WO1986003515A1

WO1986003515A1 - A method and apparatus for separating cell tissue from an animal stomach

Info

Publication number: WO1986003515A1
Application number: PCT/DK1985/000117
Authority: WO
Inventors: Erik Vilhelm Balslev; Ernst Lorenz Pedersen; Hans Henrik Zinck
Original assignee: A/S Orthana Kemisk Fabrik
Priority date: 1984-12-11
Filing date: 1985-12-10
Publication date: 1986-06-19
Also published as: DK593484A; DK593484D0; AU5312686A

Abstract

Cell tissue containing biologically active substances, such as pepsin and mucin, are found in certain areas at the inner sides of animal stomachs (27). In order to provide a starting material for the production of such biologically active substances it is desirable to separate such cell tissue from the membrane or substratum of the animal stomach to which the cell tissue is fastened. Such preparation may be obtained by pressing a plate (29) with a plurality of small, closely spaced openings or perforations (60) against such area of the inner side of the animal stomach at a pressure sufficiently high to crush the cell tissue and to extrude the crushed cell tissue (59) through the perforations (60). The necessary pressure depends on the age and kind of the slaughtered animal from which the stomach stems and may be in the range of 5-50 Kg/cm2. When the stomachs are from slaughter swines, a pressure of 30 Kg/cm2 is suitable. The pressure need not be maintained for more than a few seconds, but before the pressure is totally relieved, the strings of crushed cell tissue extruded through the perforations (60) are preferably cut adjacent to the membrane or substratum to which the cell tissue was originally fastened. This may, for example, be obtained by using a plate comprising first and second plate members (28 and 29) displaceable between a position in which the openings or perforations of the plate members are aligned, and a position in which the perforations or openings are relatively displaced.

Description

A METHOD AND APPARATUS FOR SEPARATING CELL TISSUE FROM AN ANIMAL STOMACH

The present invention relates to a method of separating cell tissue containing biologically active substances from a membrane or sub- stratum in an inner side area of an animal stomach . Such cell tissue may be used as a basic material for the production of one or more of the active substances contained therein such as pepsin and mucin .

It is known to prepare a basic material for the production of pepsin or mucin by manually or mechanically separating the mucous mem- branes of the fundus regions from swine stomachs and by subsequent¬ ly mincing these separated parts with their adhering cell tissue. From the Danish patent specifications Nos . 145264 and 146543 it is also known to separate mucous cells from animal stomachs by cutting, abrading, or scraping the cells from their supporting membrane at the fundus region, while the membrane remains fastened to the stomach . By separating the cell tissue from the supporting membrane instead of separating the membrane with the interesting tissue from the stomach, a starting material containing an increased concentration of the inter¬ esting biologically active substances could be obtained . However, in practice, it has been found that it is hardly possible to separate cell tissue from a supporting membrane in the fundus region of a swine stomach by a cutting, abrading or scraping action at a desired high rate of production without also removing parts of the membrane and possibly also other underlying layers of the stomach . By this pro- cess, in order to avoid removal of parts of the membrane, the cutting device has to be extraordinarily sharp, which, in practice, can only be achieved for a short period during processing. As soon as the cutting device becomes dull, the resistance will result in undesi red removal of parts of the membrane.

The present invention provides a method by means of which cell tissue may be efficiently separated from a supporting membrane or substratum of an animal stomach at a reasonably high rate without simultaneous removal of parts of the membrane. Thus, the method according to the invention comprises pressing a first surface of a tissue separating device having a plurality of close¬ ly spaced small perforations or bores therein into engagement with said area of the stomach at a pressure sufficiently high to crush said tissue and to extrude the crushed tissue through said perforations or bores.

It has been found that when the perforated surface of such a separat¬ ing device is pressed into engagement with the selected stomach area, the cell tissue sandwiched between the separating device and the membrane of the stomach may be crushed, and the crushed tissue material will then flow to one of the adjacent perforations or bores and escape therethrough so that the said first surface of the separat¬ ing device may come into direct contact with the membrane.

The pressure which is necessary to obtain the desired separation of cell tissue varies within a wide range and is dependent on i .a . the kind of animal which the stomach stems from, the age of the animal, and the kind of tissue being separated . However, normally, the necessary pressure is 5-50 kg/cm². By separating cell tissue from the fundus region of a stomach stemming from a slaughter swine, the pressure is normally 20-40 kg/cm² and preferably 25-35 kg/cm², and the presently preferred pressure is about 30 kg/cm². When the stom¬ ach stems from a cow, a pressure of 15-25 kg/cm² would be suitable, and when the stomach is from a young animal the pressure could be reduced to 10-20 kg/cm².

The perforations or bores opening into said first surface of the separating device should have a cross-sectional area or diameter which is relatively small and a suitable mutual spacing so as to obtain the desired crushing of cell tissue. As an example, the diameter of the bores or perforations may be 1 -3 mm, for example 1 .5 mm, and the mutual spacing of the centres of the openings may be about the double, such as about 2.5 mm.

A stomach to be treated by the method according to the invention is cut and rinsed automatically in a conventional manner in such a manner that the stomach may be flattened out so that the interesting cell tissue to be separated is found only on one side of the flattened stomach . Alternatively, the stomach, which has been cut and rinsed in a conventional manner, may have its inside turned out so that areas with interesting cell tissue are arranged on opposite sides thereof. I n that case the stomach may be compressed between two oppositely arranged perforated tissue separating members so as to simultaneously separate cell tissue from the opposite areas of the stomach .

When substantially all of the crushed cell tissue has been extruded through the perforations or bores, part of the extruded strings of tissue may tend to retract from the perforations or bores when the pressure exerted by the separating device is relieved. Such retrac¬ tion of extruded cell tissue from the perforations may be prevented by separating the extruded tissue strings from the membrane adjacent to the membrane or substratum while the pressure is at least partly maintained. Such separation from the membrane could, for example, be obtained by displacing the separating device in relation to the membrane or substratum along said first surface of the separating device. However, such displacement may cause part of the extruded material to be smeared out over the membrane resulting in a lower yield, as the pressure exerted by the separating device has to be relieved to a relatively high extent. Therefore, according to a pre¬ ferred embodiment, the tissue separating device may comprise a thin perforated plate defining said first surface, which is pressed into engagement with the said area of the stomach, and the extruded tissue strings may then be separated from the membrane at the oppo¬ site second surface of the plate. Such separation may be made in any suitable manner, for example by means of a separating blade passing across said second surface of the thin plate.

One side of the stomach opposite to said inner side area may be arranged on a supporting su rface which is preferably somewhat resi¬ lient, before the separating device is pressed into engagement with the said area of the stomach , for example by means of a hydraulic or mechanical press . The resiliency of the supporting surface may com- pensate for possible irregularities of the wall of the stomach being treated. Any kind of animal stomach having cell tissue containing interesting substances may be treated by the method according to the invention . As an example, stomachs from swine, calves, cows, lambs or sheep may be processed.

The invention also relates to an apparatus for separating cell tissue containing biologically active substances from a membrane or sub¬ stratum in an inner side area of an animal stomach, said apparatus comprising a tissue separating device defining a first surface having a plurality of closely spaced small perforations or bores defined therein, and means for pressing said first surface of the separating device into engagement with the area of the stomach at a pressure sufficiently high to crush said tissue and to extrude the crushed tissue through said perforations or bores.

Generally, the said first surface of the separating , device may have any suitable concavely or convexly curved shape substantially com¬ plementary to the shape of a supporting surface on which the stomach is placed. I n the preferred embodiment, however, the said first surface is a substantially plane surface. Furthermore, the separating device may have any desired height or thickness provided that all of the bores communicate with a suitable receiving chamber or space or a suitable discharge passage or passages so that the cell tissue pressed into the bores may be discharged therefrom.

However, the said first surface is advantageously defined on a thin plate member, which may then be supported by a suitable supporting or backing structure. The plate member may have a thickness in the order of 1 mm. I n the preferred embodiment, the said tissue separat¬ ing device comprises a pair of superposed first and second perforated members having mutual contacting surfaces extending substantially parallel to said first surface defined on said first plate member, and means for moving said second plate member in relation to said first plate member between a position in which the perforations or bores of the first and second plate members are aligned, and a position in which the perforations or bores of said plate members are relatively displaced, whereby strings of tissue extruded into the perforations may be separated from the membrane at said contacting surfaces .

The invention will now be further described with reference to the drawings, wherein Fig . 1 is a perspective view showing a hydraulic press equipped with tissue separating devices for carrying out the method of the inven¬ tion,

Fig. 2 shows in an enlarged scale a side view and partially sectional view of part of the press table and of the tissue separating devices shown in Fig . 1 ,

Fig . 3 is an exploded view of a tissue separating device, and

Fig . 4 is a sectional and fragmentary view in a further enlarged scale illustrating the function of the separating device.

Fig . 1 shows a hydraulic press 10 with a ram plate 11 having a pair of tissue separating devices 12 mounted on its bottom side. Th ree pairs of stomach supporting blocks 13 are mounted on the upper side of a circular rotatable table 14. These pairs of blocks 13 are equally angularly spaced, and each pai r may cooperate with the pair of sep¬ arating devices 12 as explained in greater detail below. The table 14 may be rotated through angles of 120° by means of a rack and pinion drive comprising a pinion 15 mounted on a shaft 16 supporting the table 14, and a rack member 17 which may be reciprocated by means of a hydraulic or pneumatic cylinder 18. The rack member 17 and the cylinder 18 are pivoted at 19 and biased counter-clockwise by a spring member 20 mounted between the frame 21 of the press 10 and an arm 22 extending from the pivot 19 in a direction opposite to that of the rack member 17 and the cylinder 18. Thus, the spring member 20 tends to keep the rack member 17 out of engagement with the pinion 15. A pneumatic or hydraulic cylinder 23, which is mounted on the frame 21 of the press and extends substantially parallel to the cylinder 18 has a triangular head 24 mounted on the free end of its piston rod . An inclined su rface 25 on the head 24 may cooperate with a corresponding surface 26 formed on the rack member 17 so as to press the rack member 17 into engagement with the pinion 15 when the head 24 is moved to an extended position , while the spring mem- ber 20 keeps the rack member out of engagement with the pinion 15 when the head 24 is in a retracted position . The function of the cylinders 18 and 23 may be controlled so that the rack member 17 is pressed in engagement with the pinion 15 during the power strokes of the cylinder 18, but kept out of engagement during the return strokes of the cylinder 18. Thus, the rack and pinion mechanism may index the table in a desired direction through angles of 120° .

Animal stomachs 27 to be treated by the press with the separating devices shown in Fig. 1 are cut and rinsed in a conventional manner, and each stomach is manually placed on a pair of supporting blocks 13 in an unfolded or flattened condition with the inner side of the stom¬ ach facing upwards as shown in Fig . 1 . The shape of each supporting block may correspond to the shape of the area of the stomach 27 from which cell tissue is to be separated as described in greater detail below.

As best shown in Fig. 3 each of the tissue separating devices 12 comprises upper and lower thin plate members 28 and 29, respective¬ ly. Each of these plate members has a perforated area 30 with an outline corresponding substantially to the outline of the supporting blocks 13 and to the stomach area from which cell tissue is to be separated. The perforated areas 30 of the plate members 28 and 29 are identical . The lower plate member 29 is mounted on the bottom side of a surrounding rectangular frame member 31 , and the upper plate member 28 Is mounted on the bottom side of a tissue discharge block 32 by means of screws 33, and a spacer plate 34 is sandwiched between the upper plate member 28 and the discharge block 32. The spacer plate 34 has a perforated area 34a with an outline correspond¬ ing to the outline of the area 30 and may for example be made from expanded metal or wire mesh .

Thus, the discharge block 32, the upper perforated plate member 28 and the spacer plate 34 form an upper unit having outer dimensions which correspond to, but are slightly smaller than the inner dimen¬ sions of the frame member 31 . The upper unit is received in the frame member 31 so that the bottom side of the upper plate member 28 abuts on the upper su rface of the lower plate member 29. Parallelly extending cylinder bores 35 each receiving a cylinder sleeve 36 therein are formed in the discharge block 32 and open into oppo¬ site vertical side surfaces thereof . A piston 37 which is provided with peripheral sealing rings 38 and an axially extending piston rod 39 is received in each of the cylinder sleeves 36 and may be displaced between an inner position in which the piston rod 39 engages with the bottom end wall of the respective cylinder, and an outer position in which the outer end of the piston 37 extends beyond the adjacent vertical surface of the discharge block 32. Pressurized fluid such as pressurized air or liquid, may be supplied to the cylinder bores 35 through supply conduits 40 and 41 . The conduit 40 communicates with a connecting bore 42 which interconnects the inner ends of the cylin¬ der bores 35 at one side of the block 32, and the supply conduit 41 communicates with a connecting bore 43 interconnecting the cylinders 35 at the other side of the block. It is understood that the tissue discharge block 32 and the upper plate member 28 fastened thereto may be slightly displaced transversely in relation to the frame member 31 and, consequently, in relation to the lower plate member 29 by alternatingly supplying pressurized fluid to the supply conduits 40 and 41 .

A resilient pressure distributing assembly 44 comprises an upwardly open tray member 45 and a plurality of parallel, cylindrical resilient bodies 46 received in the tray member. These bodies, which may, for example, be made from solid rubber or plastic material , extend radially beyond the upper edge of the tray member 45 as shown in Figs . 2 and 3. The tray member 45 is fastened to the upper surface of the discharge block 32 by means of screws 47 as shown in Fig . 2, and the cylindrical bodies 46 are covered by a cover plate 48 having depending skirt portions 49 formed so as to engage with a shoulder 50 formed on the tray member 45 as shown in Fig . 2. The cover plate 48 also comprises a depending lug member 51 having elongated reces¬ ses 52 for engaging with locking pins 53 actuated by a spring 53a and received in bores 54 formed in the tray member 45 as best shown in Fig . 3. The pins 53 engaging with the recesses 52 and the skirt portions 49 engaging with the shoulders 50 are formed so as to allow limited mutual vertical movement between the cover plate 48 and the tray member 45. Hook-shaped members 55 fastened to and extending upwards from the frame member 31 may cooperate with locking devices 56 such as spring locks, which are fastened to the cover plate 48 so as to interconnect the cover plate 48 and the frame member 31 . How- ever, the interconnection established by the locking devices 56 allows limited movement of the discharge block 32 and the upper plate mem¬ ber 28 connected thereto in relation to the frame member 31 and the lower plate member 29. The cover plate 48 may cover two separate tissue separating devices 12 as shown in Fig . 2, and these devices may be mounted in the hydraulic press 10 by fastening the cover plate 48 to the ram plate 11 of the press as shown in Figs . 1 and 2.

Each of the supporting blocks 13 is made from rubber, nylon or a similar resilient material, and a friction increasing band 57 may be fastened to the rim portion of the upper surface of the block 13 so as to define a central portion 58 having substantially the same shape and size as the perforated area 30 of the plate members 28 and 29. Each of the stomachs 27 to be processed in accordance with the method of the present invention is placed on a pair of supporting blocks 13 as described above, and when the table 14 is rotated by means of the pinion and rack mechanism 15, 17, the supported stomach 27 is moved to a position immediately below the pair of tissue separating devices 12 mounted on the ram plate 11 of the hydraulic press 10. The hydraulic press 10 is now operated so that the tissue separating devices 12 are pressed against the stomach areas supported by the blocks 13. The mutual position of the plate members 28 and 29 is such that the openings or perforations of the plate members are aligned as shown in Fig. 4. The high pressure applied to the upper surface of the stomach 27 by the lower plate member 29 causes crushing of the cell tissue at the upper side of the stomach 27 and extrusion of the crushed tissue 59 upwards through the aligned perforations or open- ings 60 of the plate members 28 and 29. The crushed tissue may further flow through the openings of the spacer plate 34 and verti¬ cally extending bores 61 formed in the bottom part of the discharge block 32 and communicating with horizontally extending manifold conduits 62 formed in the block 32. Each of these conduits is con¬ nected to a discharge hose or tube 63 th rough which the separated crushed tissue 59 may be discharged from the block 32, for example by means of a pump, not shown . When the pressure on the stomach has been maintained for a few seconds so that substantially all of the interesting cell tissue on the stomach membrane has been crushed and extruded into the aligned bores 60, pressurized fluid is supplied through the conduit 41 to the cylinders 35 at one side of the block 32. This causes a slight displacement of the block 32 and the upper plate member 28 in relation to the lower plate member 29, whereby the strings of extruded cell tissue in the openings 60 are separated from the membrane. In order to facilitate the displacement of the block 32, the pressure may be somewhat relieved prior to the separating move¬ ment. The ram plate 11 and the separating devices 12 mounted there¬ on may now be lifted to the upper position shown in Fig . 1 , and the table 14 may be rotated so as to place a new stomach 27 in position below the separating devices 12. Furthermore, pressurized fluid is supplied through the conduit 40 so as to move the discharge block 32 to a starting position in which the openings or perforations of the plate members 28 and 29 are aligned . The operation described above may now be repeated .

EXAMPLE 1

Stomachs from slaughter swine were cut and rinsed in an automatic cutting and rinsing machine of the type in which the stomachs are moved along a conveyor bar extending through the intestinal openings of the stomachs, past a rotating cutting knife and succeeding spray- ing nozzles . The cut and rinsed stomachs were dried in a centrifuge.

Ten of such cut, rinsed, and dried swine stomachs were processed in an apparatus as that shown in the drawings. The thicknesses of the upper and lower plate members 28 and 29 were 2.5 mm and 1 .5 mm, respectively, and in both plate members, the diameter of each per- foration was approximately 1 .5 mm, and the centres of the perfora¬ tions were arranged in a pattern so as to form vertices of isosceles . The mutual distance of the vertices or the side lengths of the iso¬ sceles was approximately 2.5 mm. The perforated area 34a of the spacer plate 34 was formed by a wire mesh . The diameter of the wires forming the mesh was approximately 2 mm, and the size of the meshes was about 4 mm. The size of each of the perforated areas 30 of the plate members 28 and 29 and the perforated area 34a of the spacer plate 34 was approximately 170 cm². Each of the stomachs was placed on a pair of supporting blocks 13 so that the fundus regions of the stomach was supported and with the inner side of the stomach up¬ wards . Thereafter, a pair of tissue separating devices 12 mounted on a hydraulic press was pressed into engagement with the fundus regions of the stomach as described above with reference to the drawings . A total pressure of 10,000 kg - corresponding to approx¬ imately 30 kg/cm² - was maintained for 1 -2 seconds . Thereafter the pressure was relieved to about 4000 kg - corresponding to approxi¬ mately 12 kg/cm². While this relieved pressure was maintained, pres- surized fluid was supplied to the four cylinders 35 arranged at the same side of the discharge block 32 so as to generate a total displac¬ ing force of approximately 1256 kg for each discharge block, whereby the extruded strings of crushed tissue were separated from the membrane. This displacing force of 1256 kg was necessary to over- come the frictional forces between the plate members 28 and 29 at said relieved pressu re of 4000 kg.

The separating treatment of ten swine stomachs resulted in 720 g of separated crushed cell tissue of mucous cells .

EXAMPLE 2

I n a comparative test the mucous membranes of the fundus regions were removed from ten swine stomachs similar to those processed as described in Example 1 . These membranes were manually cut and torn off in a conventional manner. The total weight of the separated mu¬ cous membranes was approximately 1275 g, and these membranes were minced .

The proteolytic activity was measu red for the mass obtained in Example 1 as well as for the mass obtained in the present Example. It was found that the activity of the mass obtained in Example 1 was in¬ creased by 40% compared to the mass obtained in the present Example 2.

EXAMPLE 3

A single stomach from a cow was trimmed for fat, cut and rinsed . The total area of the inner side of the cut and flattened stomach was approximately 2150 cm². The weight of the stomach was 1004 g . The stomach was processed by means of a tissue separating device as that described with reference to the drawings . The perforated area of the separating device covered the above total area of the stomach, and the size and spacing of the perforations were as stated in Example 1 . The separating device was pressed against the inner side of the stomach at a total pressure of 40,000 kg for a few seconds, and an amount of 390 g crushed tissue was separated . The inner side of the stomach comprised several laminae, and it was found that also cell tissue positioned on the lower side of such laminae was crushed and extruded through the perforations of the separating device.

Claims

1 . A method of separating cell tissue containing biologically active substances from a membrane or substratum in an inner side area of an animal stomach, said method comprising pressing a first surface of a tissue separating device having a plurality of closely spaced small perforations or bores therein into engagement with said area of the stomach at a pressure sufficiently high to crush said tissue and to extrude the crushed tissue through said perforations or bores.

2. A method according to claim 1 , wherein a pressure of 5-50 kg/cm² is used.

3. A method according to claim 2, wherein a pressure of 20-40 kg/cm², preferably 25-35 kg/cm², and especially about 30 kg/cm² is used.

4. A method according to claim 2, wherein a pressure of 15-25 kg/cm² or 10-20 kg/cm² is used.

5. A method according to any of the claims 1 -4, wherein the inside of the stomach is turned out and areas with cell tissue containing biolog¬ ically active substances are arranged oppositely, whereafter these areas of the stomach are compressed between two oppositely arranged perforated tissue separating members so as to simultaneously separate cell tissue from said opposite areas.

6. A method according to any of the claims 1 -5, wherein strings of crushed cell tissue extruded through the perforations or bores are separated from the membrane or substratum closely adjacent to such membranes while said pressure is at least partly maintained .

7. A method according to claim 6, wherein the tissue separating device comprises a thin perforated plate having a side surface defining said first surface, which is pressed into engagement with the said area of the stomach, the extruded tissue strings being separated from the membrane at the opposite second surface of the plate.

8. A method according to any of the claims 1 -7, wherein one side of the stomach opposite to said inner side area is arranged on a some¬ what resilient supporting su rface, before the separating device is pressed into engagement with said area of the stomach .

9. A method according to claim 8, wherein the supporting surface is surrounded by a slightly raised area .

10. A method according to any of the claims 1 -9, wherein the bio¬ logically active substances comprise pepsin and/or mucin .

Tl . A method according to any of the claims 1 -10, wherein said animal stomach is from a swine, a calf, a cow, a lamb, or a sheep.

12. An apparatus for separating cell tissue containing biologically active substances from a membrane or substratum in an inner side area of an animal stomach, said apparatus comprising a tissue separat¬ ing device defining a first surface having a plurality of closely spaced small perforations or bores defined therein , and means for pressing said first surface of the separating device into engagement with said area of the stomach at a pressure sufficiently high to crush said tissue and to extrude the crushed tissue th rough said perforations or bores .

13. An apparatus according to claim 12, wherein said first surface is defined on a thin plate member.

14. An apparatus according to claim 12 or 13, wherein said tissue separating device comprises a pair of superposed first and second perforated plate members having mutual contacting surfaces extending substantially parallel to said first su rface defined on said first plate member, and means for moving said second plate member in relation to said first plate member between a position in which the perforations or bores of the first and second plate members are aligned, and a position in which the perforations or bores of said plate members are relatively displaced, whereby strings of tissue extruded into the perforations or bores may be separated from the membrane at said contacting surface.

15. An apparatus according to claim 14, wherein each of said first and second plate members is a thin plate members, the patterns of perforation of the first and second plate members being substantially identical .

16. An apparatus according to claim 15, wherein the tissue separating device comprises a first unit including said first plate member and a frame member extending from an opposite second surface or the contacting surface thereof, and a second unit having an outer wall surface defined by said second plate member and being received in the frame member of said first unit.

17. An apparatus according to claim 16, wherein said moving means comprise at least one fluid-operated piston-cylinder assembly.

18. An apparatus according to claim 17, wherein said assembly com¬ prises a cylinder housed within -said second unit, and a piston dis- placeably arranged within the cylinder and having its outer end in engagement with the inner surface of the frame member of said first unit.

19. An apparatus according to claim 18, wherein said second unit defines one or more hollow spaces communicating with the perforations of said second plate member and with discharge conduits for dis¬ charging crushed tissue.

20. An apparatus according to any of the claims 16-19, further com¬ prising means for connecting said first unit to said second unit when received therein , said connecting means allowing a limited mutual displacement of said units along said contacting surface.

21 . An apparatus according to claim 19 or 20, further comprising a cover plate overlaying said second unit opposite to said first unit, and resilient means such as rubber cylinders sandwiched between said cover plate and said second unit.

22. An apparatus according to any of the claims 12-21 , further com¬ prising a stomach support having a supporting surface for supporting the outer surface of the stomach opposite to said inner stomach area, said supporting surface comprising a surrounding raised rim portion .

23. An apparatus according to any of the claims 12-22, further com¬ prising a mechanical or hydraulic press for pressing said first surface into engagement with said area of the stomach when positioned on said supporting surface.