Fermentation,Structure Characterization And Properties Of Pigment From Pseudomonas Stutzeri ZH-1

Microbial pigments were extensively used in many fields such as dyeing,weaving,food,medicine,cosmetics due to the characteristics of safety,nutrition and out of control the environment.In this study,the pigment was investigated from Pseudomonas stutzeri ZH-1which was first isolated and identified from the sludge of Fenhe River,Linfen,Shanxi Province.The thesis consists of four sections:the choice of cultural conditions,the choice of extraction conditions,the identification of structure,the study of physical and chemical properties.The specific contents and results were display as following:In this experiment,six culture conditions including time,temperature,initial p H,inoculation amount,liquid loading amount and medium concentration were studied as single factor experiment.Response surface optimization was investigated by four conditions(time,temperature,initial p H and liquid loading).The results were obtained as follows:time was 52h,temperature was 37?,initial p H was 7 and liquid loading was 96 m L in 250 m L Erlenmeyer flask.In this study,the secrete pathway of pigment was analyzed and the single factor experiment of extract condition of pigment was studied.The results showed that the pigment from P.stutzeri ZH-1 was secreted by extracellular and can be extracted by 40%ethanol.The extract conditions were as follows:time was 12 hours,temperature was 60?,p H was12,and the ratio of pigment solution and 40%ethanol was 1:5.The results showed that the pigment value of P.stutzeri ZH-1 was 129.58±8.75,and compared to the previous unoptimized,the yield was increased by 11.03±0.13%.The structure of pigment was analyzed by color reaction,ultraviolet(UV)spectrum and infrared(IR)spectrum.Color reaction indicate the pigment from P.stutzeri ZH-1 possess quinone structure.UV has a maximum absorption peak at 252 nm with a slope of-0.0015,and the typical absorption peak of eumelanin of the DOPA pathway is shown in infrared(IR)spectrum,indicating pigment from P.stutzeri ZH-1 is eumelanin of the DOPA pathway.Stability tests showed the pigments from P.stutzeri ZH-1 has a relatively good stability at p H>5 and thermal stability at 30?100?.The pigments all retained a residual rate above90%in many solutions such as metal ion(Mn²⁺,Na⁺,Al³⁺and Cu²⁺),H₂O₂,Na₂S₂O₈and Na SO₃.Moreover,antioxidant activity test displayed that the nature pigments has the strong scavenging activities against three free radicals including DPPH,ABTS and hydroxyl,and the highest scavenging rates at 0.5 g/L concentrations of pigments were up to 20.58±1.4%,19.23±2.8%,37.69±1.8%,respectively.The significance analysis showed that there were significant differences(p<0.05)under the conditions of p H=2?12,temperature30?100?,metal ions Ca²⁺,Mn²⁺,Na⁺,Al³⁺,Cu²⁺and Zn²⁺in the residual amount of pigment.In conclusion,the pigments from P.stutzeri ZH-1 has obvious advantages in the utilization of microbial pigments.Our work provided a new idea for the utilization of P.stutzeri ZH-1 and a good microbial source for melanin.