HK1060031A - Dietary supplements for regulating male hormone - Google Patents

Description

Dietary supplement for regulating androgens

Technical Field

The present invention relates to compositions that can ameliorate or prevent androgen deficiency and can be used as dietary supplements. These compositions contain yeast cells that can be obtained by growth in an electromagnetic field of a particular frequency and field strength.

Background

Testosterone levels in males decline with age. Testosterone deficiency is believed to be associated with hip fracture and reduced bone mass in the elderly. Testosterone replacement therapy has been used to improve the levels of this hormone.

In various treatments, some botanical preparations including ginseng, ginkgo biloba, yohimbine (pausinylia yohimbe) and a south american plant-force wood (Ptychopetalum olacoides) have been used to restore erectile function. The precursors of nitric oxide, L-arginine and androstenedione, also have utility in male sexual function. Males receiving testosterone had significant improvements in bioavailable testosterone concentration, hematocrit, right hand muscle strength, and osteocalcin concentration. Treatment with the estrogen antagonist tamoxifen has been shown to increase testosterone serum levels in mammals and oligospermic men. In addition, the antiestrogen clomiphene is used to treat decreased libido, hypogonadotropic hypogonadism and infertility associated therewith.

Summary of The Invention

The present invention is based on the discovery that: certain yeast cells can be activated by electromagnetic fields having specific frequencies and field strengths to increase the serum levels of testosterone. Thus, compositions containing these activated yeast cells can be used to treat androgen deficiency or related diseases in a subject (e.g., a human).

The 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 10500-12000MHz (e.g., 11210-11250MHz) and a field strength in the range of about 100-600mV/cm (e.g., 150-520 mV/cm). Culturing the plurality of yeast cells in the alternating electric field for a time sufficient to activate the plurality of yeast cells to treat androgen deficiency or a related disorder in the subject. For example, when ingested, the cultured yeast cells can stimulate prostate, seminal vesicle, and glandular growth (e.g., at least 10%, such as 20%, 30%, 40%, 50%, one-fold, five-fold, ten-fold, and 15-fold) of the mammal and increase testosterone secretion (e.g., at least 10%, such as 20% and 200%).

In one embodiment, the frequency and/or field strength of the alternating electric field may vary within the ranges described above during the time period. In other words, the yeast cells are exposed to a series of electromagnetic fields. Typical time periods are 50-200 hours (e.g., 60-145 hours).

The invention also includes a composition comprising a plurality of yeast cells that have been cultured under acidic conditions in an alternating electric field having a frequency in the range of 11000-12000MHz (e.g., 11200-11250MHz) and a field strength in the range of 200-600mV/cm (e.g., 240-520 mV/cm). In one embodiment, the yeast cells are exposed to a series of electromagnetic fields. Typical time periods are about 50-100 hours (e.g., 54-84 hours).

Yeast cells that may be included in the composition are all available from the China General Microbiological Culture Collection Center ("CGMCC"), a Collection Center approved by the Budapest treaty (China Committee for Culture Collection of microorganisms, institute of microbiology, China, Hai lake, 2714 letter box, Beijing, 100080, China). Yeast species that may be used include, but are not limited to, those commonly used in the food and pharmaceutical industries, such as Saccharomyces cerevisiae (Saccharomyces cerevisiae), Saccharomyces carlsbergensis (Saccharomyces carlsbergensis), Saccharomyces chevalieri (Saccharomyces chevalieri), Saccharomyces delbrueckii (Saccharomyces delbrueckii), Saccharomyces exis (Saccharomyces exiguus), Saccharomyces fermentans (Saccharomyces fermentification), Saccharomyces logarii (Saccharomyces lowarii), Saccharomyces lowarii (Saccharomyces lowaricus), Saccharomyces cerevisiae (Saccharomyces meldonii), Saccharomyces cerevisiae (Saccharomyces oviformis), Saccharomyces roseus (Saccharomyces cerevisiae), Saccharomyces cerevisiae (Saccharomyces cerevisiae, trichosporon capitatum (Trichosporon capitatum), Trichosporon cutaneum, Saccharomyces cerevisiae (Wickerhamia fluocinosens), Candida xylonatans (Candida arboreum), Candida krusei (Candida krusei), Candida lambada (Candida lambda), Candida lipolytica (Candida glabrata), Candida parapsilosis (Candida parapsilosis), Candida ferrooxidans (Candida pulcherrima), Candida rugosa (Candida rugosa), Candida tropicalis (Candida tropicalis), Candida utilis (Candida utilis), Creothechrocrelatively asbyii, Geotrichum (Geotrichum), Hansenula anomala (Hansenula polymorpha), Saccharomyces arabicum (Hansenula arabica), Saccharomyces cerevisiae (Hansenula polymorpha), Pichia pastoris (Hansenula, Hansenula polymorpha), Pichia pastoris (Hansenula polymorpha), Candida albicans (Hansenula, Hansenula polymorpha, Hansenula (Hansenula, Hansenula polymorpha, Hansenula polymorpha membrane, Hansenula (Hansenula polymorpha) and Candida solanum), rhodosporidium toruloides (Rhodosporidium toruloides), Rhodotorula glutinis (Rhodotorula glutinis), Rhodotorula microzyme (Rhodotorula minuta), Rhodotorula rubra (Rhodotorula ruba), Rhodotorula aurantiaca (Rhodotorula aurantiaca), Saccharomyces lucwigii (Saccharomyces lucwigii), and Saccharomyces cerevisiae (Saccharomyces sinense). For example, the yeast cell can be a strain of Saccharomyces cerevisiae Hanson (Saccharomyces cerevisiae Hanson) AS2.375, AS2.501, AS2.502, AS2.503, AS2.504, AS2.535, AS2.558, AS2.560, AS2.561, AS2.562, or IFFI 1048; or Saccharomyces carlsbergensis Hansen AS2.420 or AS 2.444. Other yeast species that may be used are described in table 1.

The invention also includes a composition comprising a plurality of yeast cells, wherein the plurality of yeast cells have been activated to treat androgen deficiency or a related disease in a subject. The invention also includes methods of making these compositions.

Unless defined otherwise, 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, and methods and materials similar or equivalent to those described herein can also be used in the testing or practice of the present invention. All publications and other documents 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 should not be construed as limiting the invention. Throughout the description and claims of this specification, the word "comprise", or variations such as "comprises" or "comprising", will be understood to imply the inclusion of a stated integer or group of integers but not the exclusion of any other integer or group of integers.

Other features and advantages of the invention will be apparent from the following detailed description, and from the claims.

Drawings

FIG. 1 is a schematic diagram showing a typical apparatus for activating yeast cells using an electromagnetic field. 1: a yeast culture; 2: a container; 3: a power source.

FIG. 2 is a schematic diagram showing a typical apparatus for making the yeast composition of the invention. The device comprises a signal generator and interconnected containers 1, 2 and 3.

Detailed Description

The present invention is based on the discovery that: certain yeast strains can be activated by electromagnetic fields ("EMF") of specific frequencies and field strengths to produce substances that are highly effective in increasing testosterone serum levels. Compositions containing these activated yeast cells are useful for treating androgen deficiency or related diseases. The yeast composition containing activated yeast cells can be used as a dietary supplement, for example in the form of a health drink or a pill.

Since the activated yeast cells contained in these yeast compositions have been cultured under acidic conditions at pH2.5-4.2, these cells are stable in the stomach and can be continued to travel to the small intestine. Once in the small intestine, these yeast cells are disrupted by various digestive enzymes, so that the active substance is released and can be easily absorbed.

The inventors are not bound by any theory or mechanism, and believe that: EMF activates or enhances the expression of a gene or set of genes in yeast cells, or alters the conformation and/or activity of certain cellular components (e.g., DNA, RNA, enzymes/proteins) in yeast cells, thereby causing the production of substances that can ameliorate or prevent androgen deficiency. I. Yeast strains for use in the present invention

Types of yeasts used in the present invention include, but are not limited to, Saccharomyces cerevisiae, Saccharomyces carlsbergensis, Saccharomyces chevalieri, Delbert, Saccharomyces oligosporus, Zymomyces, Saccharomyces luggensis, Saccharomyces melleus, Saccharomyces ovaginosum, Saccharomyces rouxii, Schizosaccharomyces pombe, Saccharomyces Sporoselis, Torulopsis albicans, Pseudosaccharomyces famensis, Torulopsis globulosa, Torulopsis vulgaris, Saccharomyces bayanus, Saccharomyces cerevisiae, Trichosporoides, Saccharomyces cerevisiae, Candida krusei, Candida lambdakii, Candida lamblia, Candida lipolytica, Candida parapsilosis, Candida rugosa, Candida tropicalis, Candida utilis, Creotecium ashbyii, Geotrichum, Hansenula anomyces anomala, Hansenula arabinita, Hansenula jackinthica, Hansenula pseudostellata, schneggii Hansenula, Hansenula polymorpha, Klebsiella citrullina, Saccharomyces cerevisiae, Pichia farinosa, Pichia membranaefaciens, Rhodosporidium toruloides, Rhodotorula rubra, Rhodotorula aurantiaca, Rhodotorula lodes, and Saccharomyces mellea.

Yeast strains for use in the present invention may be obtained from laboratory cultures or from publicly available culture collections, such as CGMCC and American type culture Collection, 10801 University Boulevard, Mass., VA 20110-. Non-limiting examples of strains that may be used (including the accession number of CGMCC) are Saccharomyces cerevisiae Hanseng AS2.375, AS2.501, AS2.502, AS2.503, AS2.504, AS2.535, AS2.558, AS2.560, AS2.561, AS2.562, and IFFI 1048; and karyomyces hansenii AS2.420 and AS 2.444. Other useful yeast species are described in table 1.

The preparation of the yeast composition of the invention is not limited to starting from a pure yeast strain, although it is preferred to do so. The yeast compositions of the invention can be produced by culturing a mixture of yeast cells of different species or strains.

Table 1 exemplary Yeast strains

Hanseng Saccharomyces cerevisiae (Saccharomyces cerevisiae Hansen)
					ACCC2034	ACCC2035	ACCC2036	ACCC2037	ACCC2038
ACCC2039	ACCC2040	ACCC2041	ACCC2042	AS2.1

AS2.4	AS2.11	AS2.14	AS2.16	AS2.56
					AS2.69	AS2.70	AS2.93	AS2.98	AS2.101
AS2.109	AS2.110	AS2.112	AS2.139	AS2.173
					AS2.174	AS2.182	AS2.196	AS2.242	AS2.336
AS2.346	AS2.369	AS2.374	AS2.375	AS2.379
					AS2.380	AS2.382	AS2.390	AS2.393	AS2.395
AS2.396	AS2.397	AS2.398	AS2.399	AS2.400
					AS2.406	AS2.408	AS2.409	AS2.413	AS2.414
AS2.415	AS2.416	AS2.422	AS2.423	AS2.430
					AS2.431	AS2.432	AS2.451	AS2.452	AS2.453
AS2.458	AS2.460	AS2.463	AS2.467	AS2.486
					AS2.501	AS2.502	AS2.503	AS2.504	AS2.516
AS2.535	AS2.536	AS2.558	AS2.560	AS2.561
					AS2.562	AS2.576	AS2.593	AS2.594	AS2.614
AS2.620	AS2.628	AS2.631	AS2.666	AS2.982
					AS2.1190	AS2.1364	AS2.1396	IFFI1001	IFFI1002
IFFI1005	IFFI1006	IFFI1008	IFFI1009	IFFI1010
					IFFI1012	IFFI1021	IFFI1027	IFFI1037	IFFI1042
IFFI1043	IFFI1045	IFFI1048	IFFI1049	IFFI1050
					IFFI1052	IFFI1059	IFFI1060	IFFI1062	IFFI1063
IFFI1202	IFFI1203	IFFI1206	IFFI1209	IFFI1210
					IFFI1211	IFFI1212	IFFI1213	IFFI1214	IFFI1215
IFFI1220	IFFI1221	IFFI1224	IFFI1247	IFFI1248
					IFFI1251	IFFI1270	IFFI1277	IFFI1287	IFFI1289
IFFI1290	IFFI1291	IFFI1292	IFFI1293	IFFI1297
					IFFI1300	IFFI1301	IFFI1302	IFFI1307	IFFI1308
IFFI1309	IFFI1310	IFFI1311	IFFI1331	IFFI1335
					IFFI1336	IFFI1337	IFFI1338	IFFI1339	IFFI1340
IFFI1345	IFFI1348	IFFI1396	IFFI1397	IFFI1399

IFFI1411	IFFI1413	IFFI1441	IFFI1443
						Egg-shaped variant of Saccharomyces cerevisiae Hansen (Hansen) Dekker (Saccharomyces cerevisiae Hansen Var. Ellipsoides (Hansen) and the like)
ACCC2043	AS2.2	AS2.3	AS2.8		AS2.53
						AS2.163	AS2.168	AS2.483	AS2.541		AS2.559
AS2.606	AS2.607	AS2.611	AS2.612
						Schlerya verrucosa yeast (Saccharomyces chevalieri Guillierimond)
AS2.131	AS2.213
						Deschomyces (Saccharomyces delbrueckii)
AS2.285
						Saccharomyces delbrueckii Lindner ver monogolicus (Saito) Lodder et van Rij
AS2.209	AS2.1157
						Hansenula oligospora (Saccharomyces exiguous Hansen)
AS2.349	AS2.1158
						Zymobacter (Saito) Lodder et van Rij (Saccharomyces transferati Lodder et van Rij)
AS2.286	AS2.343
						The Roger yeast van laer et Denamur ex Jorgensen (Saccha)romyces logos van laer et Denamur ex Jorgensen)
AS2.156	AS2.327	AS2.335
						Honey yeast (Fabian et Quinet) Lodder et kreger van Rij (Saccharomyces mellis (Fabian et Quinet) Lodder et kreger van Rij)
AS2.195
						Honey small oval yeast Osterwalder (Saccharomyces mellis Microllipsoides Osterwalder)

AS2.699
					Oval yeast Osteralder (Saccharomyces oviformis Osteralder)
AS2.100
					Rosemary yeast (Guilliermond) Lodder et Kreger van Rij (Saccharomyces rosei (Guilliermond) Lodder et Kreger van Rij)
AS2.287
					Saccharomyces rouxii (Saccharomyces rouxii) Bortroux
AS2.178	AS2.180	AS2.370	AS2.371
					Sake yeast Yabe (Saccharomyces sake Yabe)
ACCC2045
					Candida Mudanae (Candida arborea)
AS2.566
					Candida lambertian (Lindner et Genoud) van. uden et Buckley (Candida lambda (Lindner et Genoud) van. uden et Buckley)
AS2.1182
					Candida Krusei (Castellani) Berkhout (Candida krusei (Castellani) Berkhout)
AS2.1045
					Candida lipolytica (Harrison) Diddens et Lodder (Candida lipolytica (Harrison) Diddens et Lodder)
AS2.1207	AS2.1216	AS2.1220	AS2.1379	AS2.1398
					AS2.1399	AS2.1400
Candida parapsilosis (Ashford) Langeron et Talcum Var.Intermedia Van Rij et Verona (Candida parapsilosis (Ashford) Langeron et Talcum Var Intermedia Rij et Verona)
					AS2.491

Candida parapsilosis (Ashford) Langeron et Talci (Candida parapsilosis (Ashford) Langeron et Talci)
					AS2.590
Candida Ferrugosa (Lindner) Windisch (Candida pulcherrima (Lindner) Windisch)
					AS2.492
Candida rugosa (Anderson) Diddens et Lodder (Candida rugousa (Anderson) Diddens et Lodder)
					AS2.511	AS2.1367	AS2.1369	AS2.1372	AS2.1373
AS2.1377	AS2.1378	AS2.1384
					Candida tropicalis (Castellani) Berkhout (Candida tropicalis (Castellani) Berkhout)
ACCC2004	ACCC2005	ACCC2006	AS2.164	AS2.402
					AS2.564	AS2.565	AS2.567	AS2.568	AS2.617
AS2.637	AS2.1387	AS2.1397
					Candida utilis Henneberg Lodder et Kreger Van Rij (Candida utilis Henneberg Lodder et Kreger Van Rij)
AS2.120	AS2.281	AS2.1180
					Creothechium ashbyii (Guillermond) Routein (Erothechium ashbyi) (Creothechium ashbyii (Guillermond) Routein (Eremothecium ashbyii Guillermond)
AS2.482	AS2.482	AS2.1197
					Geotrichum candidum (Geotrichum candidum Link)
ACCC2016	AS2.361	AS2.498	AS2.616	AS2.1035
					AS2.1062	AS2.1080	AS2.1132	AS2.1175	AS2.1183
Hansenula anomala (Hansen) H et P sydow (Hansenula anomala (Hansen) H et P sydow)
					ACCC2018	AS2.294	AS2.295	AS2.296	AS2.297

AS2.298	AS2.299	AS2.300	AS2.302	AS2.338
					AS2.339	AS2.340	AS2.341	AS2.470	AS2.592
AS2.641	AS2.642	AS2.782	AS2.635	AS2.794
					Hansenula arabinitol producing Hansenula fang (Hansenula arabitol fang)
AS2.887
					Hansenula jidingensis (A.et R Sartorey Weill et Meyer) Wickerhan (Hansenula jadinii (A.et R Sartorey Weill et Meyer) Wickerhan)
ACCC2019
					Hansenula terrestris (Klocker) H et P sydow (Hansenula saturnus (Klocker) H et P sydow)
ACCC2020
					Schnegii Hansenula (Weber) Dekker (Hansenula schneggii (Weber) Dekker)
AS2.304
					Hansenula polymorpha Bedford (Hansenula subapelliculosa Bedford)
AS2.740	AS2.760	AS2.761	AS2.770	AS2.783
					AS2.790	AS2.798	AS2.866
Klebsiella yeast (stress em. Klocker) Janke in the form of lemon
					ACCC2022	ACC2023	AS2.197	AS2.496	AS2.714
ACCC2021	AS2.711
					Lipid yeast Lodder et van Rij (Lipomyces starkeyi Lodder et van Rij)
AS2.1390	ACCC2024
					Pichia farinosa (Lindner) Hansen (Pichia farinosa (Lindner) Hansen)
ACCC2025	ACCC2026	AS2.86	AS2.87	AS2.705
					AS2.803

Pichia membranaefaciens Hansen (Pichia membranaefaciens Hansen)
					ACCC2027	AS2.89	AS2.661	AS2.1039
Rhodosporidium toruloides Banno (Rhodosporidium toruloides Banno)
					ACCC2028
Rhodotorula (Fresenius) Harrison (Rhodosporidium gluttinis (Fresenius) Harrison)
					AS2.2029	AS2.280	ACCC2030	AS2.102	AS2.107
AS2.278	AS2.499	AS2.694	AS2.703	AS2.704
					AS2.1146
Rhodotorula parviflora (Saito) Harrison (Rhodotorula minuta (Saito) Harrison)
					AS2.277
Rhodotorula rubra (Demmer) Lodder (Rhodotorula rubar (Demmer) Lodder)
					AS2.21	AS2.22	AS2.103	AS2.105	AS2.108
AS2.140	AS2.166	AS2.167	AS2.272	AS2.279
					AS2.282	ACCC2031
Rhodotorula aurantiaca (Saito) Lodder (Rhodotorula aurantiaca (Saito) Lodder)
					AS2.102	AS2.107	AS2.278	AS2.499	AS2.694
AS2.703	AS2.704	AS2.1146
					Calycor hansenii (Saccharomyces carlsbergensis Hansen)
AS2.113	ACCC2032	ACCC2033	AS2.312	AS2.116
					AS2.118	AS2.121	AS2.132	AS2.162	AS2.189
AS2.200	AS2.216	AS2.265	AS2.377	AS2.417
					AS2.420	AS2.440	AS2.441	AS2.443	AS2.444
AS2.459	AS2.595	AS2.605	AS2.638	AS2.742

AS2.745	AS2.748	AS2.1042
					Grape juice yeast Beijer (Saccharomyces uvarum Beijer)
IFFI1023	IFFI1032	IFFI1036	IFFI1044	IFFI1072
					IFFI1205	IFFI1207
Saccharomyces cerevisiae (Saccharomyces willianus Sacchardono)
					AS2.5	AS2.7	AS2.119	AS2.152	AS2.293
AS2.381	AS2.392	AS2.434	AS2.614	AS2.1189
					Yeast sp (Saccharomyces sp.)
AS2.311
					Lvde yeast Hansen (Saccharomyces ludwigii Hansen)
ACCC2044	AS2.243	AS2.508
					Yeast of the Sinenses class Yue (Saccharomyces Sinenses Yue)
AS2.1395
					Schizosaccharomyces octasporum Beijerinck (Schizosaccharomyces occidentalis Beijerinck)
ACCC2046	AS2.1148
					Schizosaccharomyces pombe Lindner (Schizosaccharomyces pombe Lindner)
ACCC2047	ACCC2048	AS2.214	AS2.248	AS2.249
					AS2.255	AS2.257	AS2.259	AS2.260	AS2.274
AS2.994	AS2.1043	AS2.1149	AS2.1178	IFFI1056
					Sporobolomyces Kluyver et van Niel (spoorobolomyces roseus Kluyver et van Niel)
ACCC2049	ACCC2050	AS2.19	AS2.962	AS2.1036
					ACCC2051	AS2.261	AS2.262

Torulopsis bailii (Saito) Lodder (Torulopsis candidii (Saito) Lodder)
					AS2.270	ACCC2052
Yeast analogue Famta (Harrison) Lodder et van Rij (Torulsis Famta (Harrison) Lodder et van Rij)
					ACCC2053	AS2.685
Torulopsis globisporus (Olson et Hammer) Lodder et van Rij (Torulopsis globosa (Olson et Hammer) Lodder et van Rij)
					ACCC2054	AS2.202
Ordinary Torulopsis coccinella Lodder et Kreger van Rij (Torulsis inconetc. Lodder et Kreger van Rij)
					AS2.75
Belgium toruloides Lodder et Kreger van Rij (Lodder et Kreger van Rij)
					ACCC2056	AS2.1193
Trichosporon capitatum Diddens et Lodder (Trichosporon capittatum Diddens et Lodder)
					ACCC2056	AS2.1385
Trichosporon oryzae (de Berm et al.) Ota (Trichosporon cutaneum (de Berm et al.) Ota)
					ACCC2057	AS2.25	AS2.570	AS2.571	AS2.1374
Yeast Weeker (Soneda) Soneda (Wickerhamia fluoscescens (Soneda)
					ACCC2058	AS2.1388

Application of electric field

The electromagnetic field used in the present invention can be generated and applied by various methods known in the art. The EMF may be generated, for example, by applying an alternating electric field or an oscillating magnetic field.

An alternating electric field may be applied to the cell culture by direct contact of the electrodes with the culture medium, or by electromagnetic induction. See, for example, fig. 1. Using the method of contacting the electrodes with the medium, a relatively high electric field can be generated in the medium. Care must be taken to prevent electrolysis at the electrodes from introducing non-target ions into the medium, and care must be taken to prevent contact resistance, bubbles, or other characteristics of electrolysis from dropping the electric field level below the desired level. The electrodes should be matched to their environment, for example, Ag-AgCl electrodes are used in solutions rich in chloride ions, and the electrodes should be operated at as low a voltage as possible. For a Review, see Effects of EMF on Molecules and cells, International Review of Biology, A Survey of Cell Biology, Vol.158, Academic Press, 1995, Goodman et al.

The EMF used in the present invention may also be generated by applying an oscillating magnetic field. An oscillating magnetic field may be generated by an oscillating current through a helmholtz coil. This oscillating magnetic field in turn induces an electric field.

The EMF used in the present invention has a frequency range of about 10500MHz to 12000MHz (e.g., 11210-11250 MHz). Typical frequencies are 11217, 11224, 11231, 11237, and 11244 MHz. The electric field strength used in the present invention is in the range of about 100-600mV/cm (e.g., 150-230, 190-250, 250-270, 430-450, or 480-520 mV/cm). Typical field strengths are 177, 221, 224, 236, 241, 244, 260, 442, 507, and 512 mV/cm.

When a series of EMFs are applied to a yeast culture, the yeast culture can be maintained in the same container, using the same set of EMF generators and transmitters to vary the frequency and/or field strength. Each EMF in the series may have a different frequency or a different field strength; or different frequencies and different field strengths. These frequencies and field strengths are preferably in the ranges mentioned above. Although any number of EMFs can be used in a series, it is preferred that the yeast culture is exposed to a total of 2, 3, 4, 5, 6, 7, 8, 9 or 10 EMFs in a series.

Although yeast cells can be activated even after several hours in culture in the presence of EMF, it is preferred that the activated yeast cells be allowed to replicate and grow in the presence of EMF for a total of 50-200 hours (e.g., 80-145 hours).

FIG. 1 depicts an exemplary apparatus for generating an alternating electric field. The electric field of the desired frequency and intensity can be generated by an AC source (3), the AC source (3) being capable of generating an alternating electric field, preferably of sinusoidal waveform, in the frequency range 10-20,000 MHz. A signal generator capable of generating signals with a narrow frequency range may also be used. A signal amplifier may be used to increase the output if desired. The culture vessel (2) may be made of a non-conductive material such as glass, plastic or ceramic. The cable connecting the culture vessel (2) and the signal generator (3) is preferably a high frequency coaxial cable, and the transmission frequency thereof is at least 20 Ghz. In one embodiment, the transmission frequency is 30 Ghz.

The alternating electric field can be applied to the medium by a variety of methods including placing the yeast culture (1) in close proximity to a signal emitter, such as a wire or tube capable of transmitting EMFs. The wire or tube may be made of red copper and may be placed into the container (2) to a depth of 3-30 cm. For example, if the depth of the fluid in the vessel (2) is 15-20cm, 20-30cm, 30-50cm, 50-70cm, 70-100cm, 100-150cm or 150-200cm, the length of the metal wire from the bottom of the vessel (2) may be 3-5cm, 5-7cm, 7-10cm, 10-15cm, 15-20cm, 20-30cm and 25-30cm, respectively. The number of electrode wires used depends on the volume of the culture and the diameter of the wires. The number of wires/tubes used may be 1-10 (e.g., 2-3). Although not required, it is suggested that for a 10L volume of culture, a wire/tube with a diameter of 0.5-2mm should be used. For cultures of volume 10-100L, a wire/tube of diameter 3-5mm may be used. For a culture volume of 100-1000L, a metal wire/tube with a diameter of 6-15mm may be used. For cultures with a volume greater than 1000L, a wire/tube with a diameter of 20-25mm may be used.

In one embodiment, the electric field is applied by electrodes immersed in the culture (1). In this embodiment, one electrode may be a metal plate placed at the bottom of the container (2) and the other electrode may contain a number of electrode wires distributed evenly in the culture (1), so that an even distribution of the electric field energy may be achieved. The number of electrode wires used depends on the volume of the culture and the diameter of the metal wires. III. culture Medium

The culture medium used in the present invention contains a nutrient source that can be taken up by the yeast cells. The complex carbonaceous material may be used in a suitable form, such as sugars (e.g., sucrose, glucose, fructose, dextrose, xylose, cellulose, starch, etc.) and coal as a carbon source for the yeast cells. The exact amount of carbon source may be adjusted depending on the other components of the medium. Generally, the amount of carbonaceous material ranges from about 0.1% to 10%, preferably from about 0.1% to 5%, and most preferably about 2% by weight of the medium. These carbon sources may be used alone or in combination. It is likewise possible to add the amino acid-containing substances in suitable form (e.g.beef extract and peptone), alone or in combination. Generally, the amount of amino acid-containing substance varies from about 0.1% to about 1%, preferably from about 0.1% to about 0.5% by weight of the medium. The inorganic salts which may be added to the medium are conventional salts capable of producing sodium, potassium, calcium, sulfate, carbonate and the like ions. A non-limiting example of a nutritive inorganic salt is (NH)₄)₂HPO₄，CaCO₃，KH₂PO₄，K₂HPO₄，MgSO₄NaCl, and CaSO₄. Electromagnetic activation of yeast cells

The yeast cells of the invention are cultured in a suitable medium under sterile conditions at 20 ℃ to 38 ℃, preferably 28 ℃ to 32 ℃ (e.g., 30 ℃), in the alternating electric field or series of alternating electric fields described above for a sufficient period of time, e.g., 80 to 200 hours (e.g., 103 hours and 145 hours).

An exemplary medium is prepared by mixing 1000ml of distilled water with 20g of sucrose, 50. mu.g of vitamin B₆40 μ g of vitamin B₁₂0.20g KH₂PO₄0.2g of MgSO₄·7H₂O, 0.25g NaCl, 0.1g CaSO₄·2H₂O, 3.0g CaCO₃·5H₂O, and 2.5g of beef extract.

An exemplary device for performing the culturing is depicted in FIG. 1. Untreated yeast cells were cultured at 1X 10⁸Each yeast cell/1000 ml of the medium was added to the medium. The yeast cell may be saccharomyces hansenii AS2.504, or any strain selected from those listed in table 1. A typical activation process for yeast involves the following steps: yeast cells are grown in culture at 28-32 ℃ for 35-45 hours (e.g., 42 hours) before they are exposed to the following electric field: (1) culturing in an alternating electric field having a frequency of 11217MHz and a field strength of 190-250mV/cm (e.g., 240-242mV/cm) for 15-24 hours (e.g., 16 hours); (2) then culturing in an alternating current electric field with the frequency of about 11224MHz and the field intensity of about 220-250mV/cm (for example 235-237mV/cm) for 12-24 hours (for example 22 hours); (3) then culturing in an alternating current electric field with the frequency of 11231MHz and the field intensity of 430-450mV/cm (such as 440-444mV/cm) for 15-20 hours (such as 19 hours); (4) then culturing for 15-20 hours (for example, 17 hours) in an alternating current electric field with the frequency of 11237MHz and the field intensity of 490-520mV/cm (for example, 505-509 mV/cm); (5) finally, culturing the cells in an alternating current electric field with the frequency of 11244MHz and the field strength of about 210-230mV/cm (for example, 222-226mV/cm) for 8-12 (for example, 10 hours). Followed by culturing by various methods known in the artThe activated yeast cells are recovered from the medium, dried (e.g., lyophilized), and stored as a powder at 4 ℃. Preferably, the resulting yeast powder contains more than 10¹⁰Cells/g of cells. Adaptation of yeast cells to the gastric environment

Since the yeast composition of the present invention must pass through the stomach before reaching the small intestine, where the active ingredient is released from the yeast cells, it is preferable to culture the yeast cells under acidic conditions to acclimatize the cells to gastric juice. This adaptation process enables the yeast to have better viability in the acidic gastric environment.

To this end, the yeast powder containing activated yeast cells may be used at a level of 10g (more than 10 per gram)¹⁰Activated cells)/1000 ml were mixed with highly acidic acclimation medium. The yeast mixture is then cultured, first in the presence of an alternating electric field having a frequency of 11237MHz and a field strength of 480-520mV/cm (e.g., 510-514mV/cm), at 28-32 ℃ for 24-36 hours. The resulting yeast cells are further cultured at about 28-32 ℃ for 20-30 hours (e.g., 24 hours) in the presence of an alternating electric field having a frequency of 11244MHz, a field strength of 230-250mV/cm (e.g., 242-246 mV/cm). The resulting adapted yeast cells are subsequently recovered from the culture medium by various methods known in the art and are used in powder form (. gtoreq.10)¹⁰Cells/g) stored at room temperature or at 0-4 ℃ under vacuum.

An exemplary adaptation medium is prepared by mixing 700ml of fresh pig gastric juice and 300ml of wild chinese hawthorn extract. With 0.1M hydrochloric acid (HCL) and/or 0.2M potassium hydrogen phthalate (C)₆H₄(COOK) COOH) the pH of the acclimatizing medium was adjusted to 2.5. The pig gastric juice was prepared as follows. Newborn Holland white pigs, approximately 4 months of age, were sacrificed and their entire contents of the stomach were recovered and mixed with 2000ml of water under sterile conditions. The mixture was then allowed to stand under sterile conditions at 4 ℃ for 6 hours to precipitate food residues. The supernatant was collected for adaptation to the medium. Preparation of wild Chinese hawthorn extract in sterile strip500g of fresh wild Chinese hawthorn is dried under the condition of one piece to reduce the water content (less than or equal to 8%). The dried fruit was then ground (. gtoreq.20 mesh) and added to 1500ml of sterile water. The compound was allowed to stand under sterile conditions at 4 ℃ for 6 hours. The supernatant was collected for use in acclimatization medium. Preparation of Yeast mixtures

To prepare the yeast mixture of the present invention, the apparatus described in FIG. 2 or its equivalent may be used. The device comprises a first container (1), a second container (2), and a third container (3), each container being equipped with a pair of electrodes (4). One of the electrodes is a metal plate placed on the bottom of the container, and the other electrode comprises a plurality of electrode wires uniformly distributed in the space of the container so as to realize uniform distribution of electric field energy. All three pairs of electrodes are connected to a common signal generator.

The medium used for this purpose is a mixed fruit extract solution containing the following components per 1000L: 300L of wild Chinese hawthorn extract, 300L of jujube extract, 300L of extract from Schisandra chinensis (Turez) Bail seeds, and 100L of soybean extract. For the preparation of extracts of hawthorn, jujube and schisandra chinensis, fresh fruit is washed and dried under aseptic conditions to reduce the water content to not more than 8%. Then 100 kg of dried fruit (mesh. gtoreq.20) were ground and added to 400L of sterile water. The mixture was stirred under sterile conditions at room temperature for 12 hours, followed by centrifugation at 1000rpm to remove insoluble residues. For the preparation of the soybean extract, fresh soybeans are washed and dried under aseptic conditions to reduce the water content to not more than 8%. Then 30 kg of soybeans were ground into granules of not less than 20 mesh, and 130L of sterile water was added. The mixture was stirred under sterile conditions at room temperature for 12 hours, followed by centrifugation at 1000rpm to remove insoluble residues. To make the medium, these extracts were mixed according to the above recipe, the solution was autoclaved at 121 ℃ for 30 minutes, and cooled to 4 ℃ before use.

1000g of activated yeast powder prepared as above (part V above) was added to 1000L of mixed fruit extract solution and the yeast solution was transferred to the first vessel (I) shown in FIG. 2. Subsequently, the culture was carried out under aseptic conditions at about 28-32 ℃ for 12 hours in the presence of an alternating electric field having a frequency of 11237MHz and a field strength of 420-520mV/cm (e.g., 510-514 mV/cm). The yeast cells are further cultured in the presence of an alternating electric field having a frequency of 11244MHz and a field strength of 250-270mV/cm (e.g., 258-562 mV/cm). The incubation was continued for another 10 hours.

The yeast culture is then transferred from the first vessel (I) into a second vessel (2) containing 1000L of medium (if desired, a new batch of yeast culture can be started in the first vessel (1) which is now available) and subsequently subjected to an alternating electric field having a frequency of 11237MHz and a field strength of 220-250mV/cm (for example 233-237mV/cm) for 12 hours. The frequency and field strength of the electric field were then changed to about 11244MHz and 215-240mV/cm (e.g., 222-226mV/cm), respectively. The incubation was continued for another 10 hours.

The yeast culture is then transferred from the second vessel (2) to a third vessel (3) containing 1000L of medium and subjected to an alternating electric field having a frequency of 11237MHz and a field strength of 150-230mV/cm (e.g., 219-223mV/cm) for 10 hours. The frequency and field strength of the electric field were then changed to about 11244MHz and 160-. The incubation was continued for another 12 hours.

The yeast culture from the third container (3) is packaged into vacuum sealed bottles for use as a dietary supplement such as a health drink. If desired, the finished yeast culture can also be dried within 24 hours and stored in powder form. The dietary supplement can be administered 30-60ml each time, 3-4 times a day for 3 months, preferably 10-30 minutes before meal and bedtime.

In some embodiments, the compositions of the present invention may also be administered intravenously or intraperitoneally in the form of a sterile injectable preparation. Such a sterile formulation is prepared as follows. The sterilized healthy beverage composition was first treated under ultrasound (1000Hz) for 10 minutes, followed by centrifugation at 4355rpm for another 10 minutes. The pH of the resulting supernatant was adjusted to 7.2-7.4 using 1m naoh and then filtered through a membrane (0.22 μm for intravenous injection, 0.45 μm for intraperitoneal injection) under sterile conditions. The resulting sterile preparation was immersed in a water bath at 35-38 ℃ for 30 minutes before use.

The yeast compositions of the invention are derived from yeast used in the food and pharmaceutical industries. These yeast compositions therefore do not have the side effects associated with many pharmaceutical compounds.

Examples

In order that the invention may be more fully understood, the following examples are set forth. These examples are intended to illustrate the invention and should not be construed as limiting the scope of the invention in any way.

The activated yeast compositions used in the following examples were prepared AS described above using Saccharomyces cerevisiae Hanseng AS2.502 cultured in the presence of an alternating current electric field having electric field frequencies and field strengths AS exemplified in parentheses following the proposed use ranges listed in section IV above. Control (i.e.: untreated) yeast compositions were prepared in the same manner as described in section VI, except that the culture was performed in the absence of EMF. Unless otherwise defined, all yeast compositions and corresponding control compositions were administered to the animals by intragastric feeding. Example 1: effect of Yeast compositions on prostate, seminal vesicle and glandulae preputiales in Wistar rats

To test the ability of EMF-treated AS2.504 cells to modulate androgens, 24 healthy male Wistar rats (approximately 50g body weight, 20 days old) were selected. Under ether anesthesia, the testes of each rat were aseptically removed. These castrated rats were then randomized into equal 4 groups. The group A rats were administered 3ml daily of the activated yeast composition for 14 consecutive days. The control yeast composition was administered to group B rats at the same dose. Testosterone propionate was injected intramuscularly to group C rats in the buttocks at a dose of 25mg/kg for 14 days. Saline was injected subcutaneously or intramuscularly to group D rats at a dose of 2ml/kg for 14 consecutive days.

Each rat was sacrificed 24 hours after the last administration of the composition of interest. The seminal vesicles and prostate were withdrawn and placed in Bouin solution overnight. The adipose tissue surrounding the prostate was removed. The vas deferens, part of the urethra, and bladder were also removed from around the prostate. The remaining prostate was weighed and immersed in 70% ethanol overnight. The urethra was completely dissected from the prostate and seminal vesicle. The wet weights of prostate and seminal vesicle were recorded (table 2). The glandulae preputiales were also retrieved from the pubis. The wet weight of the removed glandulae preputiales was also recorded (table 2).

TABLE 2 Yeast compositions for prostate gland in Wistar rats,

Effects of seminal vesicles and glandulae preputiales

Group of	Compositions for administration	Prostate and seminal vesicle	Glandula preputipes
				A	EMF-treated AS2.504 (3 ml per day)	112.5±14.6	120.9±18.9
B	Unactivated AS2.504 (3 ml per day)	42.6±13.6	23.1±11.4
				C	Testosterone propionate (25 mg/kg per day)	91.5±12.8	98.6±15.8
D	Saline (2 ml/kg per day)	43.8±5.8	22.7±10.7

The results in table 2 show that the composition comprising EMF-treated AS2.504 cells (1) was able to stimulate the growth of prostate, seminal vesicle, and glandulae preputiales, whereas the control yeast composition was not; and (2) superior to testosterone propionate in stimulating the growth of prostate, seminal vesicle, and glandulae preputiales. Example 2: effects of yeast compositions on prostate, seminal vesicle and glandulae preputiales in mice

40 male NIH-grade castrated mice (approximately 25-30g each) were randomly assigned to 4 equal groups. Three days after castration, each mouse in group a was administered the activated yeast composition twice daily at a dose of 1.5 ml/dose for 7 consecutive days. Each mouse in group B was administered the control composition at the same daily dose for 7 consecutive days. Testosterone propionate was injected intramuscularly to group C mice every other day at a dose of 20mg/kg in the buttocks, i.e.: on days 1, 3, 5 and 7 for a total of 7 days. Saline was orally administered to group D mice at a dose of 1.5ml/kg for 7 consecutive days.

At the end of treatment, each mouse was sacrificed. Their seminal vesicles and glandulae preputiales were withdrawn and the surrounding adipose tissue was stripped. Seminal vesicles and glandulae preputiales were weighed and the results are shown in Table 3.

Yeast composition for treating prostate and seminal vesicle in mouse

And effects of the glandulae preputiales

Group of	Seminal vesicle (mg/100g body weight)	Glandulae preputiales (mg/100g body weight)
			A	361.3±66.1	356.6±61.2
B	39.1±18.7	107.5±41.2
			C	268.2±56.2	323.7±50.6
D	38.6±19.3	105.4±37.6

The results in table 3 show that the activated yeast composition (1) stimulates the growth of seminal vesicles and glandulae preputiales, whereas the control yeast composition does not; and (2) superior to testosterone propionate in stimulating the growth of seminal vesicles and preputial glands. Example 3: effect of Yeast compositions on Testosterone levels in Wistar rats

To test the effect of EMF-treated AS2.504 cells on testosterone levels, 36 healthy male Wistar rats (290 ± 30g) (10-12 months of age) were randomly divided into 6 equal groups. Each rat was treated according to the procedure outlined in table 4.

TABLE 4 treatment procedure for Wistar rats

Group of	Cyclophosphamide (20mg/kg) was injected 5 days before administration	Administration and duration of the composition
			A1	Is not injected	The activated yeast composition is administered at 3ml per day for 28 days
B1	Is not injected	The control yeast composition was administered at 3ml per day for 28 days

A2	Injection of drugs	The activated yeast composition is administered at 3ml per day for 28 days
			B2	Is not injected	The control yeast composition was administered at 3ml per day for 28 days
C	Injection of drugs	Distilled water is administered at 20ml per day for 28 days
			D	Is not injected	Distilled water is administered at 20ml per day for 28 days

Each rat was sacrificed at the end of treatment. Blood samples were collected from each rat and blood concentrations of testosterone were determined by standard Radioimmunoassay (RIA) methods. The results obtained are shown in Table 5 below.

TABLE 5 influence of Yeast compositions on Testosterone levels in Wistar rats

Group of	Testosterone concentration (ng/dl)
		A1	206.3±46.32
B1	146.12±43.32
		A2	172.42±52.53
B2	89.11±32.14
		C	87.31±23.51
D	145.28±34.34

The results in table 5 show: the activated yeast composition increased testosterone secretion, whereas the control yeast composition did not.

While several embodiments of the present invention have been illustrated, it will be apparent that modifications can be made to these basic concepts to yield other embodiments utilizing the compositions and methods of the present invention.

Claims (9)

1. A composition comprising a plurality of yeast cells, wherein said plurality of yeast cells are characterized by: the plurality of yeast cells are capable of stimulating the growth of prostate, seminal vesicle, and glandulae preputiales and increasing the secretion of testosterone in a subject as a result of having been cultured in the presence of an alternating electric field having a frequency range of 10500-12100MHz and a field strength range of 100-600mV/cm, as compared to yeast cells not so cultured.

2. The composition of claim 1 wherein said frequency range is 11217-11244 MHz.

3. The composition of claim 1 wherein said field strength is in the range of 150 and 520 mV/cm.

4. The composition of claim 1, wherein the yeast cell is selected from the group consisting of: saccharomyces cerevisiae, Saccharomyces carlsbergensis, Delbergi, Saccharomyces microsporum, Saccharomyces fermentans, Saccharomyces luogonii, Saccharomyces mellea, Saccharomyces ovatus, Saccharomyces rouxii, Saccharomyces sake, Saccharomyces uvarum, Saccharomyces williamsii, Saccharomyces carlsbergensis, Schizosaccharomyces pombe, Saccharomyces Sporobolomyces, Torulopsis pombe, Pseudobulbus fasciatus, Candida globosa, Candida vulgaris, Saccharomyces baileyi, Trichosporoides, Trichosporon, Saccharomyces cerevisiae, Candida xylostellata, Candida krusei, Candida lambertii, Candida lipolytica, Candida parapsilosis, Candida ferroportionto, Candida rugosa, Candida tropicaligenes, Candida utilis, Candida lamblia, Candida parapsilodosia, Hansenula pseudochinensis, Hansenula polymorpha, Hansenula pseudomondii, Hansenula pseudochinensis, Hansenula polymorpha, Saccharomyces submenus, and Saccharomyces limonum, grease yeast, Pichia farinosa, Pichia membranaefaciens, Rhodosporidium toruloides, Rhodotorula rubra, Rhodotorula aurantiaca, Saccharomyces rouxii, and Saccharomyces mellea.

5. The composition of claim 1, wherein said yeast cells are cells of a strain deposited at the China center for general microbiological culture Collection with a deposition number selected from the group consisting of Saccharomyces cerevisiae Hanseng AS2.375, AS2.501, AS2.502, AS2.503, AS2.504, AS2.535, AS2.558, AS2.560, AS2.561, AS2.562, and IFFI1048, and Saccharomyces carlsbergensis Hanseng AS2.420 and AS 2.444.

6. The composition of claim 1, wherein the composition is in the form of a tablet, a powder, or a health drink.

7. The composition of claim 1, wherein the composition is in the form of a health drink.

8. A method for treating androgen deficiency in a subject, comprising orally administering to said subject the composition of claim 1.

9. A method for preparing a yeast composition comprising culturing a plurality of yeast cells in the presence of an alternating electric field having a frequency in the range of 10500-12000MHz and a field strength in the range of 100-600mV/cm, wherein the composition is capable of treating androgen deficiency in a subject.