil pro?les,due to the selective pressure repre-sented by the presence of fresh phenanthrene.
All the cultures showed DGGE pro?les that indi-cated a di?erent bacterial species composition,as evi-denced by the presence of peculiar bands in each culture(Fig.1).Sorensen similarity values calculated from DGGE pro?les revealed that there were signi?cant di?erences in species composition of cultures from each native and treated soil(S=0.33for C B-BT vs.C B-AT and 0.25for C I-BT vs.C I-AT).Some bands were in common among enrichment cultures,indicating the presence of similar bacterial species,such as band‘‘g’’in C B-BT, C B-AT and C I-BT.Other bands were visible in the enrich-ment culture DNA pro?les and in the corresponding soil samples(band‘‘a’’in C B-BT and C B-AT,in B-BT and B-AT,and band‘‘c’’in C I-BT and C I-AT,in I-BT and I-AT).All these bands belong to species that could be relevant in situ phenanthrene degraders and that have been enriched during the transplant procedure.Four bands(‘‘d’’,‘‘e’’,‘‘f’’and‘‘g’’)were in common among DNA pro?les of C B-BT and C B-AT,thus con?rming their presence in the native and treated Belgian soils(Fig.1).
The di?erences encountered in the DGGE pro?les could re?ect the di?erent degradative kinetics of the four cultures.The presence of di?erent species could assure a probable existence of di?erent mechanisms for e?cient assimilation/uptake of soluble or solid phenanthrene.
Colonies with di?erent morphologies were isolated from the fastest degrading culture C B-BT after growth on0.1·tryptic soy broth agar plates.Representative strains of C B-BT,identi?ed on the basis of1200nucleo-tides sequence homologies with entries in GenBank-EMBL databases,belong to:Achromobacter xylosoxidans (100%),Methylobacterium sp.(99%),Alcaligenes sp. (99%),Rhizobium galegae(99%),R.aetherovorans (100%),Aquamicrobium de?uvium(100%)and Stenotro-phomonas acidaminiphila(100%).When these strains were checked for the capability of growing on100mglà1 crystalline phenanthrene as sole C source,the growing strains had di?erent growth behaviour.While R.aether-ovorans produced a di?use turbidity of culture broths (data not shown),Methylobacterium sp.grew in contact with the phenanthrene crystals,as revealed by micro-scopic examination.This implies that the low solubility of phenanthrene was limiting the growth,and the few cells freely present in the culture broth were probably those sloughed o?from the crystal surfaces.
The presence of strains within the culture C B-BT dur-ing the time course of phenanthrene degradation was
Table4
Values of phenanthrene(200mglà1)disappearance constants
calculated for the cultures enriched from the study soils
Culture k1(dà1)k2(dà1)R2
C B-BT0.369–0.95
C B-AT0.0200.2970.99
C I-BT0.1130.5100.99
C I-AT0.076–0.98
k1and k2calculated by a non-linear regression routine
according the equation X t=X0exp(àkt)where X t is the con-
centration of phenanthrene in mglà1,k is the uptake or trans-
formation constant and t is time.
V.Andreoni et al./Chemosphere57(2004)401–412409
followed by DGGE analysis.During the degradation process,no change was evidenced in the bacterial com-ponents of C B-BT (Fig.3)but some bands increased their relative intensity.
C B-BT bands were correlated to the isolated strain bands (Fig.3)on the basis of their electrophoretic mobility.Theoretically,bands at the same position in the electrophoresis pattern contain DNA fragments with identical sequences.Band ‘‘h’’had the same electropho-retic mobility of R.aetherovorans ,band ‘‘l’’the same of R.galegae and Aquamicrobium de?uvium ,band ‘‘m’’the same of Methylobacterium sp.,band ‘‘n’’the same of Alcaligenes sp.and of one of the two bands of A.xylos-oxidans and band ‘‘p’’the same of the other band of A.xylosoxidans .Bands corresponding to R.aetherovorans and A.xylosoxidans increased their relative intensity during phenanthrene degradation suggesting that these strains represent active members of the culture and are likely involved directly or indirectly in the utilization of phenanthrene as C and energy sources.
The overlapping of ampli?ed PCR products cannot con?rm that sequences of these isolates are identical to the sequences of corresponding DGGE enrichment cul-ture bands.
Band corresponding to St.acidaminiphila has never been retrieved in culture C B-BT DGGE pro?les.This could be due either to its low cell number in the culture or to the DNA applied extraction method.Conversely,species corresponding to bands ‘‘a’’and ‘‘g’’in the DGGE pro?les of culture C B-BT were not recovered among isolates,and their sequence types were identi?ed as Pseudomonas and Arthrobacter ,respectively.The ampli?cation of these bands may be due to biases in selective PCR ampli?cation (Heuer and Smalla,1997).The bands corresponding to P.putida and Ralstonia sp.have approximately the same relative intensity dur-ing incubation time,suggesting that these species do not increase during phenanthrene degradation.
4.Conclusions
The results,here presented,all indicate that soils highly contaminated by hydrocarbons displayed di?er-ent microbiological properties.In particular the higher/the lower the pollutant content,the smaller/the greater are the activities of some enzymes related to nutrient cycling and the viable bacterial cell numbers.The di?er-ent microbiological properties of the soils probably re?ect the di?erent bacterial diversity as assessed by DGGE pro?les of the 16S rDNA genes.
Phenanthrene-degrading mixed cultures were en-riched from all soils except the old heavily contaminated German soil.When tested in liquid batch systems using solid phenanthrene as C and energy source,cultures showed di?erent kinetic behaviours probably because of a di?erent species composition,as evidenced by DGGE 16S rDNA pro?les.The presence of di?erent species could indicate a probable existence of di?erent mechanisms for e?cient assimilation/uptake of soluble or solid phenanthrene,as observed for C B-BT culture that contained more than one phenanthrene-degrading bacterium.The simultaneous presence in the culture of Rhodococcus and Methylobacterium strains might be ex-plained with the capability to use phenanthrene under di?erent conditions such as dissolved,solid associated,and perhaps surfactant-associated,according to di?erent substrate-degrading strategies.C B-BT culture also con-tained bacteria that do not use phenanthrene,suggesting that the phenanthrene-degraders themselves may be associated with bacteria using metabolites of phenanth-rene.The presence of some DGGE bands with the same electrophoretic mobility and the presence of degrading strains belonging to the same species in all the enrich-ments are indicative of their degradative role in the cul-tures.The isolation of bacteria from B-BT soil,that are able to grow on phenanthrene,is consistent with the ob-served decrease of PAH and phenanthrene contents of soil after the biotreatment and suggests that aerobic phenanthrene biodegradation was occurred.The
?nding
Fig. 3.DGGE analysis of V3fragments obtained from uncharacterized bacterial culture C B-BT and bacterial isolates from the https://www.doczj.com/doc/075067500.html,nes T 0(P )to T 8(P )show the pro?les obtained from C B-BT after 0,2,4and 8day growth in presence of phenanthrene;lane V 3MIX contains the separation pattern of a mixture of fragments of seven isolates,i.e.,Alcaligens sp.(lane 1);Rhizobium galegae (lane 2);Methylobacterium sp.(lane 3);Stenotrophomonas acidaminiphila (lane 4);Aquamicrobium de?uvium (lane 5);Achromobacter xylosoxidans (lane 6)and R.aetherovorans (lane 7).
410V.Andreoni et al./Chemosphere 57(2004)401–412
that a number of bacteria identi?ed in culture C B-BT de-grade phenanthrene supports this assumption.The iso-lation of R.aetherovorans and Methylobacterium sp. can be consistent with the hypothesis that di?erent phen-anthrene-degraders inhabiting soils and enrichment cul-tures may be adapted to di?erent phenanthrene bioavailabilities.The use of these species in microcosm bioaugmentation trials could help in evaluating their in situ catabolic behaviour to degrade phenanthrene in highly polluted soils.
Acknowledgments
This research was supported by Ministero dell?Uni-versita′e della Ricerca,Italy,Programmi di Interesse Nazionale PRIN2002-2003.Dr.Fornaro E.of ENVIR-OREM,Lugane,Switzerland is thanked for the kind supply of Belgian and Italian soil samples and for the determination of their phenanthrene content.DiSSPA Contribution no.049.
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昭通市中医医院关于发挥中医药特色优势和提高中医临床疗效的绩效考核实施方案（试行） 为进一步加强我院中医药内涵建设，坚持以中医药为主的办院方向，保持和发挥中医药特色优势，提高临床疗效和学术水平，把我院建设成功能完善、中医中药特色突出的三级中医医院，我院依照医院中长期发展规划、中医院评价标准及评估细则和实施方案，特制定发挥中医药特色优势和提高中医临床疗效的鼓励和考核制度： 一、加强中医中药人才培养，有计划配备的中医中药人才 根据国家中医药管理局关于中医医院发挥中医药特色优势，加强人员配备的要求，结合我院实际情况，加强专业队伍建设。 （一）优化卫生技术人员结构，按计划配备充足的中医药人员。今后我院引进主要以中医、中西医结合为主的卫生技术人员，现有进修学习人员主要安排到上级中医医院进修学习，争取在十
三五计划内使中医类别执业医师（含执业助理医师）占执业医师比例大于60%；中药专业技术人员占药学专业技术人员的比例大于60%；护理人员系统接受中医药知识和技能岗位培训的比例不低于70%。每个临床科室执业医师中至少有60%中医类别执业医师（麻醉科、口腔科除外）。 （二）定期组织医、护、药人员进行中医基础理论和基本技能培训，并熟练掌握各种技能。每年要求我院全体护理、药学人员系统主动学习中医中药理论基础知识，提高中医护理工作技能。 （三）结合我院实际，要求现有领导班子成员，认真学习中医知识，参加中医管理培训教材。医院主要负责人、业务管理领导和医务、护理、药剂、教学、科研部门的主要负责人经过中医药政策、中医药知识和管理知识的系统培训。对参加中医中药学历提高学习的人员，给予一定的补助和奖励。 （四）临床科室负责人中有计划调整充实中医类别执业医师
治妇科病最常用的7种养生中药 治妇科病最常用的7种养生中药。女人天生爱美，然而受妇科病的困扰，严重了影响了美丽与健康。这是因为妇科病不仅损害身体健康，还会影响容颜，使脸上长痘痘、肌肤暗淡无光等。人们往往采用中药治疗方法。下面介绍治妇科病最常用的7种养生中药。 1、当归：李时珍认为，当归是女人调血的要药。从我国南北朝开始，当归就被视为补血活血珍品。补血调经、活血止痛、泽颜润肤、生肌强体、延年益寿。在保护女性健康方面极其重要。 2、枸杞子：是我国最早记载的一味妇科用药，可滋阴养血，益肝补肾，能明目润肤，乌发养颜。《重庆堂随笔》评价它：“专补心血，非他药所能及。”《中药大辞典》认为它滋肾、润肺，补肝，明目。 3、黄芪：能够增强皮肤营养和皮肤的抗菌能力，防止皮肤老化，促进毛发生长，防止脱发。它含有多种氨基酸及人体必需微量元素和叶酸等，具有全面营养作用。中医认为，它能够补中益气，对气虚血脱、崩带及一切气衰血虚之症有疗效。 4、白芍：《日华子本草》评价它：“治风补涝，主女人一切病，并产后诸疾。”《唐本草》说它“益女子血。”现代中医认为，它能够养血柔肝、缓中止痛，对月经不调，崩漏，带下有效果。 5、珍珠：珍珠历来是名贵的中药材，对皮肤有特殊的滋养保健作用，能保持颜面细腻白嫩，并可促进人体细胞再生，防止衰老，延缓皱纹产生。中医认为，它具有养阴安神、镇心熄风清热、解毒生肌等功效，可治眩晕、耳鸣、烦躁、头痛、失眠、惊悸等病症。 6、芦荟：对女性来说，芦荟是最熟悉不过的美容佳品。中医看来，芦荟能治疗清热、通便、热结便秘、妇女闭经等症。 7、川芎：川芎在唐朝《日华子本草》中有着很高的评价：“治一切风，一切气，一切劳损，一切血，补五劳，行气开郁，活血止痛，对经闭、难产、产后瘀阻块痛等有效。壮筋骨，调众脉，养新血……”《医学启源》：“补血，治血虚头痛。” 以上7种中药都是最常见的药材，在治疗妇科病方面有很好的治病养生疗效。有些中药可以用来泡水喝。女性朋友选择中药时，一定根据自身体质及症状等因素而定。
如何正确煲中药 煲中药就是简单的“3碗水煮成1碗”？非也！最近与朋友讨论起如何煲中药，才发现许多年轻的广州人，虽然一有什么“头晕身热”都会习惯地煲中药喝，但到底怎样煲中药才是正确，往往只是一知半解，道听途说。其实，煲中药，还真是一件有技术含量的事！ 煎煮前——— ●清洗 中药材是否需清洗，这是很多人的疑问虽然很多中药饮片看起来表面会有些灰黑，其实在出售前都经过了加工炮制，所以煎煮之前一般无需清洗。如果实在觉得草药有些泥沙，可以用水迅速漂洗一下，但切忌浸洗，以免一些水溶性成分丢失，以及一些细小种子类的药材（如车前子等）被冲走流失。 ●浸泡 清洗步骤不能浸泡中草药，但煎煮之前，却需要有个浸泡药材的过程。 煎煮前用凉水浸泡药材约半小时，可以使水溶性成分析出在汤水中，同时也能增加汤药的浓度。冬天可以用20-30度的温水浸泡，以缩短煎煮时间，但切不可用开水浸泡，以免某些植物细胞中的蛋白质受热凝固，或是部分高分子物质形成胶体，不利于有效成分析出。 浸泡时间不宜超过1小时，特别是在夏天，浸泡时间过长会很容易引起酸败。 煎煮中——— ●用水 传统的“3碗水煮成1碗”，其实不是个科学的标准。因为不同处方的药味多少、药量大小各有不同，不同药材吸水量也有不同。如果真的有人一边煲药，一边不断把药汁倒来倒去，作为煲中药的标准，这样瞎折腾其实也不可能煲出最佳效果的中药。 应以水浸过药材面2-3cm为佳，或者用手轻轻摁住药材，水面刚好漫过手背。而不是机械地用3碗水煮药。通常一些花草类的药物吸水量较大，在浸泡半小时后水位下降，可以另加凉水至标准水位，再开始煎煮。 ●火候 一般的中药应先用武火，煮沸后改为文火。控制火候的意义在于，若火候过强，水分蒸发过快，影响有效成分的析出，亦易焦糊。 但一些治疗外感的中药，可以在煮沸之后不改文火，继续用武火煎煮15分钟左右即可。 ●时间 中药煎煮时间，应根据不同药物和疾病性质、有效成分溶出的难易和用药情况而定。楼步青说，沸腾后再用文火煲药的时间，一般中药，头煎应在20-25分钟，二煎15-20分钟；解表类中药，头煎10-15分钟，二煎10分钟；滋补类中药，头煎应在30-40分钟，二煎25-30分钟。 如果有“大头虾”不慎煎煮时间过长，令药汤太浓，这时可以加些白开水再煮沸，就可以避免有效成分反渗透的问题。 ●复煎 许多老人家习惯于一副中药“返煎”三四次，楼步青说，一般而言，一副中药在煎煮两次后，所含有效成分已大为降低，故以煎煮两遍为佳。但滋补类的中药，可以煎煮三次。而一些药量较大的处方，也可以煎煮三遍。 但需注意的是，如果将头煎与二煎的药液分别服用，这样未能将药效发挥至最佳。应该将头煎与二煎的药液混合，分早晚两次服用。同样，煎煮三遍的药液也相应地改为一天3次服用。 煎煮后——— ●立即滤取 药汤煎煮好，应趁热过滤倒出，不宜久置锅中。否则含胶体过多的药液，随温度下降产生胶凝，难以过滤，
中成药临床应用指导原则
目录 前言 (2) 第一部分中成药概述 (3) 一、中成药的剂型 (3) 二、中成药分类 (4) 三、中成药安全性 (5) 第二部分中成药临床应用原则 (6) 一、中成药临床应用基本原则 (6) 二、联合用药原则 (6) 三、孕妇使用中成药的基本原则 (7) 四、儿童使用中成药的基本原则 (7) 第三部分各论 (8) 一、解表剂 (8) 二、泻下剂 (8) 三、和解剂 (8) 四、清热剂 (9) 五、祛暑剂 (9) 六、温里剂 (9) 七、表里双解剂 (10) 八、补益剂 (10) 九、安神剂 (10) 十、开窍剂 (11) 十一、固涩剂 (11) 十二、理气剂 (11) 十三、理血剂 (12) 十四、治风剂 (12) 十五、治燥剂 (12) 十六、祛湿剂 (13) 十七、祛痰剂 (13) 十八、止咳平喘剂 (13) 十九、消导化积剂 (14) 二十、杀虫剂 (14) 第四部分中成药临床应用的管理 (15) 一、含毒性中药材的中成药临床应用的管理 (15) 二、中成药不良反应的监测 (15) 三、开展中成药临床应用监测、建立中成药应用点评制度 (15)
前言 为加强中成药临床应用管理，提高中成药应用水平，保证临床用药安全，国家中医药管理局会同有关部门组织专家制定了《中成药临床应用指导原则》（以下简称《指导原则》）。《指导原则》由四部分组成，第一部分为中成药概述；第二部分为中成药临床应用基本原则；第三部分为各类中成药的特点、适应证及注意事项；第四部分为中成药临床应用的管理。 《指导原则》是为适应中成药临床应用管理需要而制定的，是临床应用中成药的基本原则。每种中成药临床应用的具体要求，还应以药品说明书、最新版本的《中华人民共和国药典》、《中华人民共和国药典-临床用药须知-中药卷》为准。在医疗工作中，临床医师应遵循中医基础理论，根据患者实际情况，选用适宜的药物，辨证辨病施治。 第三部分各论中为更好地说明各类中成药的特点，列举了部分中成药，列举的药物是《国家基本药物目录》中的药物和《国家基本药物目录》未包括但又属临床常用的中成药。 中药注射剂的临床应用及使用管理，《指导原则》提出了具体要求，同时还应遵照《卫生部关于进一步加强中药注射剂生产和临床使用管理的通知》（卫医政发…2008?71号）执行。
养生中药材有哪些 很多人在养生的时候都会选择用到中药材来帮助自己改善，养生的方法有很多种，但是如果选择中药材来帮助自己养生的话就可以得到很不错的效果，而养生的时候选择中药材也不是什么药材都有养生的效果，这个还得我们细心的挑选，选择专业正规的药材才可以让我们得到养生的好处，那么养生的中药材有哪些呢? 1、人参粥：用人参压成粉末3克、粳米60克、用砂锅煮成粥，可食用。它有益元气，补五脏，生津液、抗衰老的作用。 2、人参茶：用人参10克、大枣10枚，用开水冲泡15分钟后代茶饮用，它有大补元气，安神益智的作用。
3、大枣养生保健应用：大枣津浓厚，其味甘美，营养丰富，药力平和，即是寻常之食品，也是常用之药品，久服或入药膳，确有补气血，益脾胃，通九窍，和百药，润肤养颜，强志延年等养生保健功效。凡体质虚弱或欲邀请书衰延年者均可食用。民间有谚曰：“一日吃三枣，一辈子不显老。”确为经验之谈。它具有护肝、抗肿瘤、中枢抑制、增强肌力等作用。服用大枣的 方法很多， 简单易行的方法有： 1、大枣粥：大枣10枚，茯神15克，小米100克，先 煮大枣及茯神，去渣，后下米煮粥。温食。 2、大枣人参汤：大枣5枚，吉林参(或高丽参)6克。 大枣人参放炖盅内，隔水炖煮1小时。分两次，温热食。人参连用2-3次，救治虚脱，人参加至15-30克，如法炖后，顿服。 在养生的时候这些中药材都是可以的，中药材养生的时
候药材必须要根据医生的要求来给自己配量，让自己吃到最健康的中药材，中药材用来养生的时候也可以选择其他的方法来帮助自己养生，养生的时候也可以多吃一些食物来帮助自己养生，但是最好的方法就是多运动运动，这样才可以得到养生的效果。
煎中药方法大全 煎中药方法大全:煎中药方法大全：一元一教你如何煎中药煎中药最好用 砂锅、砂壶或搪瓷锅，忌用铁锅。砂锅受热均匀，不会使中药的有效成分起化学变化而降低药效。一剂中药是由多味药物配起来的，每味药的性能各不相同，凡注明“先煎 煎中药最好用砂锅、砂壶或搪瓷锅，忌用铁锅。砂锅受热均匀，不会使中药的有效成分起化学变化而降低药效。一剂中药是由多味药物配起来的，每味药的性能各不相同，凡注明“先煎”者要先煎15分钟，再加入其他药。“后下”者要在药煎好以前5～10分钟放入。“包煎”者要用布袋包好再放入锅内同煎。“溶化”者则置于煎好的药液中稍加文火使其溶解。“冲服”的药是用煎好的药液送服。煎头煎药时，加冷水超过药面1～2横指，浸泡半小时，其有效成分易于煎出。用大火煎沸后，再用小火煎20～30分钟，滤渣备用。煎二煎药时水量要少些，沸后再煎15～20分钟。药品质地坚实者要多煎5～10分钟。滋补药可煎煮40～60分钟。清热解表药应少煎5～10分钟。头煎和二煎药液的量，以共计一茶杯左右为宜，混合后分两次服用。 中药汤剂，因其适应中医辨证施治,随症加减的原则,具有制备简单,容易吸收等特点，在中医临床上的应用是非常广泛的。但是,拿到一包包的中药,很多人还是会问：中药怎样煎煮呢？需要注意些什么呢？ 其实很简单，对一般中药来说，煎煮中药无非就是通过加热煎煮使中药的有效成分溶解到水里去，然后通过喝药汤达到用药的目的。 只是，整个煎煮过程中需要注意以下一些问题： 一煎前的浸泡药物在煎煮前最好能浸泡1~3小时,令药材变软,细胞膨胀，使煎药时更易煮出其有效成分。 二煎药的器具最好是选用砂锅。此外，也可选用搪瓷锅,不锈钢锅和玻璃煎器。但是不能使用铁锅,铜锅,因为铁、铜化学性质不稳定,在煎煮中药时容易发生化学反应。 三煎药的水量加水太少，可导致药物煎煮浸出不透或容易煮干；加水太多，又会导致药液太多,服用不方便。因中药质地数量的不同,不可能规定出一个统一标准的加水量，只能说做到加水适量。一般来说，可以参考下面的加水方法：将药物放入锅内,第一次煎煮的加水量以水超过药物表面3~4厘米，第二次煎煮的加水量以超过药物表面2~3厘米为宜。 四煎煮的次数实践证明,汤剂煎煮两次能够煎出所含成分的80%左右,所以一般药物最好能煎两次。煎好第一次以后，倒出药液，其药渣加水再煎一次，然后把两次煎得的药液混合起来。 五煎药的火候一般在未沸腾前采用武火，至煮沸后再改用文火,保持在微
近年来，中医西医化，中医院特色不浓，优势发挥不够，中医药治疗率偏低，居民选择中医药治疗意向偏低，中医院对当地医疗市场占有份额不高，综合效益欠佳等，这些问题不但直接制约着我院的持续发展，也影响着政府和社会对中医事业的看法。对此，我院结合本地域的现状，进行了影响中医药特色优势发挥和提高中医临床疗效的关键问题调研分析： 一、中医药特色不明显 （一）、原因分析： 医院是公益性事业单位，但财政的投入少之以少，医院长期实行自负盈亏的核算体制，医院、医生不得不为收入而工作，这就导致中医医生并不以中医手段来治病，很多采用西医为主、中医为辅的治疗方式，开药也以开西药为主，而价格低廉、简便灵验的中医药和中医特色疗法逐渐被冷落。 （二）、针对性措施： （一）突出中医药特色，发挥中医药优势 1、我院认为，中医院应该发挥中医药特色优势，紧紧围绕“中” 字做文章。以开展中医诊疗方案为切入点，将中医药特色与优势贯穿全院工作。中医特色与优势的发挥和凸显，是中医医院的办院之基。 针对农村人口多、贫困人口多、经济欠发达的县情，全面向农村基层群众推广使用中医药服务，减轻了患者就医负担。在对临床工作的考核中，突出中医特色，做到年初有计划，年中有评估，年底有总结。同时不断建立健全各项规章制度，建立科学合理的评价体系。
2、通过经济杠杆保证中医药特色和优势。我院建立新的考核机制，在每个科室体现中医特色和优势服务，并树立典型，奖优罚劣，动态管理，相互促进。针对过去有些科室很少使用中草药的情况，医院反复要求采用“中西医结合”的治疗原则，并与绩效工资挂钩，调动临床医生使用中草药的积极性和主动性。中西医学比较而言，优势应该体现在疗效更高、疗程更短、方法更简易、价格更低廉、毒副作用更小方面。中医药治疗心脑血管疾病、糖尿病、病毒性疾病、急慢性肝肾疾病、慢性支气管炎、哮喘、骨伤疾病、各种痛症、烧伤、肛肠疾病、妇科疾病、神经衰弱等等，确实有一定的优势，其优势表现在全程或一个或多个环节上。我院针对具有优势的疾病，运用中医药为主或中西医结合进行治疗。针对这些有优势的疾病，建设专科专病，形成中医专家群，专门诊疗技术、专门的协定处方或制剂，取得明显优于其他疗法的临床疗效。中医药的学术水平、临床疗效、社会认可度、市场选择度将会进一步得到提高，中医行业的核心竞争力也将会进一步增强。 3、把突出中医特色优势放在管理工作首位。中医院姓“中”应上升到医院发展战略的高度，要落实到医院发展的各个环节中去。一是深入开展“西医学中医，中医学经典”工作；二是加强临床科室中医特色发挥的动态管理，按月考核，加大奖惩力度，处罚考核较差的科室。三是切实加强独具中医特色的临床教学工作，使学生能够在夯实中医基础理论的同时，掌握多项中医药诊疗技术。四是不断加强中医药科研课题管理，将中医药特色与优势是否发挥作为申报科研课题的重要指标来评定推荐。五是定期组织开展各类中医药特色岗位练兵和技术比武活动，发掘和培育一批中医实用技术
业务学习培训2016年3月25日 中成药临床应用指导原则 第二部分中成药临床应用原则 一、中成药临床应用基本原则 1.辨证用药依据中医理论，辨认、分析疾病的证候，针对证候确定具体治法，依据治法，选定适宜的中成药。 2.辨病辨证结合用药辨病用药是针对中医的疾病或西医诊断明确的疾病，根据疾病特点选用相应的中成药。临床使用中成药时，可将中医辨证与中医辨病相结合、西医辨病与中医辨证相结合，选用相应的中成药，但不能仅根据西医诊断选用中成药。 3.剂型的选择应根据患者的体质强弱、病情轻重缓急及各种剂型的特点，选择适宜的剂型。 4.使用剂量的确定对于有明确使用剂量的，慎重超剂量使用。有使用剂量范围的中成药，老年人使用剂量应取偏小值。 5.合理选择给药途径能口服给药的，不采用注射给药；能肌内注射给药的，不选用静脉注射或滴注给药。 6.使用中药注射剂还应做到： （1）用药前应仔细询问过敏史，对过敏体质者应慎用。 （2）严格按照药品说明书规定的功能主治使用，辨证施药，禁止超功能主治用药。 （3）中药注射剂应按照药品说明书推荐的剂量、调配要求、给药速度和疗程使用药品，不超剂量、过快滴注和长期连续用药。 （4）中药注射剂应单独使用，严禁混合配伍，谨慎联合用药。对长期使用的，在每疗程间要有一定的时间间隔。 （5）加强用药监护。用药过程中应密切观察用药反应，发现异常，立即停药，必要时采取积极救治措施；尤其对老人、儿童、肝肾功能异常等特殊人群和初次使用中药注射剂的患者应慎重使用，加强监测。 二、联合用药原则 （一）中成药的联合使用 1.当疾病复杂，一个中成药不能满足所有证候时，可以联合应用多种中成药。 2.多种中成药的联合应用，应遵循药效互补原则及增效减毒原则。功能相同或基本相同的中成药原则上不宜叠加使用。 3.药性峻烈的或含毒性成分的药物应避免重复使用。 4.合并用药时，注意中成药的各药味、各成分间的配伍禁忌。 5.一些病证可采用中成药的内服与外用药联合使用。 中药注射剂联合使用时，还应遵循以下原则： 1.两种以上中药注射剂联合使用，应遵循主治功效互补及增效减毒原则，符合中医传统配伍理论的要求，无配伍禁忌。 2.谨慎联合用药，如确需联合使用时，应谨慎考虑中药注射剂的间隔时间以及药物相互作用等问题。
冬季中药养生配方 冬季中药养生配方 风寒感冒： 艾叶15、苏叶15、桔梗10、麻黄10、生姜5片。风寒感冒者往往喷嚏、流鼻涕、周身紧痛、恶寒、口淡、没有胃口、恶心呕吐、大便溏烂。在感冒的急性期在家泡泡脚能帮助尽快恢复。 防手脚冰凉： 当归羊肉汤：当归、生姜、葱白适量与羊肉同熬即可。太子参鸡汤：太子参、生姜、大枣、葱白与鸡同熬即可，这两款药膳都有温经活血、通筋活络、益气养血的作用。另外，还可用茄子根、秦艽、路路通、桑枝、桂枝单方或组合熬水泡洗。 护肤： 胡桃芝麻饮胡桃30克，芝麻20克，牛乳、豆浆各200毫升，白糖适量。将胡桃仁、芝麻研为细末，与牛乳、豆浆混匀，煮沸饮服，白糖调味，分作2份，早晚各1份，每日1剂。可补益虚损，生津润肠，润肤消斑。 防冻疮： 当归四逆汤(当归15克，桂枝12克，赤芍10克，细辛、通草、甘草各6克，大枣8枚)煎服。使阳气通、寒气散、气血通畅，则冻疮痊愈。 活血御寒：
以15克西洋参、10克田七和约50克猪蹄，加水炖1小时而成的汤，也具有补气、活血、通络的功效，适用于高脂血症、高血压、糖尿病、冠心病及骨关节炎患者。 防口干舌燥： 将莲子芯用开水沏还可以有效地去火，不要过浓也不要过淡，日饮二三次可预防口干舌燥、虚火上升、嗓子疼痒、声音嘶哑、脑觉昏沉等。同时还可治疗咳嗽。 冬季养生中草药 1、黄芪 补气升阳、益精固表、利水退肿，适用于自汗、盗汗、浮肿、内伤劳倦、脾虚、泄泻、脱肛及一切气衰血虚之症。但有高热、大渴、便秘等实热症者忌用。民间流传着“常喝黄芪汤，防病保健康”的顺口溜，意思是说经常用黄芪泡水当茶喝，具有良好的防病保健作用。黄芪茶能补中盖气，而且补而不腻，可改善气虚和贫血，增强体质，延年益寿，若熊再加上枸杞、党参、茯芩等药材一起冲泡，效果更好。 2、西洋参 补气养阴、清火生津，适用于肺虚劳嗽、久咳、喘咳、咯血、失音等症。激烈活动后疲劳乏力、大汗虚脱者服用不错。西洋参补气，性凉，所以服用后不会上火，气阴两虚之人尤宜。气虚之人主要表现为神疲乏力、少气懒言、呼吸短促、动则汗出、声音低微等;而阴虚则会表现出一派虚热之象，如舌质红、口干舌燥、眼干、手心发热等。这类人服用西洋参最简单的方法就是把西洋参切成片，或者去药店买些现成的西洋参片，每天拿两片
常见的中药煎煮方法 中药煎煮质量的好坏直接影响药物治病的疗效，我国历代名医都十分重视中药煎煮方法。汉代医家张仲景将煎煮用水分为雨水、千扬水等多种；徐灵胎认为：“煎药之法，最宜深讲，药之效不效，全在乎此”；李时珍指出了药液煎煮不当的不良后果：“凡服汤药，虽品物专精，修治如法，而煎药者鲁莽造次，水火不良，火候失度，则药亦无功。”这些都说明古人已认识到煎煮过程中有诸多因素影响煎煮质量，而煎煮质量的好坏直接影响了中药药效的发挥。 中药煎煮过程中要发生两种变化：一是药物有效成份的溶出；二是药物中各种生理活性成份进行化合反应。因此，汤剂的煎制方法有许多特殊的讲究。 可见中药的煎煮方法对于有效地利用药物和提高治疗效果十分重要。中药的合理煎煮可以充分地发挥药物的作用，对于防治疾病均有重要意义。中药的煎煮是多方面的，主要包括： 1.清洗
中草药大都是生药，在出售之前一般都进行了加工炮制，煎煮之前一般没有必要淘洗。如果的确觉得草药有些脏，可在浸泡前迅速用水漂洗一下，切勿浸泡冲洗，以防易溶于水的有效成份大量丢失，从而影响中药疗效。 2.器具 一般选用砂锅，搪瓷器皿次之，千万不要用铁锅或铝锅等金属器皿。详见前发博文：煎中药，用什么容器最好？ 3.浸泡 中药饮片煎前浸泡既有利于有效成份的充分溶出，又可缩短煎煮时间，避免因煎煮时间过长，导致部分有效成份耗损、破坏过多。多数药物宜用冷水浸泡，把药物倒入药锅内摊平，然后加常温水——室温水浸泡60分钟，轻压药材时水高出药平面约2厘米。以药材浸透为原则。夏天气温高，浸泡时间不宜过长，以免腐败变质，冬季可以长些。特别需要注意的是浸泡中药绝对不能用沸水浸泡。 4.用水 煎药用水必须无异味、洁净澄清，含矿物质及杂质少。一般来说，凡人们在生活上可作饮用的水都可用来煎煮中药。一般可用清澈的泉水、河水及自来水，井水则须选择
十大泡脚之中药泡脚配方大全 一、泡脚的优点：1、安全；2、无痛苦；3、无毒副作用；4、廉价；5、有效；人们常说“春天洗脚，升阳固脱；夏天洗脚，暑湿可祛；秋天冼脚，肺润肠濡；冬天冼脚，丹田温灼”,哈，这些生活小窍门应该好多人都知道的吧；6、方便；7、舒适。 二、泡脚水的组成：泡脚水—般取自来水、河水、等为基水。通过泡脚治病的可根据不同疾病加入不同的药物。 三、泡脚水的温度，泡脚水的温度以30~38度为宜，但最好不要超过40度。 四、泡脚容器的要求：1、质地应无害、安全、保温性能好。2、高度一般泡脚盆的高度最好超过20厘米［没过踝关节］。3、结构可买一些微电脑浴脚器。 五、泡脚时间的要求：1、每天安排泡脚几次：如一般保健泡脚，每天一次即可；如患有某种疾病每天至少2次以上。2、每天什么时间泡脚为宜：如2次，一般上午lO 点1次，晚上睡前1次，因为睡前泡脚对消除疲劳大有好处可使人睡得更甜容易进入“倒床不复闻钟鼓”的境界。广为流传的“饭后三百步，睡前一盆汤”、“睡前洗脚，胜吃补药”。 3、每次泡脚时间多长为宜：一般为30分钟以上，但对于如慢性风湿性关节炎、慢性高血压等要适当延长一些。每次具体时间还需根据泡脚者的年龄、性别、疾病情况等及泡脚后的感受来逐渐调整。 六、泡脚与按摩的关系：传统的泡脚不包括按摩，现在有了新变化，有些厂家已生产出泡脚与按摩同时进行的泡脚盆，如没买到可用手做一些足底按摩或一些其它按摩这样结合起来效果最好。 七、泡脚的作用：1、清洁皮肤的作用。2、扩血管作用。3、降低血液粘稠度。4、缓解肌肉痉挛。5、镇静作用。 八、泡脚的注意事项：l、注意卫生［最好以家庭为主］。2、切忌求快。3、切忌三天打鱼、两天晒网，要坚持不断才能受益终生。4、儿童禁止泡脚。5、某些急性感染性疾病禁止泡脚。6、出血性疾病禁止泡脚，［包括急性外伤出血，如泡脚会引发意外后果不堪设想］。 高血压：钩藤40克、夏枯草30克、桑叶20克、菊花20克。 失眠：磁石60克、菊花20克、黄芩15克、夜交藤30克。 肝脾肿大：三棱15克、莪术15克、延胡索15克、乌梅10克。 遗精、早泄：仙鹤草40克、黄芩10克、丹皮10克、芡实30克、女贞子30克、狗脊15克、桑葚30克、知母12克、黄柏12克。 足跟、足踝关节痛：寻骨风30克、透骨草30克、鸡血藤30克、乳香10克、没药10克、血竭10克、王不留行15克。 将泡脚方剂煎汤至2000毫升左右，水温保持在40摄氏度上下为宜。
熬中药的最佳方法！在家自己熬中药，放心又省钱 引导语：现在虽然西药已经很普遍的被运用，但是还是很多人对中药深信不疑。尤其对慢性病，很多人还是选择中药治疗。很多大医院都提供了中药代煎服务，殊不知为了提高煎药的效率，很多都只熬一次装袋了事，浪费了药材。在家自己熬中药，放心又省钱。 如何选择熬药的锅 1.熬中药最好的是陶瓷器皿中的砂锅、砂罐，因其化学性质稳定，不易与药物成分发生反应，并且导热均匀，保暖性好。 2.其次可用白色搪瓷器皿或不锈钢锅。 3.煎药忌用铁、铜、铝等金属器具。因为金属元素容易与药液中的中药成分发生化学反应，可能是疗效降低，甚至产生毒副作用。 煎药前的浸泡 药物在煎煮前一定要浸泡使有效成分易于浸出。一般以花、叶、茎类为主的药物，浸泡时间一般为15分钟。以根、种子、根茎、果实类为主的药物浸泡时间一般为半小时。
煎煮次数 以两次或三次为宜。 熬药方法 1.先把药放在砂锅里面，根据药的多少加水，加的水必须漫过中药(千万不要加少了，否则熬中药容易靠干)。大约泡20分钟左右。泡的过程中最好不时用搅拌棒搅一下，这样泡的均匀一些。天热时可以加凉水泡，天冷时用凉水要延长浸泡时间，或者可用温水，这样效果更好邮箱。 2.中药泡好后，放在煤气炉子，或者蜂窝炉子上面开始熬制。在中药熬开之前用大火，熬开之后转为小火慢慢熬，小火熬制大约20分钟。看着表，到时间后，如果药汤还是很多，可以再继续熬一会。切记千万不能把中药熬干了。熬完后，可用一根筷子放在砂锅沿上挡住药渣，再用药淋子过滤，这样效果更好， 3.第二遍熬制时，可以加温水，加的水应是第一次的一半。也是熬20分钟左右，熬完看看药汤多不多，如果不多就可以直接倒出来。若药汤较多，可以再多熬一会儿。 4.有时间的话可以熬第三遍，加的水是第二遍的水的一半。但是一般情况下，熬两遍就可以了，第三遍的药力很低了，但可以熬完用药汤泡脚。 5.在倒中药时，一定要小心烫手。盛中药的器具最好是陶瓷的或不锈钢的，这样中药不易与器具发生化学反应。 煎煮时间
1,要选好煎熬中药的容器. 煎熬中药最好是沙锅,陶瓷瓦罐（铝制品,搪瓷器也可用）,忌用铁器.因为陶瓷化学性质稳定,在药物水煎复杂的化学应中,不会“干扰”药物的合成与分解,导致影响药效.而我们常用的铁锅容器在药物煎煮过程中,极易同中药内所含的鞣酸质,甙类等成份起反应,造成药物的疗效降低或失效,以至发生反作用,所以不宜使用.一定不要不锈钢或铁锅熬中药.因中药中含有多种生物碱以及各类生物化学物质,尤其在加热条件下,会与不锈钢或铁发生多种化学反应,或使药物失效,甚至产生一定毒性. 2,要掌握正确的煎煮法. 药物入锅后,先用凉水浸泡半小时,使药的有效成份易煎出.放水量要注意,一般放水要高出药面少许,治水肿病的药宜少放水；小孩药要少放水,发汗药可多放水.放水要一次放足,不可中途加凉水,切不可用沸水煮药,以免药物表面蛋白质变性,而影响有效成份析出.煎中草药时,为了使药煎透,最好是加盖煎.尤其是含有挥发性成份的中草药,如薄荷,苏叶,藿香,佩兰等,更要盖好盖,并要在短时间内煎好,以减少有效成份的挥发；有些贵重药物,如人参,鹿茸等也要盖住,并要用文火细煎.煎药要掌握好火候.一般未沸前用急火,沸后用文火.如解表发汗的药,猛火煮沸3～5分即可；熟地,山萸之类补益药则宜用文火煎,煮沸后再煎20分钟左右.此外,绒毛类药物及散剂煎煮时宜做成布包入锅,以减少绒毛对喉的刺激.对于抓的特殊药物,先煎,后煎,冲服,包煎等,都要遵医嘱. 3,要掌握服用方法. 中草药有“冷服”,“热服”之说,服药时间也有讲究.解表药一般宜温服,为了达到发汗的目的；祛寒药也宜热服；解毒药,止咳药,清热药则应冷服；滋补药宜空腹温服,易于消化吸收,但量不宜太多；安神药在睡前半小时,以加强药物作用；脾胃虚弱者宜饭后服药,对胃肠有较强刺激的药物更应饭后服；泻下药须空腹时服,而不宜于夜间服用,大便通畅后则应停药；糖尿病人口渴时服,不拘时间；驱虫药早晚空腹时服,利于驱虫；口腔咽喉病人宜含药,充分发挥药物局部作用；呕吐病宜少量多次饮药,减轻胃的负担,或先服姜汁少许,以降逆止呕；小孩及体弱患者,药量宜少；妇女孕期服药更要谨慎.中药不宜用茶水和乳汁送服,因茶叶,乳汁易和某些药物发生化学作用,降低药效. 4,煎煮中药的要求. ①每次将一剂中药饮片材料放入煲内,加入清水,观察加水能否浸满药面,不足时可稍加水量. ②一般浸泡半小时使中药饮片的有效成分易于煎出(如赶时间,此步骤可略去). ③先用猛火煎至充分沸腾1-3分钟.然后收至小火,煎20-30分钟使之成一碗,用消毒纱布或咖啡格滤渣倒入杯内,温热服用. ④一次将药物煎好后,可以将首剂和再煎的药物混匀,以便药效均衡. 最后一定注意服用的时间,应当仔细询问医生,记在纸上.
十大泡脚之中药泡脚配方大全 1.、泡脚的优点：1、安全；2、无痛苦；3、无毒副作用；4、廉价；5、有效；人们常说“春天洗脚，升阳固脱；夏天洗脚，暑湿可祛；秋天冼脚，肺润肠濡；冬天冼脚，丹田温灼”,哈，这些生活小窍门应该好多人都知道的吧；6、方便；7、舒适。 2.、泡脚水的组成：泡脚水—般取自来水、河水、等为基水。通过泡脚治病的可根据不同疾病加入不同的药物。 三、泡脚水的温度，泡脚水的温度以30~38度为宜，但最好不要超过40度。 四、泡脚容器的要求：1、质地应无害、安全、保温性能好。 2、高度一般泡脚盆的高度最好超过20厘米［没过踝关节］。 3、结构可买一些微电脑浴脚器。 五、泡脚时间的要求：1、每天安排泡脚几次：如一般保健泡脚，每天一次即可；如患有某种疾病每天至少2次以上。2、每天什么时间泡脚为宜：如2次，一般上午lO点1次，晚上睡前1次，
因为睡前泡脚对消除疲劳大有好处可使人睡得更甜容易进入“倒床不复闻钟鼓”的境界。广为流传的“饭后三百步，睡前一盆汤”、“睡前洗脚，胜吃补药”。3、每次泡脚时间多长为宜：一般为30分钟以上，但对于如慢性风湿性关节炎、慢性高血压等要适当延长一些。每次具体时间还需根据泡脚者的年龄、性别、疾病情况等及泡脚后的感受来逐渐调整。 六、泡脚与按摩的关系：传统的泡脚不包括按摩，现在有了新变化，有些厂家已生产出泡脚与按摩同时进行的泡脚盆，如没买到可用手做一些足底按摩或一些其它按摩这样结合起来效果最好。 七、泡脚的作用：1、清洁皮肤的作用。2、扩张血管作用。 3、降低血液粘稠度。 4、缓解肌肉痉挛。 5、镇静作用。 八、泡脚的注意事项：l、注意卫生［最好以家庭为主］。2、切忌求快。3、切忌三天打鱼、两天晒网，要坚持不断才能受益终生。 4、儿童禁止泡脚。 5、某些急性感染性疾病禁止泡脚。 6、出血性疾病禁止泡脚，［包括急性外伤出血，如泡脚会引发意外后果不堪设想］。 高血压：钩藤40克、夏枯草30克、桑叶20克、菊花20克。
标准的水煎中药法 “水煎中药好？还是科学中药强？”的确，科学中药服用起来方便、也比较经济，而水煎中药(简称水药)则虽然“料理”起来比较麻烦，但是也由于水煎药的“量身定作”、以及“现煮现喝”，所以药效方面往往比较令人满意，对于重病、急病或慢性病的治疗，水药的效果是相当迅速的，相对地，如果只是感冒、肠胃炎等轻微的疾病，只要用科学中药就相当有效，不必一定要“花钱”喝水药才行。 问题是，水煎药材一定是四碗水煮成一碗吗？其实不然，由于水煎药材的成分不同、煎煮的方法当然要“因药而异”，才能充分发挥每味药材的药效；临床上，常见的药材煎煮法有下列几种： 1. 标准煮法：药材为一般的补药(例如包含有当归、黄芪、熟地) 步骤一：先将药材放入锅内(不可使用铁锅或铜锅)，加入适量的水(超过药面约2公分、大约4碗水)，浸泡半个小时。 步骤二：以武火(大火)煮沸后，再以文火(小火)煮到剩一碗水(约250ml.)，将煮好的药液倒出于碗中，再重复用三碗水煎煮成8分，前后两次混合服用。 2. 快煮法：药材含有挥发性中药(例如银花、菊花等治疗感冒的中药) 步骤一：先将药材放入锅内，加入适量的水(刚好盖过药面，约三碗水)，浸泡半个小时。 步骤二：以武火(大火)煮沸后，再以文火(小火)煮10分钟，不可打开盖子，煮一次即可服用，不必重复一次。 3. 后下法：药材中含有不耐煮的中药(例如天麻、钩藤、麻黄等) 步骤一：先用四碗水浸泡药材约半个小时。 步骤二：以武火(大火)煮沸后，再以文火(小火)煮到剩一碗水(约250ml.)时，再将不耐煮的中药(一般会另外包)投入再煮5分钟，将药液倒出于碗中即可服用。 4. 先下法：药材中有一些需久煮的矿物类(例如：滑石、石膏)。 步骤一：先将矿物类的中药(另外包)加入六碗水，先煎30分钟后，再加入其它药材。 步骤二：以武火(大火)煮沸后，再以文火(小火)煮到剩一碗水(约250ml.)，将煮好的药液倒出于碗中服用即可。 5. 另煎法：药材中含有一些名贵药材(例如人参) 步骤一：先将一般药材加四碗水放入锅内，浸泡半个小时，大火煮沸后，小火煮到剩一碗水，放凉。 步骤二：将名贵药材的药材，加一碗水，于电饭锅中蒸30分钟后，取出与上述药液相混，即可服用。 “良药苦口”，中药本来就不太好吃，何况是水煎中药？可是，如果真的病情需要服用水煎中药，那么，至少也要“料理”正确、水药的药效才会彻底出来，所以，如果有机会服用水煎中药时，别忘了问问您的医师、该如何煎煮您的中药，可不是一味地“四碗煮成一碗”！
中国传统养生十大滋补品+滋补养生中药50味 中国传统养生十大滋补品 冬季是滋补的时候，中华传统养生佳品有着全方位的养生功效。你知道传统的养生滋补品都有哪些吗？让我们一起来看看，为爸妈、为家人、为自己选好冬季滋补品，过一个暖暖的冬天。 一、冬虫夏草- 高原瑰宝稀世仙草冬虫夏草，冬则为虫，夏则为草，聚天地之灵气而形成，是上等滋补品，生长在海拔数千米的青藏高原高寒地带，以青海和西藏所产为佳品，又以青海玉树所产为上品之代表，被誉为“草虫之王”。冬虫夏草性温暖，补精益髓，具有良好的免疫调节功能，能减轻放化疗的毒副作用，能够抑制肿瘤、抗病毒、防衰老及保护心、脑、肾、肝细胞等，适宜亚健康人群、癌症患者、肾气不足者、月经不调者、三高人群等。 二、燕窝- 造化珍馐虚劳圣药燕窝，燕科动物分泌物筑垒而成的巢穴，形似元宝，是传统的名贵滋补食品，有“东方珍品”之美称。燕窝甘淡平，对肺的调养极有益处。有组织修复和提高免疫功能，其固有成分能促进生长发育，延缓衰老及安胎。微信号：YujiaoWeikan另外，燕窝对加快癌症患
者手术后康复及减轻放疗副反应有一定功效。 三、海参- 八珍魁首海中人参海参，又名海鼠、海男子，是生长在海洋底层岩石上或海藻间的一种棘皮动物，有着神奇的自我修复功能，被誉为长寿之神。海参有着无与伦比的营养结构，具有高蛋白、低脂肪的特点，能够滋阴补血，健阳补肾，润燥调经，养胎利产，具有显著的抗肿瘤活性及免疫调节作用，其抗氧化作用优于天然抗氧化剂维生素C。 四、西洋参- 北美灵参补益尚品西洋参系五加科人参属，是生长于北美原始森林之中的古老植物，素有“森林活化石”之称。受墨西哥暖流和五大湖影响，周围地区空气相对湿润、气温偏冷、雨量充沛，特别适宜西洋参生长。西洋参性寒，味甘微苦，除补气作用之外，更具泻火除烦、养胃生津之功，专补人参之不补者。具有广泛的保健功效，可以改善中枢神经功能，增强记忆力和人体免疫力，保护心血管系统，调节胰岛素分泌、促进糖代谢和脂肪代谢，对糖尿病有辅助治疗作用。 五、灵芝- 神芝还魂瑞草延年灵芝，自古以来被民间视为吉祥、富贵、美好的象征，素有仙草、瑞草之称，是中医药宝库中的珍品，是滋补强壮、固本扶正的珍贵中草药。我国
老中医解读：中药该怎么熬，才能熬出药效 熬中药真的是一门学问，说难真的不难，只要掌握正确的方法，就能很轻松的熬出最好的中药。相信很多东西是我们这些平常人想问但是又不敢问的。今天我们特地请教了我们的神农中医馆馆长曹教授，让他给我们讲讲如何如法熬药、服药，药效翻倍。 首先曹教授跟我们谈起，确实现在有多数药店都提供代煎中药服务，一般花费两三个小时就可以取到机器熬好的中药，拿到中药后可以放到冰箱里，随时取用，非常方便。但是呢，这样搞出来中药虽然效果不错，但除非实在没有时间、没有条件，中药最好还是自己熬，毕竟这样搞出来的药效估计只有90%左右。因为熬药是有很多细节，代煎服务不一定给你全部都注意到，除非她真的很用心在做。 熬药的步骤： 一：买回中药，别急着就把中药洗完就拿去熬药，可以先将中药用冷水浸泡半个小时。需要先煎、后下或者一些粉末状的药材则不用浸泡。 二．第一次熬药：用大火先把中药煮开，再用文火煮20-30分钟，然后就把熬出来药导入大碗中。后下药则在第一次熬药中的最后几分钟放下药煲里面。 三．第二次熬药：在上面的药渣中加入水，用大火煮开，然后用文小火煮跟第一次熬药时同样的时间，然后熬出来的要倒入碗中。 四．把上面两次煎熬出的中药混合，混合均匀，分2次喝完。 熬药小技巧 一．我们在熬药的时候，一定要用陶瓷制的药煲，千万不能使用金属器物。毕竟基本很多中药材都是草木，而金克木，金属制的器皿是能改变中药材的药性。而土生万物，陶瓷制的药煲属土。 二．我们每次熬药的水量我们曹教授是推荐三碗水熬成一碗水，这样能保障药的药效，每一次熬药的时候，保证加水的量，千万不能边熬药边加水，这跟煲汤是同样的道理。 三．熬药的时间，指煮开后用文火慢煮的那段时间，一般是二十到三十分钟。总的来说中药熬的时间太长，反而会降低中药的药效。
中医中药在外科手术中的临床运用中医中药在外科手术中的临床运用 1手术前中医药的辅助治疗 1.1改善患者全身营养状况 许多腹部外科疾病潜在发病时间长，致使患者伴有营养不良、低蛋白血症等。这些并发症直接影响病人接受手术治疗及术后效果。对这些患者近年来有人主张采取胃肠外营养等治疗措施。这些措施能改善病人全身营养状况，但因治疗费用昂贵且并发症多而难于推广。中医理论认为这些需手术治疗的病人往往存在着各种“虚证”，采取“虚则补之”的治疗法则，能改善机体全身状况，为手术创造良机。按照中医理论“整体观念、辨证论治的特点”选用以十全大补汤、补中益气汤加减”对手术前的气血虚证患者具有良好的疗效；采用静脉点滴参麦或黄芪注射液治疗术前虚证病人，也获得较好疗效。在乳腺癌手手术前给予患者积极的中医药治疗，可改善机体一般状况，增强体力，调理因疾病引起的肝肾功能障碍，有利于手术进行，对控制肿瘤的发展和潜在的转移也有帮助。 1.2增加胃肠动力和洗涤肠胃 在消化道手术中，采取通里攻下法选用大承气汤或调胃承气汤加减做术前准备，此类中药有明显增加胃肠动力和洗涤肠胃积滞的作用外，还有改善脏器血流及腹膜吸收，促进术后肠功能早期恢复，预防肠源性感染和内毒素血症的作用。许多外科危重病
人，由于感染、中毒、失血等因素存在，导致病情恶化，甚至休克。冒然施行手术往往适得其反。若在采取输血、补液、抗感染、抗休克治疗措施的同时加以中医辨证论治，往往能为手术治疗创造条件 采用清热解毒、通里攻下等药物组成的“清胆汤”和“消胰汤”治疗急性梗阻性化脓性胆管炎和出血坏死性胰腺炎合并休克病人，成功率达到80%～90%；对创伤性休克和过敏性休克患者，使用独参汤加味再辅以输血、输液等综合抗休克措施，可使血压迅速上升，为手术的实施提供了良好保证。 2手术后中医药调理及营养支持治疗 2.1营养支持和免疫调理作用 肠道除了具有传统认为的营养功能之外，还具有内分泌、免疫和屏障功能。肠道的屏障功能是防止肠源性感染的重要环节。近年来，用中药和肠内营养治疗胰十二指肠切除术后的患者，发现胰十二指肠切除术后早期实施中药和肠内营养支持治疗有助于胃肠功能的早日恢复、营养状况的改善和免疫功能的提高，并有助于减少术后并发症的发生。采用中药黄芪注射液与全肠外营养，联合应用可改善围手术期梗阻黄疸病人营养状况及提高病人免疫的功能。术后常规服用十全大补汤具有增高病人血浆白蛋白、血红蛋白的作用，可做为外科营养治疗之良方。 2.2术后中医药恢复肠道功能的作用 腹部外科疾病术后常出现腹胀，肠粘连等症状，选用自拟扶正还原汤（主要由大黄、白术、枳实、厚朴、莪术、槟榔、甘草组成）水煎内服或从胃管注入，治疗98例腹部手术后出现的腹胀
中药养生的十大药材 何首乌——滋补良药 推荐明星：王菲。王菲生产后的调理秘方中，何首乌是一味主药。每天吃，使她恢复了孕前的身材。 青春奇效：何首乌最青春的功能是乌发，经炮制后，它可以补肝肾、益精血，而且不寒不燥，实为乌发良品。后来西医研究又发现了它降脂、降糖、防止动脉粥样硬化、减少体内细胞氧化的功能。 如何使 用： ●首乌洗发：60 克，用水煎煮，水色成浅黄即可使用，先用普通洗发水洗净头发，然后用何首乌汁浸洗。 ●首乌蛋：何首乌60 克、鸡蛋2 只。将何首乌洗净、切片，放人砂锅内，加进鸡蛋，煮至蛋熟，捞出去掉蛋壳，再放锅内煮片刻，加盐、味精即可。 黄芪——最能补气 推荐明星：赖雅妍。台湾“大长今”每到换季之时，就用黄芪水沐浴，以增强抵抗力，防止感冒。 青春奇效：中医认为，能补一身之气，可以用来缓解气虚乏力、中气不足。现代研究发现，黄芪还能增强代谢，使机体细胞生长旺盛，同时消灭体内自由基，预防衰老。
如何使用： ●代茶饮：黄芪5~6 片，用沸水冲泡饮用，可以缓解身体困倦、无力、气短。 ●煨枣：取大枣30 个、生黄芪30 克，一同煨煮。可以提高免疫功能，增强体质。 人参——补充元气 推荐明星：袁咏仪。她表示怀孕时，妈妈教她用水冲蜜糖和人参喝，所以产后的袁咏仪中气十足。 青春奇效：提及的抗衰老功能，是最没有悬念的一个。据现代药理研究，人参可以激化中枢神经系统，具有提高脑力和体力的作用，并能迅速消除疲劳。 如何使用： ●人参茶。人参片3 克，用沸水冲泡后代茶饮。能强心安神，迅速消除疲劳。 ●人参粥。人参3 克、粳米100 克，用文火煎熬至熟。适宜在秋冬季早晚空腹食用，用来补充元气，调和五脏。 三七——调血妙品 推荐明星：姚明。姚明最近一次受伤为胫骨有骨裂的迹象，为了促进愈合，营养师开出了用三七炖肉鸽的食疗配方。 青春奇效：在中医史上，三七很早就出名了，但一直被作为止血、定痛、消肿的伤科补品来关注。近些年才发现，它还可以扩张血管、