EXAMPLE I From 201.4 liters of Chaetoceros algae culture having a cell density ranging from ***** cells per milliliter of water, 22.5 grams of ground wet algae-paste was obtained. The wet paste was taken into 600 milliliters of water and centrifuged at 10,000 RPMs for approximately 20 minutes. The supernatant liquid was then loaded onto an Amberlite XAD-2 column (sigma, 30x7 cm) and eluted with 2.0 liters of the following gradient: (1) water, (2) 10% aqueous MeOH, and (3) 30% aqueous MeOH, (4) 50% aqueous MeOH, (5) 70% aqueous MeOH, (6) 90% aqueous MeOH, (7) MeOH and (8) CH2Cl2. These fractions were tabulated in Table I1 below and show that identified fractions were placed in petri dish cultures of methicillin resistant ~~hvlococcus aureus. As can be seen from Table 11, activity against ~vlococcu~ aureus was observed in fractions 5-7 with fraction 6 yielding the highest activity. Activity is indicated by the dark portion around the white circle of antibiotic which was introduced into the petri dish. The observations as to activity were taken after 12 hours.
Activity is indicated in fractions 5 through 8, with the highest activity being prese in fraction 6. The activity of the various fractions against Vibrio vulnificus can be seen from the table. Activity was strongest in fractions 5 through 8, with the highest activity occurring with fraction 6. As can be seen from Table 11, the most active fractions against each of the organisms listed are fractions 5 to 8, with fraction 6 obtained from a 90% by volume aqueous methanol eluant indicating the highest activity. All of the fractions were concentrated under reduced pressure to provide the crudematerial. As a result of activities observed in fractions 5 through 8, these fractions were combined and loaded onto a YMC-ODs-120A (reverse-phase silica gel, C18; 20 X 2.5 cm)eluting with the following gradient, the fraction number in the table being indicated with the same numbers: (1.1) 20% aqueous MeOH, (2.1) 40% aqueous MeOH, (3.1) 60% aqueous MeOH, (4.1) 80% aqueous MeOH. and (5.1) MeOH. Five fractions of 250 to 30milliliters were collected and tested for activities using previously prepared cultures of methicillin resistant ~taphvlococcus mreus, vancomycin resistant ~nterococcus and Vibrio vulnificus, respectively. The results indicate the efficacy of five fractions against each of the indicated pathogens. It is interesting to note that although fraction 2.1 did not show activity against Vibrio vulnificus 32 after 12 hours, shows fraction 2.1 is strongly effective against Vibrio vulnificus 32 when the experiment proceeds up to 24 hours. The results are tabulated for convenience in Table I11 below.
As indicated in Table 111, the active fractions were 2.1, 3.1 and 4.1. Fractions 2.1 and 3.1 were combined because they exhibited very similar UV spectra. Fraction 4.1 was not combined with other fractions because its UV spectra indicated that it was different from the other fkactions. Further separation of the combined fractions of 2.1 and 3.1 by HPLC reverse-phase silica gel (C8,250 x 10 mm, 10微米; 40% MeOH: 50% MeOH: 10% MeCN: H2O: MeOH, at a flow rate of 2.0 milliliter per minute and UV detection at 220 nm) provided five additional major fractions 1.2-5.2. Fractions 2.2, 3.2,4.2 and 5.2 appear to be effective against methiciilin resistant Staphvlococcus aureus, while fractions 4.2 and 5.2 are effective against vancomycin resistant enterococcus and fractions 2.2 and 3.2 are effective against Vibrio vulnificus 32. The results of this are tabulated in Table IV below.
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