Yeast compositions for converting bio-available nitrogen in...

Chemistry: molecular biology and microbiology – Micro-organism – per se ; compositions thereof; proces of... – Fungi

Reexamination Certificate

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C435S254200, C435S255100, C435S255200

Reexamination Certificate

active

06391617

ABSTRACT:

FIELD OF THE INVENTION
The invention relates to the use of yeast cells that are highly efficient in incorporating biologically available nitrogen in a culture medium into their own biomass. These yeasts are useful in waste treatment, and can be obtained by growth in electromagnetic fields with specific frequencies and field strengths.
BACKGROUND OF THE INVENTION
Environmental pollution by urban sewage and industrial waste water has posed a serious health threat to living organisms in the world. Currently, the most common methods for large-scale water treatment include the activated sludge technology and the biomembrane technology. These technologies rely on the innate abilities of myriad natural microorganisms, such as fungi, bacteria and protozoa, to degrade pollutants. However, the compositions of these natural microbial components are difficult to control, affecting the reproducibility and quality of water treatment. Moreover, pathogenic microbes existing in these activated sludge or biomembranes cannot be selectively inhibited, and such microbes usually enter the environment with the treated water, causing “secondary pollution.”
Further, most of the current technologies cannot degrade harmful chemicals such as pesticides, insecticides, and chemical fertilizers. These technologies also cannot alleviate eutrophication, another serious environmental problem around the world. Eutrophication is usually caused by sewage, industrial waste water, fertilizers and the like. It refers to waters (e.g., a lake or pond) rich in mineral and organic nutrients that promote a proliferation of plant life, especially algae, which reduces the dissolved oxygen content or otherwise deteriorates water quality. Eutrophication often results in the extinction of other organisms.
SUMMARY OF THE INVENTION
This invention is based on the discovery that certain yeast cells can be activated by electromagnetic fields having specific frequencies and field strengths to convert biologically available nitrogen, a major environmental pollutant, to intracellular nitrogen (i.e., the yeast cells incorporate biologically available nitrogen in their environs into their own biomass). Compositions comprising these activated yeast cells can therefore be useful for waste treatment, for example, for treatment of sewage, industrial waste water, surface water, drinking water, sediment, soil, garbage, and manure, to reduce the content of available nitrogen in the waste. Waste treatment methods using these compositions are more effective, efficient and economical in preventing eutrophication than conventional methods.
This invention includes a composition comprising a plurality of yeast cells that have been cultured in an alternating electric field having a frequency in the range of about 660 to 680 MHz (e.g, 662, 664, 666, 668, 670, 672, 674, 676, or 678 MHz) and a field strength in the range of about 0.1 to 350 mV/cm (e.g., 140-320 or 120-290 mV/cm). The yeast cells are cultured for a period of time sufficient to substantially increase the capability of said plurality of yeast cells to convert biologically available nitrogen in a culture medium into intracellular nitrogen. In one embodiment, the frequency and/or the field strength of the alternating electric field can be altered within the aforementioned ranges during said period of time. In other words, the yeast cells can be exposed to a series of electromagnetic fields. An exemplary period of time is about 12-420 hours (e.g., 192-304 or 226-412 hours).
This invention also includes a composition comprising a plurality of yeast cells that have been cultured in an alternating electric field having a frequency in the range of about 2160 to 2190 MHz (e.g; 2170 to 2185 MHz) and a field strength in the range of about 0.1 to 350 mV/cm (e.g., 140-320 mV/cm) for a period of time sufficient to substantially increase the capability of said plurality of yeast cells to convert ammonium in a culture medium into intracellular nitrogen.
Yeast cells that can be included in this composition can all be obtained from the China General Microbiological Culture Collection Center (“CGMCC”), a depository recognized under the Budapest Treaty (China Committee for Culture Collection of Microorganisms, Institute of Microbiology, Chinese Academy of Sciences, Haidian, P.O. BOX 2714, Beijing, 100080, China). Useful yeast species include, but are not limited to,
Saccharomyces cerevisiae, Candida tropicalis
and
Geotrichum candidum
. For instance, the yeast cells can be of the strain
Saccharomyces cerevisiae
AS2.196, AS2.336, AS2.400, AS2.416, AS2.423, or AS2.982
; Saccharomyces willianus
Saccardo AS2.152 or AS2.614
; Candida tropicalis
AS2.1387; or
Geotrichum candidum
AS2.498.
This invention also embraces a composition comprising a plurality of yeast cells, wherein said plurality of yeast cells have been activated such that they have a substantially increased capability to convert bio-available nitrogen in a culture medium into intracellular nitrogen as compared to unactivated yeast cells.
As used herein, “biologically available,” “bio-available,” or “biologically assimilable” nitrogen refers to nitrogen that is readily available, useable, or assimilable by living organisms for survival and/or growth. Exemplary bio-available or bio-assimilable nitrogen includes, but is not limited to, NH
4
+
, N
3

and NO
2

, other water-soluble inorganic nitrogen-containing compounds, and organic nitrogen-containing compounds.
A “substantial increase” means an increase of more than 10 (e.g., 10
2
, 10
3
, 10
4
, 10
5
, or 10
6
) fold.
A “culture medium” refers to a medium used in a laboratory for selecting and growing a given yeast strain, or to liquid or solid waste in need of treatment.
Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Exemplary methods and materials are described below, although methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present invention. All publications and other references mentioned herein are incorporated by reference in their entirety. In case of conflict, the present specification, including definitions, will control. The materials, methods, and examples are illustrative only and not intended to be limiting.
Other features and advantages of the invention will be apparent from the following detailed description, and from the claims.


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K. Asami et al., “Real-Time Monitoring of Yeast Cell Division by Dielectric Spectroscopy”,Biophysical Journal, 76, pp. 3345-3348 (1999).
E.K. Balcer-Kubiczek et al., “Expression Analysis of Human HL60 Cells Exposed to 60 Hz Square-or Sine-Wave Magnetic Fields”,Radiation Research, 153, pp. 670-678 (2000).
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A.M. Gonzalez et al., “Effects of an Electric Field of Si

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