Ways Out of the BSE Crisis - Alternative Solutions

 

by Professor Dr.Dr.h.c. Randolph Riemschneider, Life B.Fel.[1]

Instituto Central de Quimica da Universidade Federal de Santa

Maria (UFSM), Santa Maria, RS, Brazil

Institute of Biochemistry, Free University of Berlin (FUB), Germany

 

[Editor’s Note: This paper is presented as Part II of a two-part series]

 

Keywords: Animal extracts: synthetic, cell culture based - plant extracts: yeast, cereal, glycoprotein based -  chemicytorrhysis - activation of cell metabolism, fermentation and respiration increasing factor; (WARBURG tests), ASO- and ASD-inhibition - applications in cosmetics, in medicine, as animal food additive, as food drug  -  whitening effect  -  proteinfree extracts  -  trade marks – BSE[2]

 

 

Owing to the pronounced increase in fermentation, the invention therefore makes it possible to carry out an aerobic fermentation process where a preparation of the invention is used to promote fermentation especially during the initial phase.

In discussions with master brewers at several breweries, the author learned that an acceleration of the brewing process is by no means desirable. A question asked by the author in this respect was answered as follows: "Dear Professor, we have time."

 

Automatic registration of fermentation:

We developed a special equipment to control the production and quality of Y 20 - the basis for several mentioned preparations – i.e. “Fermentationstest with automatic registration of the fermentation process under the influence of Y 20”: Illustration 2. This simple and rel. quick method (9-11 hours) is superior to other tests like animal tests, manometry (WARBURG), microbiology – and well to reproduce.

Registration curve in Diagram 5:

 

 

 

 

 

Illustration 2:

 

 

 

Procedure:  Die 4 auf 30° C vorgewärmten, mit Rührstäben gleicher Dimension versehenen Gefässe (1) bis (4) werden beschickt mit je 30,0g Glukose, je 5,0g Caseionpepton, tryptisch verdaut  (Merck), je 0,6g MgSO4•7H20, je 450ml vorgewärmtes Leitungswasser. Nach Zugabe von 3,0g Y 20 pur in Gefäss (1) und (2) und nach 15 Min. Zugabe 0,6g Trockenbackhefe in Gefäss (1) bis (4) wird jeweils auf 500ml  aufgefüllt. Dann werden die Magnetrührer eingeschaltet und die Gefässe geschlossen. 0-Stellung der Waage und Start aller Schreiber. Die Gefässe (3) und (4) sind Kontrollen: Kurve 2.

Beispiel in Diagram 5.

 

Für die Gefässe (1) und (2) wird die Zeit des Erreichens der maximalen CO2-Entwicklung (Knickpunkt der Kurve 1) abgelesen; im allgemeinen nach 9-11 Stunden:

% Wirksamkeit von Y 20 =  (Knickpunkt der Kurve 1)  :  (Knickpunkt der Kurve 2)   x 100

 

 

 

 

V  5.3  Claims of the PR China-patent  91105975.1

             in Chinese Plate 8a (document)                               in German Plate 8b (document)

 

 

 
 

re: claim 27, see V 5,4        re: claim 30, see V 6,1 and 6,2

 

 

 

V  5.4:   ATP formation (8a,b)[3]

 

In Diagram 6a and b you find the chronological progress of ATP formation in an incubated formulation (Diagram 6b in German language):

 

Plant cell preparations of the invention DE 4042157 C 2 (8a) show a surprising and pronounced influence on ATP formation in suitable systems such as a yeast suspension containing glucose and phosphate.

 

If a formulation of 1 g of baker's yeast, 4.0 ml of phosphate buffer having a pH of 7.4, 0.1 ml of yeast cell preparation (prepared as in Example 1), 1 mg of magnesium gluconate and 0.1 ml of manganese sulfate are incubated at 30°C for 60 minutes, and 2.0 ml of a solution of 30 mg of adenosine and 200 mg of glucose are then added and the mixture allowed to react at 37°C and a pH value of 7.4 for up to 270 minutes, a significant yield of ATP is recorded after some time, while the phosphate and adenosine content is reduced accordingly. Diagram 5 shows the measurement results of the increased ATP formation after 90', 120', 180', 240', and 270', all of the samples being worked up and the resulting ATP yields, the adenosine quantities consumed and the phosphate decrease being measured in each case. Without the addition of the cell preparation of the invention, practically no ATP production is measured under otherwise identical conditions (dotted line in diagram 5a,b).

 

Thus the invention using a plant cell preparation opens up a very simple and commercially practicable way to biochemical ATP formation in a yeast suspension medium to which adenosine and phosphate are added. Other nucleotides such as GTP, CTP and TTP may be prepared analogously.

 

Diagram 6a:

 

Diagram 6b:

 

 

 

 

 

 


V  6.1: 

TLC-chromatograms from DE 40 42 157 C2, fig. 3-6

 

In Fig 3 resp Fig 5 there are the TLC of the identified amino acids resp nucleo acid components of a yeast cell preparation prepared according to DE 40 42 157 C2. Fig  4 and 6: TLC of the starting materials.

Fig 3a. amino acid TLC of a dialysate Y 20 – starting material to develop the cosmetic additive CYTOCATALYZER; cf. claim 30 in Plate 8b.

 

 

 

Key to Fig 3-6:

 

 

Experimental details in PROJ XXIII, “Re-reading- 66 years chemistry” (20)

 

 

 


 

V   6.2:

CYTOCATALYZER – a yeast cell preparation based on Y 20

 

In the sections IV 1-3 several patents are described which are the basis for the production of metabolic active plant cell preparations; in this case from yeast, Saccharomyces we received the above mentioned starting material Y 20 which served to develop the prolteinfree cosmetic additives called CYTOCATALYZER  - in trade for many years in Japan.

Here follow Certificates1 and 2 of CYTOCATALYZER IKD and YSC:

 

Certificate 1:         CYTOCATALYZER IKD

Analytical data:     Appearance                           clear solution

                               Odor                                      deodorated

                               pH                                          6,7

                               dry substance                         4,1 %

                               N                                            0,4 %

                               metabolic activity              >  2,0

                               (WARBURG-factor)

Identification:         amino acids                          positive (ninhydrin)

                                peptides                                positive (biuret)

                                nucleo acid components      positive (TLC)

Puritiy:                    heavy metals                     < 20 ppm

                                As                                      <  2  ppm

                                sterility                                 germ free

 

 

Certificate 2:          CYTOCATALYZER - YSC

Analytical data:      Appearance                            clear solution

                        odor                                        deodorated

                                pH                                           6,7

                                dry substance                          7,1 %

                                N                                             0,6 %

                            aminoacidoxydase-                positive

                               inhibition

                        metabolic activity                > 2,0

                               (WARBURG-factor)

Identification:        amino acids                             positive (ninhydrin)

                        nucleo acid components        TLC

                        peptides                                  positive (biuret)

Purity:                            heavy metals                       < 20 ppm

                        As                                       <   2  ppm

                        sterility                                   germ free             

 

 

The CYTOCATALYZER preparations  passed all toxicological and dermatological test without showing any negative results, i.e. examinations like DL50 (mice), chronic toxicity (rats), teratogenity (rabbits), tests according to TURNER, BÜHLER, DRAIZE (eye,skin), PATCH, AMES. Tests explained in Table 6 in IV 4.


 

CYTOCATALYZER  IKD  -  16 amino acids determined

 

 

 


VI
  APPENDIX 

 

VI 1 

Cosmetic composition with a whitening effect, method for the production and use thereof  (19)

EP 03 718 674.9 – PCT/EP 03/01874  from 24.2.2003 (inventor: the author) 28p,  15 claims in Plate 10,

 

Abstract:

The invention relates to cosmetic compositions with a whitening effect, containing at least one enol lactone of the following general formula (I in Plate 9) thus cited and/or one or several tautomers thereof, also having the following formulae (Ia-Ic) thus cited or a salt, especially an alkaline earth salt, alkali salt or ammonium salt as an active ingredient. The invention also relates to a method for the production and use thereof for preventing the formation of brown skin spots and/or for lightening (whitening) or removing brown skin spots which are already present, especially for treating melanoses in mammals including humans.

 

 

Plate 9: General formula I and its tautomers Ia- Ic

 

Plate 10: Claims (document)

 

1. Kosmetische Zusammensetzung mit"Whitening-Effekt", die als Wirkstoff mindestens ein Enol-Lacton der nachstehend angegebenen allgemeinen Formel (I in Plate 9)  und/oder eines oder mehrere seiner Tautomeren der Formeln (la-ic) (la) (Ib) (Ic) worin Ri und R2 gleich oder voneinander sind, wobei Ri für Phenyl, C1-4-Alkyl-phenyl, insbesondere Benzyl, C1-4-Alkyl, C1-4-Alkenyl oder C14-Alkinyl steht, und R2 für COOH oder COOR (worin R die für Ri angegebenen Bedeutungen hat) oder für die gleichen Reste wie Ri steht, und worin die OH-Gruppen gegebenenfalls teilweise oder vollständig durch Halogenidreste, insbesondere Cl und/oder Br oder durch Oxyglycosid-, Thioglycosid-und/oder Aminoglycosid-Reste ersetzt sein können, und/oder ein oder mehrere Salze, insbesondere Erdalkaii-, Alkali-und/oder Ammoniumsalze, davon enthält.

2. Kosmetische Zusammensetzung nach Anspruch 1, dadurch gekennzeichnet, dass sie als Wirkstoff mindestens ein Enol-Lacton der in Anspruch 1 angegebenen Formeln (I), (la), (lob) und (Ic) enthält, worin Ri und R2 gleich oder verschieden sind und für CH3 oder C2H5 stehen.

3. Kosmetische Zusammensetzung nach einem der Ansprüche 1 bis 2, dadurch gekennzeichnet, dass sie als Wirkstoff 3-Acyl-3,4-dihydro-6-alkyl-2H- pyran-2, 4-dion der Formel (I) bzw. (la, lb, lc), worin Ri und R2 für CH3 oder C2H5 stehen, oder ein Salz, insbesondere ein Natriumsalz, desselben enthält.

4. Kosmetische Zusammensetzung nach einem der Ansprüche 1 bis 3, dadurch gekennzeichnet, dass sie den (die) Wirkstoff (e) der Formel (I) bzw.

(la, lb, Ic) in einerwirksamen Menge, vorzugsweise in einer Menge von 0,01 bis 5,0 Gew.-%, besonders bevorzugt in einer Menge von 0,1 bis 1,0 Gew.-%, speziell in einer Menge von 0,15 bis 0,5 Gew.-%, bezogen auf das Ge- samtgewicht der kosmetischen Zusammensetzung, enthält.

5. Kosmetische Zusammensetzung nach einem der Ansprüche 1 bis 4, dadurch gekennzeichnet, dass sie als zusätzliche (s) Additiv (e) Aldohexonsäuren, Ketohexonsäuren, Weinsäure, Citronensäure und/oder Ascorbinsäure enthält.

6. Kosmetische Zusammensetzung nach Anspruch 5, dadurch gekennzeichnet, dass sie die zusätzlichen Additive in einer Menge von 0,01 bis 5 Gew.-%, vorzugsweise in einer Menge von 0,1 bis 0,5 Gew.-%, bezogen auf das Gesamtgewicht der kosmetischen Zusammensetzung, enthält.

7. Kosmetische Zusammensetzung nach einem der Ansprüche 1 bis 6, dadurch gekennzeichnet, dass sie als weitere(s) Additiv(e) S-haltige Aminosäuren und/oder S-haltige Oligopeptide, insbesondere Glutathione, enthält.

8. Kosmetische Zusammensetzung nach Anspruch 7, dadurch gekennzeichnet, dass sie die weiteren Additive in einer Menge von 0,01 bis 3,0 Gew.- %, vorzugsweise in einer Menge von 0,1 bis 0,5 Gew.-%, bezogen auf das Gesamtgewicht der kosmetischen Zusammensetzung, enthält.

9. Kosmetische Zusammensetzung nach einem der Ansprüche 1 bis 8, dadurch gekennzeichnet, dass sie als weitere(n) Wirkstoff(e) keratolytisch wirksame Verbindungen, insbesondere Keratase und/oder alpha-Hydroxysäuren (AHA-Säuren), enthält.

10. Kosmetische Zusammensetzung nach Anspruch 9, dadurch gekennzeichnet, dass sie als weitere Wirkstoffe Rubiginsäure, Hydracetsäure, Comensäure, Chelidonsäure und/oder Mekonsäure enthält.

11. Kosmetische Zusammensetzung nach Anspruch 9 oder 10, dadurch gekennzeichnet, dass sie den (die) weiteren Wirkstoff(e) in einer Menge von 0,01 bis 5,0 Gew.-%, vorzugsweise in einer Menge von 0,1 bis 0,5 Gew.-%, bezogen auf das Gesamtgewicht der kosmetischen Zusammensetzung, enthält.

12. Kosmetische Zusammensetzung nach einem der Ansprüche 1 bis 11, dadurch gekennzeichnet, dass sie außerdem Sorbitol, Harnstoff, ein Succinat, Ethanol, Dexpanthenol sowie übliche kosmetische Zusätze, Hilfsstoffe, Träger und dgl. enthält.

13. Kosmetische Zusammensetzung nach einem der Ansprüche 1 bis 12, dadurch gekennzeichnet, dass sie in Form einer Lotion, einer Creme, einer Salbe, einer Lösung, eines Gels oder eines Sprays vorliegt.

14. Verfahren zur Herstellung'der kosmetischen Zusammensetzungen nach den Ansprüchen 1 bis 14, dadurch gekennzeichnet, dass man einen oder mehrere Wirkstoffe der Formel (I) und/oder Tautomere und/oder Derivate davon einer in an sich bekannter Weise hergestellten Lösung oder Dispersion der übrigen Bestandteile der kosmetischen Zusammensetzung zusetzt, nachdem die Lösung bzw. Dispersion auf einen pH-Wert &lt; 7,0, vorzugsweise zwischen 6 und 7, insbesondere zwischen 6,3 und 6,7, eingestellt worden ist, und das da- bei erhaltene Gemisch dann in üblicher Weise entlüftet, homogenisiert, kalt rührt und gegebenenfalls über einen Salbenstuhl laufen lässt.

15. Verwendung der kosmetischen Zusammensetzung nach den Ansprüchen 1 bis 14, insbesondere der darin genannten ADAPD-Wirkstoffe der allgemeinen Formeln I, la, lb, Ic, insbesondere von Methyl-ADAPD und/oder Ethyl-ADAPD, zur Verhinderung der Bildung bzw. zur Abschwächung oder Entfernung von braunen Hautflecken, insbesondere zur Behandlung von Melanosen, bei Mensch und Tier.

 

 

VI  2

Inhibition of enzymes breaking down amino acid by Y 20

 

Y 20 inhibits the amino acid-cleaving enzymes produced by enterobacteria (e.g. E. coli) such as amino acid oxidases (ASO) and amino acid decarboxylases (ASD): Plate 11a.

Escherichia coli is hardly able to cover its N requirement from non-protein sources (in contrast to lactobacillus). Therefore, E. coli  produces enzymes such as ASO, ASD et al. which, in turn, "destroy proteins". Y 20 inhibits these amino-acid cleaving enzymes and thus takes away sources supplying protein to E. coli.

 

Plate 11a:   Scheme for amino acid-cleaving enzymes such as amino acid oxidases L-ASO : (E 1.4.3.2)) and amino acid decarboxylases (L-ASD)

 

 

This E. coli-inhibiting activity of Y 20 is significant for its application in the animal feed industry, because it involves regulation of the intestinal flora. Proof was provided by the following experiments conducted in cooperation with Dr. M. Z. Abedin, Higher Institute of Technology, Brack, Libya.

 

In vivo experiment with E. coli: The ASO and ASD activities in the substrate of a liquid E. coli culture were measured both before and after the addition of Y 20.

The result showed 61 % inhibition for ASO, and 25 % inhibition for ASD.

 

The determination of the cell count of the samples treated with Y 20 showed that the E. coli count was by 1 to 2 powers of ten lower than in the control samples.

 

Thanks to the inhibition of the activity of ASO or ASD, a higher nutrient supply is available to the intestinal wall. The metabolic activity increased by Y 20 can fully exploit this additional nutrient supply with the result that nutrient uptake is improved and growth accelerated.

 

 

Experimental data to VI 2:

 

Plate 11b: Determination of the amino acid oxydase activity (WARBURG method)

 

Gl. 1: Summengleichung;   Gl. 2 und 3: Reaktionsgleichungen

Das gemäß Gl. 3 entstehende H2O2 würde von Versuch zu Versuch unterschiedlich zerfallen; darum Katalase-Zusatz, um H2O2 zu entfernen. Als Meßgröße dient Sauerstoff, der im WARBURG-Versuch bestimmt wird: ½ O2

 

Versuchsbeschreibung:

Zur Bestimmung des ASO-Hemmfaktors einer Y 20-Lösung werden nach Füllplan die vorbereiteten Lösungen 1 bis 8 in eine WARBURG-Apparatur gefüllt, so daß die Messungen erfassen:

L-ASO + L-Phenylalanin  (3,4)

L-ASO + Y 20  (5,6)  (ohne L-Phenylalanin)

L-ASO + Y 20 + L-Phenylalanin (7,8)

 

 

Table 12a :                  Füllplan:                        Manometernummern: 1 bis 8  (Lösungen in ml)

Lösungen   Füllort                1,2          3,4         5,6         7,8__________

3                 HR                     3,00        2,40       2,65       2,15

6                 HR                                                  0,25       0,25

1                 SA                                     0,1        0,1         0,1

2                 HR                     0,1          0,1         0,1         0,1

4                 HR                                    0,5                       0,5

5                 ZE                      0,2          0,2          0,2        0,2__________

 

Lösungen:     1)          L-ASO

 2)     Katalase-Suspension (1 : 10 verdünnt, Puffer pH 7,8)

             3)    Pyrophosphat-Puffer-HCL ph 7,8 (50 ml 0,2 M Na2P2O7 + 15 ml

0,5 M HCl

 4)     L-Phenylalanin (1 mMol/50 ml Puffer pH 7,8)

 5)     6 N KOH

                                    6)    Y 20-Dialysat

 

 

Die Zylindereinsätze (ZE) werden mit Filterpapierstreifen versehen, die mit 6n KOH getränkt sind. Die Lösungen werden bei 37°C equilibriert, dann wird die Enzymlösung aus SA in den die Testlösung bzw. eine „Blindlösung“ enthaltenen Hauptraum gegeben, um die Reaktion zu starten. Der µl-Verbrauch an Sauerstoff, in Abhängigkeit von der Zeit aufgetragen, ist ein unmittelbares Maß für den Hemmfaktor, wenn man mit dem  Sauerstoffverbrauch der „Blindversuche“ korreliert. Im allgemeinen erfolgt die zur Bestimmung des Hemmfaktors benutzte Ablesung nach 60 Min.

Zur Auswertung bildet man die Summenkurve der Meßkurven „ASO + L-Phenylalanin (3,4)“ und „ASO + Y 20 (5,6)“: Der Sauerstoffbedarf ist auf die in Y 20 enthaltenen Aminosäuren zurückzuführen, dann setzt man die Summenkurve, die ein Maß für die theoretisch zu erwartende Sauerstoffaufnahme ist, in Beziehung zur Kurve „ASO + L-Phe + Y 20 (7,8)“: Im Fall von Y 20 ergibt sich ein Minderverbrauch an Sauerstoff, der den prozentualen Hemmfaktor ausdrückt.

Die für Y 20 im Versuch Diagram 6 gefundene Inhibitorrate der L-Aminosäure-oxydase liegt bei 61 %.

 

 

 

Diagram 7:   

 Aminosäureoxydase-Hemmung durch Y 20

 

                                                                         O2 -Verbrauch in µl

253 µl

 

 
    ASO + L-Phe             (3,4)                                   106 µl

    ASO + Y 20               (5,6)                                   147 µl

    ASO + L-Phe + Y 20  (7,8)                                   155 µl______________                            

 

                                                         253  :  155  =  100  :  x

    Prozentualer Hemmfaktor                     x   =    61 %

             

                 

Tabelle 12b:    Füllplan

Manometer,                   1         2         3         4         5         6         7         8          Füllort

Gefäß Nr.

Acetatpuffer                3,0     3,0         2,0      2,0      2,5      2,5      1,75    1,75      HR

L-Glu in Puffer             --        --        0,75     0,75     --         --       0,75    0,75     HR

L-GDO in Puffer          --        --        0,25     0,25    0,25    0,25    0,25    0,25      SA

Y 20                             --       --           --         --        0,25    0,25    0,25    0,25     HR

 

                 Acetatpuffer pH 4,65  :  49,0 ml  1nNaOH + 100 ml 1n Essigsäure

                 L-Glu im Puffer          :   1,0 mg L-Glu in 1,0 ml Acetatpuffer

                 L-GDO-Lösung          :   125 mg L-Glutaminsäuredecarboxylase Typ III in 5 ml Acetatpuffer

 

 

 

Tabelle 12c:  Meßwerte

                                                    1           2           3           4           5           6           7           8

Manomertervol.                               --           --       1,332     1,409    1,323    1,346    1,309    1,352

Gefäßvol.                                                               14,362   14,218  13,949  14,024  14,353  13,041

Zeit  Min.     0                                  0         -1        +1           +1           0          0            0           0

                   10                                +2          0        +72         +67      +111    +115      +101      +104

                   20                                +4        +2        +87         +87      +  74    +  77      +  54      +  60

                   30                                +4        +1        +88         +92      +103    +105      +  74      +  80

                   40                                +3        +2        +88         +91      +108    +110      +  84      +  90

h korr.                                                   +3              +84         +87      +216    +222      +181      + 190

 

µl CO2                                                                           109,9                  274,9                  252,9

(Mittelwert)

                                                                                      I:                            II:                        III:

getestet                                                TB                L-GDO + L-Glu     L-GDO + Y 20   L-GDO + L-Glu + Y 20

 

 

 

 

VI 3

Ascitomycetes  taken into consideration in the patents ref (5,7,8) and used as starting material for the described preparations [cf. following diagram 1 in IV 1]

 

Schizosaccharomycetes (millet beer in Afrika, also to be found in molasses), endomyces lactis (milk mould, important for cheese maturing), saccharomyces (yeasts,  i.e. bakery-, brewery-, and bottom-fermented yeasts), saccharomyces cerevisiae var.ellipsoideus (vine yeasts), S. carlsbergensis (bottom fermented beer yeasts), S.fragilis JÖRGENSEN, S.lactis DOMBROWSKI, S.rosei GUILIERMONT, S.biosoprus NAGARISHI, Hansenula-yeasts as H.anomala HANSEN-SYDOW; trichosporon cutaneum OTA (occurring in bakery and fodder yeast), Candida tropicalis and C.utilis HENNEBERG L. and KR (Torulopsis utilis), C.Krusei (cream yeast) BERKHOUT; Kloeckera JANKE (vine musts), Sporobolomycetes (grain, water, hay, straw), Pichia polymorpha KLÖCKER (Pichia HANSEN).

 

 

VI  4

BSE detection: urine test – ultrafiltration technique – BSE free countries

 

As soon as the first BSE-reports were published and the BSE crisis had occurred, the author engaged in the following activities in Brazil in co-operation with: BRASTON Company, directed by Dr.F.R.Pesserl, and with CONSULTING DEVELOPMENT ENGINEERING under Dr.M.Faria. We received the possibity to experiment with infectious BSE material at the fazenda of Professor Dr.Jose Mariano da Rocha Filho, former Vice-Chancellor of UFSM and our long-standing friend. Mariano took care of all necessary safety measures which the intended experiments with BSE-infected cattle called for; as physician he was very familiar with all steps to be taken.

 

a )    In co-operation with Dr.Pesserl the author looked for a method to detect BSE in living animals as early as possible: Development of an ante-mortem diagnosis of BSE-infected cattle (1986-89)

b )    The author checked ultrafiltration technique to eliminate “infectious BSE-components” – in co-operation with the dres.Faria and Pesserl

c  )   He identified countries which were free  of BSE such as Brazil, New Zealand, Australia.

 

re a) The comparison of the “protein profiles of the urine” of healthy cattle with the urine of cattle infected with BSE made BSE detection possible (Riemschneider, Pesserl 1989) and has been put to use succesfully by us at that time: Plate 12.

 

            After the author had learned that a “Research Institute for Animal Epidemics” had been established on the island of Riems in the Baltic Sea, he tried to establish contact. His inquiry "in which organs and body cells of cattle with BSE prions had been found" was not answered, and no reaction came to the "information regarding a method of detecting BSE (protein profile of the urine)" he submitted (letters from July 23, 2005 on).

 

re b)     In a 34 page expert opinion of CONSULTING DEVELOPMENT ENGINEERING in 1993 regarding the absence of BSE in the proteinfree thymus extract OMNITHYMUS, Charge STK 1993-1, initiated and supervised by the author, the activity of the ultra filtration technique employed for this extract was established, cf. 3 documents: i) Summary,  ii) list of contents of the 1993 expert opinion, iii) certificate regarding the examination of the OMNI­THYMUS extract supplied to Japan in 1996 concerning infectiousness and a possible content of BSE prions: BSE-free (next pages).

 

       Until December of the year 2000, i.e. until the import ban imposed by the Japanese government came into effect, preparations produced from calf thymus serum in Germany (14) had been supplied to Japan if the absence of BSE was guaranteed by

       -    a urine test

       -    proving that the ultra filtration technique used during the manufacture of the preparations was effective; see documents belonging to i) and  ii) next page

       -    and, where possible, obtaining the animal organs from BSE-free countries.

 

Since no prions were found in porcine organs between 1986 and 2008, requests for proteinfree porcine organ extracts and porcine collagen increased from 2001 onwards and could be fulfilled (3).

As early as the 1950s, the author had developed extracts from porcine placentae for the Brazilian industry (3), i.e. long before bovine placenta extracts became en vogue; see the publication: "Porcine-based organ extracts guarantee full substitution of bovine extracts" (23).

 

 

 

Plate 12:   Announcement of the round-table conference in Rio de Janeiro, directed by M.M.Faria, Chairman of CONSULTING DEVELOPMENT ENGINEERING:

 

 

 

Invitation to a round-table conference on the subject:

                    

“Ante-mortem diagnosis of BSE-infected cattle”

     

In the library of CONSULTING DEVELOPMENT ENGINEERING, in Rio de Janeiro on November 12th, 1989 will be given the report of the experiments, entitled above, carried out from Riemschneider and Pesserl in Rio Grande do Sul 1986-1989 under the patronage of Professor Dr Jose Mariano da Rocha Filho at his fazenda.

Initiator and project leader: Professor Dr.Dr.R.Riemschneider, member of the Brasilian University Universidade Federal de Santa Maria (UFSM), in Santa Maria, RS, Brazil, and retired professor of the Free University of Berlin (FU), Institute of Biochemistry. Dr.F.R.Pesserl is directing the BRASTONE Company in Curitiba, PR, Brazil, acting as lecturer giving the report of the mentioned authors Riemschneider and Pesserl, in Portuguese, entitled  as above written.

 

It will be presented the new ante-mortem test based on the body fluid urine applying the method of special gel-electrophoresis *  to compare the urine protein profiles of cattle – including experimental data in detail. The authors used the urine of 10 calves, 5 of them infected with BSE at 3 months of age, 5 of them as controls; during 18 months urine samples were taken nine times.

 

Rio de Janeiro, November 1989

 

 

This round-table talk is not public – only for invited guests to be kept secret

 

______

 *   after suitable  preparatory treatment of the urine samples (instantly deep frozen after taking)

  

 

We refrained then from any publication for following reasons:

- a)   to avoid complications with the Brasilian Health Department because of the experiments with BSE infected material in Brazil; especially also to protect Prof Mariano’s private initiative,

- b)   to avoid also complications with the rather inflexible German authorities.

- c)   the author remembered well the attacks of the radical students, starting 20 years ago (68), and will never forget the “science-destroying” new university law of 1969 concerning the so-called democratisation of the FU and the consequences – so that the author found himself compelled to transfer some of his current important research projects to Brazil and to call on highly industrial help in Brazil and other countries, especially Japan.

 


Three documents (mentioned in VI 4)

 

i)    Summary of the 1993 expert opinion

       regarding the ultrafiltrated thymus extract OMNITHYMUS[4] tested on infectious properties and possible BSE-prions – test animal: Syrian golden hamster, observed 365 days, microscopical anatomic inspection of their brain – 34p report.

This report and the report iii) were initiated and supervised by the author who worked yearly some months in Brazil as a member of the UFSM in Santa Maria, RS.

It is quite important to point out that in Germany in the 90ies, it would have been very difficult to receive official Government permission to carry out experiments like these because of the “Danger of working with BSE infected material” and  the “animal protection lobby”.

 

Result: Extract, free of infectious particles like scrapie.   

 

 

 

 

 


ii)  List of content of the OMNITHYMUS expertis  [summary in i)]

 

 

 

Long-winded and complicated expertises like the above documented one and preliminary BSE-tests made it possible to continue with the production and export of animal based preparations used in medicine and cosmetics - without any interruption – in spite of the BSE-crisis.

 

Reading the 34 pages document with one year lasting  Syrian golden hamster experiments you must take into consideration the situation of 1985/86: In England, thousands and thousands of cows had to be clubbed, in spite of the prohibition calves were exported to Holland and other countries; the nature of the pathogen particle responsible for BSE was not known then; there was a certain similarity of the symptoms of BSE with the JAKOB-CREUTZFELD syndrome and scrapie (Australia).  Summarized under the term TSE = transmissible spongiforme encephalopathies

 

All this then called for immediate  action – cost what it may.


 

iii)        First page of the 1996-Certificate -  regarding the examination of OMNITHYMUS, prepared according to the prescription of the author  -  33 pages report, similar to i) and ii).

          Result: Extract free of infectious particles like scrapie or scrapie-likes.

 

 

 

 

Conclusion   

The methods of preparation described have been used successfully in different fields, especially in combination with each other, but also in combination with native preparations. As mentioned above, a pharmaceutical preparation on the basis of a synthetic organ extract has been put to clinical use as long as 14 years before the BSE crisis.

 

Based on the patents described, the following preparations were developed:

 

Cytocatalyzers  as proteinfree cosmetic additives: yeast-based

Seruryel, Tricell as proteinfree cosmetic additives: cereal-based

Colezzo as food-drug: cereal-based

H 2000, Y 2000 as animal food additive (yeast-based)

Aphrodisiac (yeast based)

Collaplant PO as collagen substitute (glycoproteine-based)

Canivit for dogs (yeast based)

Omnithymus S,  a synthetic proteinfree extract (thymus analogon)

 

Depending on requirements, each of the three patent groups (5,7,8) achieved optimal solutions. The author has investigated the “active principles” of the reaction products received by the methods described in these patents (5,7,8). The results will be published later. In the EU-patent (1) these “active principles” are already used; see the component description a) - c) in (1).

The patent (16) permits an increase of the whitening effect of synthetic or native preparations.

 

REFERENCES  -  by the author (co-workers in brackets)

 

 (20)    Re-reading  - 66 years chemistry: XXVI PROJECTS (in preparation) here: PROJ XXII and XXIII

                

(21a)    The negative influence of regular intact yeast (baker's yeast) on the potency of male mice when given with the drinking water.

            Lab report June 1938, 15 pages, abbreviated version published in 1938 in the school magazine of Wilhelm Gymnasium, Hamburg-Dammtor (co-workers: A. Suhr, H. Kahl)

            These orientation experiments were carried out during the period May 1937 to June 1938 with the approval of the administrative board of Matthias-Claudius-Gymnasium, financed by the family of the author's schoolmate A. Suhr from Wilhelms-Gymnasium.

 

(21b)   "Yeast and yeast extracts - Saccharomyces cerevisiae HANSEN – BIOCOMPLEX YEAST"

            Lecture held by the author in December 1939 at the Chemical Colloquium of the Department of "Analytical Chemistry" of the Chemical Institute of Göttingen University (discussion chaired by Profs. Dr. J. Goubeau and Dr. G. Rienäcker), duration of the lecture: 60 minutes.

            General lecture, state of the literature until 1938 by Chemical Abstracts, jointly prepared with A. Suhr (student of medicine) and A. Kersting (student of chemistry)

            On the topic: BIOCOMPLEX YEAST: No natural product deserves being called a bio-complex more than yeast. State-of-the-art chemical analyses have shown that beer yeast or baker's yeast, respectively, are the plants with most active ingredients we know so far; possibly to be compared only with cereal grain [e.g. rye, wheat (11,28,29)]; cf. PROJECT XXII in (20).

 

(22)     Lecture "Test method for examining aphrodisiacs: First tests for identifying and isolating the "factor of yeast inhibiting the potency of mice", held by the author at the Colloquium of the Institute for Physiological Chemistry at Berlin University in April 1948 [discussion chaired by Prof. Dr. K. Lohmann, discoverer of ATP; cf re 5) KARL LOHMANN in (15)].

 

            The results of "Aphrodisiacs", Comm. I to VIII (20), were presented and discussed. By special invitation of Prof. Lohmann, the following were present:

            - Nobel Prize winner Otto Warburg, Berlin,

            - Prof. Dr. J. Just, Institute of Commercial Fermentation, Berlin, and Institute for Organic Chemistry, of Berlin University,

            - Dr. A. Heymons, Director of Riedel de Haen, Berlin,

            - Prof. Dr. E. Thilo, Director of the Institute for Inorganic Chemistry of Berlin University,

            - Prof. Dr. K. Noack, Dean of the Department of Mathematics and Natural Sciences at Berlin University and Director of the Botanical Institute at the same address.

            In this address to the above-mentioned personages, the author first spoke in public about the "unexpected results" of the WARBURG experiments obtained with such "yeast extracts" he had prepared while "looking for the factor inhibiting the potency of mice"; a very high respiration-increasing factor of 3 to 4 had been found for these plant extracts. Under corresponding test conditions, the factors found for animal extracts (like extracts from foetal calf serum) were only between 1.8 to 2.4      (12-14); cf. PROJ. XXIII (20).

            More details on the communication series "Aphrodisiacs" are found there, too.

 

(23)     Porcine based organ extracts guarantee full substitution of bovine extracts

            http://www.vevy.com/relata/issues.articles  2001, 3p  [co-author: M. Heisler]

International Electronic Journal on Dermopharmacological Research, Dermopharmaceutical   Technology and Related Cosmetic Subjects

 

(24)                 “The animal food additive: S 2000 (Y 2000), based on Y 20-suspension” spray dried”

Lecture given in Moscow on Oct 1988 in presence of

-   invited competent Russian scientists from the different parts of the USSR,  

-   industrialists (fodder industry),

-    fur farmers,

-    authorized persons of the Government.

-    bankers and German guests

The lecture was held by the author two times in the aula of the Deutsche Bank and simultaneously translated into the Russian language, 120 min, 25 diapositives. Summary of the lecture in Plate 5.

The author handed over the special report mentioned in (8c) to everybody.

 

(25)     Lecture identical with (24), held in Gdansk on March 1989, invited from the Polish industry: NAVIMOR, Ul.Jana Hewelnsza,11.

Present: Professor Dr. Czeslaw Lewicki, Agricultural and Technical Academy in Olsztyn, Poland Institute of animal Nutrition, Inz.Jerna Zurawicz, Dyretor Zukladow, Odanskie Zaklady Dbobiarskie, W Zukowie, Zukowo,  Dr.inz. Stanislaw Blaziak, Manager of Central, Laboratory of the Feed Industry, Lublin, Poland, NAVIMOR:  Direktor K. Krzymowski, A. Tmaszewski, Mrs V. Waiss, L. Drobotowicz

from BACUTIL: Direktor J. Krzaczek, Gdansk

 

Negotiations for Y 2000, continued succesfully in Berlin on Sept 19 and 20, 1989 with the Polish delegation of Landwirtschaftl.-Technische Akademie, Olsztyn:  Prof. Cz. Lewicki, Prof. A. Faruga, Mgr. D. Mikulski, Zootechn. Institut, Krakau: Dr. Doc. J. Koralewski Akademie der Wissenschaften: Jablona/Warschau Dr. B. Pastuszewka Zentrallabor für Futtermittel, Lublin: Dr. St. Blaziak;

BACUTIL, Gdansk:  Dir. J. Krzaczek, NAVIMOR: Dir. K. Krzymowski, Tomaszewski and Mgr. L. Drobotowicz, lawyer

 

Prof.Riemschneider repeated his lecture held in Gdansk in March.

Mikulski, Koralewski, and Krzaczek reported about “Massentierzuchtversuche mit Y 2000 and a running chicken field experiment with 6000 t added the new food animal additive Y 2000;

Mrs Pastuszewaka spoke about her lab trials with mkice, rats, rabbits, fur animals;

Mr Blaziak about results with pigs and fish.

 

(26)     Lecture identical with (24), held in Szombatlei, Hungary, on Aug 1989

Discussion of the pig experiments carried out in Brazil: Plate 5a,b.

Daraufhin sind in der Zeit vom 20.4. – 14.6.1990, initiiert durch diesen Vortrag, mit H 2000-Prämix

(20 % Y 20) – genannt PROVAL - in der Produktions-gemeinschaft Gyözolom in Lajoskomárom (Schweinemästerei) mit 306 Nagy magyar fehér + Sved lapály [große ungarische Weisse u. Schwedische Niederung] Ferkel-Versuche durchgeführt worden. Die brasilianischen Versuchs-ergebnisse aus den Jahren 1969 bis 1970 konnten bestätigt werden.

 

(27)     Three lectures in Dayin Village, Jinghai Tianjin, China in Nov 1992

invited from the Comprehensive Agricultural Corporation, Tianjnin; simultaneous tranlated into Chinese language

Lecture I and II:  S 2000 als Futtermittelzusatzstoff zum Produkt Feed BE Yeast,

einer chinesischen Futterhefe – Eigenschaften und Ergebnnisse der Feldversuche mit Hühnern, Ferkeln und Rindern aus Brasilien, Japan, USSR  (24), Polen (25), Ungarn (26) und der Schweiz.“ Present:

- Mr. Li Feng Zhang, Chairman of the board – General Manager Hua Da Forage Industrial Corp.,         Tianjin

- Mr. Zhou Zhijiang, Professor and Vice-President of the Preparatory Committee of  Man-made Fiber Industry Association China

- Mr. Liu Baoshun, Associate Professor, Beijing Research Institute for Nutritional Resources

- Mr. Wang King Min, Attead Office Director, Daqui Comprehensive Agricultural Corporation Tianjin

- Mr. Yu Sao Wu, Deputy Manager, Hua Da Forage Industrial Corporation, Tianjin

- Mr. Song Bao, Deputy Manager Factory Director, Daqui Poultry and Aquatic Breeding Farm, Tianjin

- Mr. Kauer, Kauer-Industrie Consult and Trading GmbH, Berlin

- Mrs. Kauer-Ong of Kauer Industrie Consult and Trading, also interpreting (German, English, Chinese)

Lecture III  : „Probiosis versus Antibiosis“

After lecture II a long discussion took place about the results of the field experiments  concerning the combination of BE with S 2000 (FEED  BE – an animal feed yeast produced in China).

The last mentioned lecture was given for the first time on Sept 1963 in S.Paulo, Brazil, then 1990 in Sudermühlen: SCHÜLKE & MAYR Workshop.

          

(28)    Preparation of different plant extracts and test on their cell metabolism increasig properties. Extracts from: rye, wheat, cactus, tea, papaya, ginseng ecc

Lab reports, from  Dez 1947 on

 

(29)     Intermediate metabolism – organ and plant extracts

                Bull. I – LV (1946-2005): PROJECT XXIII in  (20)

            here:

            Bull.X: Cell culture experiments: Wound healing by plant extracts like wheat, rye, ginseng, cattail, 15 series experiments,

                        ms 1973 (with R.Lipp, W.H.Chik)

Bull. XLV: Plant extracts from placenta of triticale, rye, wheat, corn, barley, oat, rice ecc - preparation and properties - interesting for cosmetics: CELLRYEL (SERURYEL)

                        ms Dec 1987, 22 p (secreted)

            Bull. XLVI : New cosmetic additives  based on rye and wheat: CELLRYEL, WHEATIN – activity data  (with U.Rotter, M.Azhar) -  first draft of a patent application.

            Ms Jan 1988

            Expert opinions about activity of CELLRYEL by Hwa Sook Chang, Hongkong, by M.Azhar, Islamabad, Pakistan

 

 

In memoriam:

This article is dedicated to the author’s long-standing Brazilian friends M.M. Faria and J.M. Faria, who unfortunately died in the crash of a private plane belonging to colleagues.

 

 



[1]     Adress correspondence to Prof. Dr. Dr. R. Riemschschneider, D-14001 Berlin, Postfach 1164, Germany

[2]     BSE = bovine spongiforme encephalopathie (10)

[3]      See Plate 8b (claim 27)

[4]  Die vom Verfasser für OMNITHYMUS entwickelte Herstellungsvorschrift umfasste folgende Stufen: Zerkleinern der gewaschenen Drüsen - Suspension in Ethylether bei -20° C  -  Zentrifugieren (explosionssichere Trommelzentrifuge) – Dialyse des Zentrifugats – Standardisierung – Sterilfiltration. Ab 1986 ist dieser, seit 1970 nach Japan exportierte proteinfreie Organextrakt zusätzlich ultrafilriert worden (Gutachten); später wurden darüber hinaus noch Thymusdrüsen verwendet, die aus einem BSE-freien Land stammten (bis zum Inkrafttreten des neuen Gesetzes im Dezember 2000).

 



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