壳聚糖复合生物防腐剂的抑菌效果研究

本实验研究了以壳聚糖及其金属锌螯合物为主复配的五个生物防腐剂配方以及配方中的主要组分在不同pH条件下的抑菌效果。研究选定的五种指示菌是大肠杆菌(Escherichia coli)8099、金黄色葡萄球菌(Staphylococcus aureus)ATCC6538、枯草杆菌黑色变种(Bacillus subtilis)ATCC9372、白色念珠菌(Candida albicans)ATCC10231和黑曲霉(Aspergillus niger ATCC16404),采用平板培养计数法检测了从加入不同种类的菌悬液至含抑菌剂和培养基的试管中培养0、4、24、48、72和96h的活菌残留数,以活菌残留数的对数值随时间变化的趋势表示抑菌效果。结果表明:配方A、B、D、E在p H6.0以下可以抑制五种指示菌。配方A～E和单组份壳聚糖金属锌螯合物对大肠杆菌、金黄色葡萄球菌和枯草芽胞杆菌的抑制效果好,而对羟基苯甲酸乙酯主要对枯草芽胞杆菌和黑曲霉的抑制较好,对其余三种菌的抑制效果较差,所有配方对白色念珠菌的抑菌效果较其它四种指示菌的抑制效果差,这是由于实验的起始菌悬液浓度较高造成的,同一配方在起始菌悬液浓度和作用pH值均低的情况下抑菌效果好。     

植物精油对微生物的抑菌效果评估研究

本实验以大肠杆菌(Escherichia coli)8099、金黄色葡萄球菌(Staphylococcus aureus)ATCC6538、枯草杆菌黑色变种(Bacillus subtilis)ATCC9372、白色念珠菌(Candida albicans)ATCC10231和黑曲霉(Aspergillus niger ATCC16404)为指示菌,采用纸片法和试管法评估了42种植物精油的抑菌效果,结果显示植物精油对真菌和革兰氏阳性细菌的抑杀效果好,其中有11种精油对5种指示菌的抑杀效果较好,由强至弱的顺序为:肉桂、百里香、柠檬草、香茅、香茅油、雪松、山鸡椒、天竺葵、桂皮油、芫荽、罗勒、山苍子油、欧薄荷、茴香。     

基于琼脂扩散法的抗菌肽定量测定方法优化

抗菌肽在试验室研究、生产实践以及安全性评价过程中,存在定量测定方法不统一,结果重复性差,缺乏比较依据等问题。以硫酸多粘菌素B为代表,以铜绿假单胞菌为指示菌,采用琼脂扩散法建立抗菌肽浓度对数值与抑菌圈直径之间的线性关系,以重复性和精密度为指标,对指示菌菌龄、菌浓、琼脂浓度和培养基厚度等影响因素进行优化,确定琼脂扩散法测定抗菌肽效价的最佳测定条件为:指示菌的最佳培养时期为稳定期,浓度为10~6 cfu/mL,培养基厚度为3.15 mm(Φ90 mm,20 mL),琼脂浓度为1.5%。在此条件下,选用其它抗菌肽和指示菌进行验证,结果重复性好,精密度高(R~2=0.997),该研究将为抗菌肽在试验和生产过程中的定量测定提供参考和依据。     

琼脂扩散法定量测定多黏菌素对霉菌的抑菌活力

本文研究抗菌肽抑制霉菌的活力定量测定方法,选用多黏菌素B和产黄青霉作为研究对象,采用琼脂扩散法优化指示菌浓度、抗菌肽添加量、培养时间等影响活力定量测定的条件,进一步以黑曲霉为指示菌进行方法验证;通过建立抗菌肽浓度对数值与抑菌圈直径之间的线性方程进行抗菌肽抑菌活力的计算。研究结果显示,琼脂扩散法测定多黏菌素B抑制霉菌活力的最佳条件:90 mm平板添加PDA培养基15 mL,指示菌孢子浓度(1～5)×106个/m L,多黏菌素B添加量100μL,4℃预扩散时间6～10 h,28℃培养时间24h。以黑曲霉为指示菌进行方法验证时发现方法重复性好、精密度高。在此条件下,多黏菌素B对产黄青霉和黑曲霉的抑菌活性分别为524.81 U/mg和605.76 U/mg。该研究为抗菌肽在试验和生产过程中的活力测定提供参考和依据。     

产脂肪酶极端丝状真菌的筛选及其酶学特性

从南非德班市及其周边地区采集的76个土样中筛选得到99株嗜碱丝状真菌和51株嗜热丝状真菌,利用橄榄油乳化法以及对硝基苯酚法筛选得到三株具有底物专一性的菌株F4-173、F4-262、F8-67,经ITS分子生物学鉴定方法分别鉴定为Aspergillus fumigatus、Penicillium verrucosum、Penicillium chrysogenum。其中,Aspergillus fumigates F4-173可专一性分解菜籽油,其最适作用条件40℃、p H8.0,Penicillium verrucosum F4-262可专一性分解对硝基苯酚月桂酸酯,其最适作用条件为40℃、p H8.0,Penicillium chrysogenum F8-67可专一性分解大豆油,其酶液最适作用条件为50℃、p H6.5。     

解淀粉芽孢杆菌HRH317菌株抗菌肽发酵条件优化及其抑菌活性研究

本文以抑菌圈直径为指标,采用单因素试验和响应面法对解淀粉芽孢杆菌HRH317菌株产抗菌肽的发酵条件进行优化,并对该抗菌肽的抑菌谱及其最小抑菌浓度(MIC)进行了研究。得到最佳发酵条件为：pH7.5,装液量为50 mL,接种量3%,培养时间13h、温度38.5℃、转速145r/min,抗菌肽含量441.56±1.24 μg/mL。在此条件下抗菌肽的抑菌圈直径达15.95 mm。针对9种指示菌的抑菌谱结果表明,抗菌肽对枯草芽孢杆菌、金黄色葡萄球菌抑制效果最好,其次是霉菌、酵母菌。其中,抗菌肽对枯草芽孢杆菌、金黄色葡萄球菌的MIC为 3.45 μg/mL,大肠杆菌、白地霉、巴氏醋酸杆菌MIC为6.9 μg/mL,禾谷镰孢菌、黄曲霉、假丝酵母MIC为 55.20 μg/mL,红酵母MIC为110.40μg/mL。     

不同培养基对黑曲霉菌丝生长影响的研究

通过选择察氏培养基不同种类碳源、pH值和外源添加物,观察黑曲霉（Aspergillus niger）菌丝生长状况,并测定菌丝球干质量和直径,分析黑曲霉菌丝最适生长条件。结果表明,察氏培养基最佳碳源为葡萄糖,60 h形成较大菌丝球,外表光滑,菌丝球干质量和直径分别为0.023 g和0.155 cm;培养基最佳pH值为8.0,60 h形成外表光滑的菌丝球,且直径较大,菌丝球干质量和直径分别为0.018 g和0.147 cm;培养基中最佳外源添加物为吐温-80,60 h菌丝成球且密度密集,菌丝球干质量和直径分别为0.006 g和0.106 cm。因此,黑曲霉菌丝最适生长条件为察氏培养基中添加葡萄糖,pH值为8.0,外源添加物为吐温-80。     

白色金针菇摇瓶发酵培养基的优化研究

以菌丝球干质量、密度、直径为考察指标,利用单因素试验和L9(34)正交试验对白色金针菇摇瓶发酵培养基进行优化。结果表明,最适摇瓶培养基配方为：葡萄糖10 g/L、红糖13 g/L、黄豆粉3 g/L、KH2PO4 0.5 g/L、MgSO4 0.5 g/L、VB1 0.01 g/L。在此条件下,白色金针菇菌丝球干质量达到15.48 g/L,菌丝球密度66.48个/mL,菌丝球直径为1.45 mm。     

响应面优化发酵磨盘柿褐变条件

黑色食品(Mat Black,MB)与其同类非黑色食品相比,具有较高的营养价值和保健功能。本课题以成熟的柿子为原料,通过接种可食用微生物来探究磨盘柿发酵加工黑柿的最佳工艺条件,为进一步加工柿子黑色产品提供一定技术参考。将黑曲霉(Aspergillus niger)、毛霉(Mucor sp.)、根霉(Rhizopus sp.)、米曲霉(Aspergillus oryzae)接种到经预处理后的柿子浆中恒温发酵,比较4种真菌对发酵柿子褐变程度的影响,得到Asp.niger是柿子黑色产品的最适微生物。通过单因素和响应面试验优化Asp.niger发酵柿子褐变条件,得到最佳发酵条件为:500.00 g柿子浆中接入Asp.niger菌悬液的量8 mL (1×10-4 g/mL)、温度33.5℃、时间72.5 d,在此条件下褐变度可达到5.62。本文比较4种真菌对发酵柿子褐变程度的影响,通过单因素和响应面实验优化Asp.niger发酵柿子褐变条件,得到影响褐变的显著因素和适宜条件,可为柿子黑色产品开发提供技术借鉴。     

响应面法优化黑曲霉产抗菌物质的发酵培养基

以抑菌圈直径为响应值,采用响应面法对黑曲霉（Aspergillus niger）xj产抗菌物质的发酵培养基进行优化.首先通过单因素试验确定最适碳源和氮源分别为麦芽糖和麦麸,并初步考察了麦芽糖、麦麸、无机盐、生长因子及表面活性剂的最适添加量.通过Plackett-Burman设计筛选出影响抗菌物质活力的显著因素：无水硫酸镁、氯化钙和吐温-80.最后通过中心复合设计及响应面分析确定产抗菌物质的的最优发酵培养基组分为麦芽糖30.00g/L、麦麸7.50g/L、酵母膏1.00g/L、磷酸氢二钾1.00g/L、无水硫酸镁0.2982g/L、吐温-80 0.24mL/L、微量元素液2.00mL/L、氯化钾0.50g/L.采用滤纸片扩散法检测优化后的发酵液抗菌活性,抑菌圈直径为（27.26±0.41）mm,比未经优化测得的抑菌圈直径提高了83％.     

Chemical Composition and Antimicrobial Activity of the Essential Oil from the Peel of Shatian Pummelo (Citrus Grandis Osbeck)

The chemical composition of the essential oil obtained from the peel of Shatian pummelo was analyzed by gas chromatography/mass spectrometry. Twenty-one components were identified. The monoterpenes and sesquiterpene hydrocarbons were the principal compound groups with 96.64% (w/w) of the total oil, among which, limonene was observed dominant (89.96 ± 1.64%), followed by β-myrcene (4.49 ± 0.38%), α-pinene (0.63 ± 0.05%), 3-carene (0.48 ± 0.04%), caryophyllene (0.47 ± 0.04%), and other minor constitutes. Esters, alcohols, ketones, aldehydes, and ether represented 3.15% of the total oil. Results by the disc diffusion method and minimum inhibitory concentration determination method showed that the essential oil contained a wide-spectrum antimicrobial activity against Penicillium chrysogenum ATCC 10106, Bacillus subtilis ATCC 21616, Staphylococcus aureus ATCC 25923, and Escherichia coli ATCC 25922, with their inhibition zones ranging from 8.27 ± 1.07 mm to 20.71 ± 1.50 mm. The MIC values were ranging from 4.69 to 37.50 μL/mL. However, no inhibition effect was observed on the Aspergillus niger ATCC 16888.     

Direct contact membrane inoculation of yeasts and moulds for evaluating preservative efficacy in solid cosmetics

A direct contact membrane inoculation technique for yeasts and moulds was used to evaluate the preservation efficacy and antimicrobial activity of Germall 115 and Germall II in pressed eye shadows. Test organisms on membrane filters were placed in direct contact with cosmetics at room temperature under humid conditions. Growth on membranes was removed daily, or as appropriate, and cultured on potato dextrose agar containing lecithin and Tween 80. Linear regression analysis was used to determine product preservation efficacy. Average D values of 1 and 3 days for Candida albicans American Type Culture Collection (ATCC) 10231 and 17 and 29 days for Aspergillus niger ATCC 16404 were obtained on two eye shadows we prepared (in-house eye shadows) with parabens and either Germall II or Germall 115 as preservatives. A decimal reduction time (D value) of 6–7 days was calculated for the yeast on a commercial eye shadow preserved with parabens and Germall 115. A. niger multiplied on six of seven replicates of this commercial product to attain a nearly 3 log₁₀ increase in 20 days. On one replicate, A. niger showed a 1 log₁₀ increase in the first 10 days, and then decreased linearly (r =— 0.95) to <10 colony-forming units per membrane by day 24. The method used with C. albicans and A. niger was then used with bacteria. The method was sensitive enough to differentiate the antimicrobial activity of the Germall 115 and Germall II against fungi but not against bacteria. The in-house and commercial products were preserved most effectively against the three bacteria tested and least effectively against the mould.     

Supercritical fluid extraction of antioxidant and antimicrobial compounds from Laurus nobilis L. Chemical and functional characterization

Extraction of laurel leaves by using supercritical carbon dioxide was carried out on a supercritical fluid (SF) pilot-scale plant. The extraction pressure and temperature were set to 250 bar and 60°C, respectively, using a 4% of ethanol as modifier. The employed apparatus, owing to a two-stage separation, allowed us to obtain two different fractions (F1 and F2), whose antioxidant and antimicrobial activities were investigated. Two different methods, β-carotene bleaching test and DPPH^• free radical–scavenging assay, were carried out to determine the antioxidant activity. Moreover, antimicrobial activity of laurel fractions was tested against Staphylococcus aureus ATCC 25923, Bacillus subtilis ATCC 6633, Pseudomonas aeruginosa ATCC 10145, Escherichia coli ATCC 11775, Candida albicans ATCC 60193 and Aspergillus niger ATCC 16404. Minimum inhibitory concentration (MIC) and minimal bactericidal and fungicidal concentration (MBC) were obtained. Both fractions showed a similar antioxidant activity, although it was slightly higher for the fraction recovered in separator 2. However, antimicrobial activity against the microorganisms tested was only found when fraction 2 was used. Staphylococcus aureus was the most sensitive microorganism to this fraction, with maximal inhibition zones (25 mm) and the lowest MBC values (1.25 mg/ml), whereas the least susceptible was the fungi Aspergillus niger. In order to determine the compounds responsible for the antimicrobial activity, fraction 2 was analysed by GC–MS; results obtained showed that most of the compounds identified in the supercritical extract have been previously described to show antimicrobial activity; among them, the major compound found in the supercritical extract corresponded to a sesquiterpene lactone of the germacrolide type (6-epi-desacetyllaurenobiolide) previously described in laurel.     

壳聚糖及其金属锌配位络合物的抑菌性能研究

本研究在合成一种壳聚糖锌配合物后,测试了合成的壳聚糖锌、合成原材料壳聚糖的抑菌效果,以及不同pH条件对抑菌效果的影响,并比较了壳聚糖锌和常见的食品防腐剂苯甲酸钠、山梨酸钾对大肠杆菌ATCC8099、金黄色葡萄球菌ATCC6538、枯草杆菌黑色变种ATCC9372、白色念珠菌ATCC10231和黑曲霉ATCC16404的最低抑菌浓度。结果表明:合成的壳聚糖锌与合成前的壳聚糖相比抑菌性能大大提高;合成后的壳聚糖锌对细菌的抑菌效果与常见的食品防腐剂苯甲酸钠、山梨酸钾相当,但对真菌的抑制时效没有苯甲酸钠、山梨酸钾好。同时系统的pH对壳聚糖锌和壳聚糖都有较大的影响,酸性条件下抑菌效果较好。     

Chemical composition and antifungal activity of essential oil from Cicuta virosa L. var. latisecta Celak

The essential oil extracted from the fruits of Cicuta virosa L. var. latisecta Celak was tested in vitro and in vivo against four foodborne fungi, Aspergillus flavus, Aspergillus oryzae, Aspergillus niger, and Alternaria alternata. Forty-five different components accounting for 98.4% of the total oil composition were identified by gas chromatography-mass spectrometry. The major components were γ-terpinene (40.92%), p-cymene (27.93%), and cumin aldehyde (21.20%). Antifungal activity was tested by the poisoned food technique against the four fungi. Minimum inhibitory concentration against the fungi was 5 μL/mL and percentage inhibition of mycelial growth was determined at day 9. The essential oil had a strong inhibitory effect on spore production and germination in all tested fungi proportional to concentration. The oil exhibited noticeable inhibition on dry mycelium weight and synthesis of aflatoxin B? (AFB?) by A. flavus, completely inhibiting AFB(1) production at 4 μL/mL. The effect of the essential oil on inhibition of decay development in cherry tomatoes was tested in vivo by exposing inoculated and control fruit to essential oil vapor at a concentration of 200 μL/mL. Results indicated that the essential oil from C. virosa var. latisecta (CVEO) has potential as a preservative to control food spoilage.     

癸酸聚甘油酯的制备及性能研究

采用静置稳定性、离心稳定性、耐热稳定性等乳化性能指标评价油水比为1:1时癸酸聚甘油酯的乳化性能,并测定其各项指标处于最佳时的最小添加量;采用琼脂扩散法测定指示菌对癸酸聚甘油酯的敏感性;二倍试管稀释法测定其对指示菌的最小抑菌浓度(MIC)。结果表明:癸酸聚甘油酯乳液的类型为水包油(O/W)型,癸酸聚甘油酯添加量为4%时,乳液各项乳化性能指标达到最佳;癸酸聚甘油酯对指示菌均有良好的抑制作用;对黑曲霉、酿酒酵母、金黄色葡萄球菌、大肠杆菌和枯草芽孢杆菌的MIC值分别为0.16mg/mL、0.16mg/mL、0.32mg/mL、2.5mg/mL和2.5mg/mL。     

黄花蒿挥发油的体外抑菌活性研究

采用水蒸气蒸馏法提取黄花蒿挥发油,通过体外抑菌实验,利用纸片法和试管法分别测定其抑菌活性和最低抑菌浓度(MIC)。比较黄花蒿挥发油对12 种常见细菌和真菌的抑菌活性。结果表明：黄花蒿挥发油对12 种实验菌表现出不同的抑制活性。真菌对黄花蒿挥发油具有更强的敏感性。黄花蒿挥发油对真菌的抑制作用比较稳定,由强到弱依次为黑曲霉、啤酒酵母菌、产黄青霉、马青霉、毛霉、青霉和根霉。     

Effects of Ozone Treatment on Botrytis cinerea and Sclerotinia sclerotiorum in Relation to Horticultural Product Quality

ABSTRACT:  Botrytis cinerea  and  Sclerotinia sclerotiorum  are fungal pathogens that cause the decay of many fruits and vegetables. Ozone may be used as an antimicrobial agent to control the decay. The effect of gaseous ozone on spore viability of  B. cinerea  and mycelial growth of  B. cinerea  and  S. sclerotiorum  were investigated. Spore viability of  B. cinerea  was reduced by over 99.5% ( P  < 0.01) and height of the aerial mycelium was reduced from 4.7 mm in the control to less than 1 mm after exposure to 450 or 600 ppb ozone for 48 h at 20 °C. Sporulation of  B. cinerea  was also substantially inhibited by ozone treatments. However, ozone had no significant effect on mycelial growth of  S. sclerotiorum in vitro . Decay and quality parameters including color, chlorophyll fluorescence (CF), and ozone injury were further assessed for various horticultural commodities (apple, grape, highbush blueberry, and carrot) treated with 450 ppb of ozone for 48 h at 20 °C over a period of 12 d. Lesion size and height of the aerial mycelium were significantly reduced by the ozone treatment on carrots inoculated with mycelial agar plugs of  B. cinerea  or  S. sclerotiorum . Lesion size was also reduced on treated apples inoculated with 5 × 10⁶ spores/mL of  B. cinerea , and decay incidence of treated grapes was reduced. The 450 ppb ozone for 48 h treatment had no significant effect on color of carrots and apples or on CF of apples and grapes. Ozone, an environmentally sound antimicrobial agent, inactivates microorganisms through oxidization and residual ozone spontaneously decomposes to nontoxic products. It may be applied to fruits and vegetables to reduce decay and extend shelf life.