阅读说明：本技术 用于测量胶原包装材料的热收缩性的装置和测试方法 (Device and test method for measuring thermal shrinkage of collagen packaging material ) 是由 胡安 阿古斯蒂·索雷特·马顿 于 2020-06-16 设计创作，主要内容包括：一种用于测量胶原膜的热收缩性的装置和方法,其中,该装置设置有水热容器桶和用于加热水浴的可调节加热器、温度探针、用于所述膜的试样的抓取夹、构造为搁置在桶的边缘上并且容纳上抓取夹的第一杆、固定到桶的底部的一组滑轮和用于将试样的张力传递到桶的外部的第二杆、设置有位移传感器的位移测量单元和设置有用于获取、处理和显示位移和温度数据的程序装置的处理器。该装置简单、经济、可靠,能够提供与通过诸如DSC的量热技术获得的数据相当的数据,并且便于在同一过程中对多个样品进行测试。(An apparatus and method for measuring the heat shrinkage of a collagen film, wherein the apparatus is provided with a hydrothermal receptacle tub and an adjustable heater for heating a water bath, a temperature probe, a grip clamp for a sample of the film, a first rod configured to rest on the edge of the tub and to receive the upper grip clamp, a set of pulleys fixed to the bottom of the tub and a second rod for transmitting the tension of the sample to the outside of the tub, a displacement measuring unit provided with a displacement sensor and a processor provided with program means for acquiring, processing and displaying displacement and temperature data. The device is simple, economical, reliable, capable of providing data comparable to that obtained by calorimetric techniques such as DSC, and facilitates testing of multiple samples in the same process.)

1. An apparatus for measuring heat shrinkage of a collagen film, comprising:

a hydrothermal vessel barrel (2) and an adjustable heater (3) for heating a water bath;

a temperature probe (8) within the container;

upper and lower gripping clamps (4) for at least one sample of the membrane (1);

a first bar (6) configured to rest on the rim of the bucket and to receive the upper grip clamp such that the specimen hangs from the first bar and extends into the bucket;

a set of pulleys (10) fixed to the bottom and second bar of the bucket and at least one wire (9) attached to each lower gripping clamp (4) to transmit the tension of the specimen to the outside of the bucket through the set of pulleys;

a measuring unit (11) for measuring the displacement of the test specimens, which measuring unit is provided with a displacement sensor (17) for each test specimen, wherein each sensor is attached to each wire (9);

a processor (13) provided with program means for acquiring, processing and displaying said displacement and temperature data.

2. The device for measuring heat shrinkage of a collagen membrane according to claim 1, wherein said device is provided with a ring (7) to fix said upper gripping clip to said rod and said lower gripping clip to said wire.

3. The device for measuring the heat shrinkage of a collagen film according to any one of the preceding claims, wherein the cross section of the barrel is polygonal or circular.

4. The device for measuring the heat shrinkage of a collagen film according to any one of the preceding claims, characterized in that said measuring unit (11) is provided with an analog displacement measuring element, allowing a visual reading of said displacement.

5. The device for measuring the heat shrinkage of a collagen film according to any one of the preceding claims, wherein said first rod (6) is circular.

6. The device for measuring the heat shrinkage of a collagen film according to any one of the preceding claims, wherein said heating system is provided with means for thermal agitation by a magnet.

7. A method performed with the apparatus according to any one of claims 1 to 6, comprising the steps of:

a sample of at least one collagen membrane in the form of a rectangular strip was prepared,

immersing the sample in the bath solution for 15 minutes until the water content of osmotic absorption is balanced;

calibrating the starting position of each sample to zero;

once the bath heating ramp was selected and the approximate initial shrinkage temperature of the collagen was known, a pre-heated bath was introduced until nearly all of the sample was covered and heating by the heater was initiated;

collecting, by the processor and the programming device, a digital signal caused in the sensor by the contraction of the sample and a digital signal from the temperature probe.

Technical Field

The present invention belongs to the field of food, and in particular, it relates to an apparatus and method for measuring the heat shrinkability of a collagen film manufactured as a food packaging material, mainly meat.

Background

For over a century, some thermal properties of native collagen have been known, such as shrinkage when heated at certain temperatures. Pairoy (Flory) et al, who have made extensive studies in this regard in the 1950 s, proposed a mechanism of contraction. Among them, they describe how to generate thermal contraction of collagen by the transition between the crystalline and amorphous phases of this protein; and they demonstrated that the transition is completely similar to that involving the fusion of other crystalline polymers. This process is described as "gelling" or also as "denaturation".

Collagen has a structure that exhibits a hierarchical structure, which is characteristic of biological systems. The chemical structure of collagen occurs through a helical "procollagen" of three chains linked by hydrogen bonds; "microfibers", having the five-fold alignment of the aforementioned substances, with directional repetition of about 64nm, and collagen "fibrils", consisting of microfibers filled with a tetragonal lattice. In biological tissue, the fibrils are surrounded by an additional fibril matrix consisting mainly of mucopolysaccharides. Recent studies (Admir Master et al; 2015; Max Planck Institute, Mass.) further showed that water is an integral part of the helical structure of collagen, accounting for approximately 60% by weight of the material.

It has been proposed that the onset of collagen denaturation begins with the unwinding of the triple helix. At a certain temperature, called Ts, the contribution of thermal energy causes the triple helix to melt synergistically and the collagen molecules change from a linear helical shape with a high elongation to a much smaller average length random sphere, which is observed macroscopically as a very significant contraction of the collagen fibrils. Thus, there is a high correlation between the observed thermal aspects (heat absorption, enthalpy of denaturation, etc.) and mechanical aspects (degree of shortening, tension produced, etc.), since both are single processes, a reflection of the melting of the crystalline structure of the collagen molecules.

Subsequently, during the course of the temperature increase, hydrolysis of the thermally labile crosslinks may occur, while the maintenance of the thermally stable crosslinks is responsible for the residual tension in the collagen fibrils.

It has also been shown that the thermal properties of collagen vary with the age and environmental conditions of the animal. Shrinkage temperature tests, which link the molecular structure to the stability of collagen, are conventional in the tanning industry to assess the adequacy of the tanning process and are carried out under predetermined humid conditions (hydrothermal), since the thermal properties of collagen depend on the water content of the material.

From the thermodynamic analysis, at the beginning of the transformation reaction of the intact procollagen triple helix structure to a disordered structure, which occurs at a certain temperature, it is observed that the activation free energy increases with the increase in the isoelectric stability of the protein, depending on the pH of the medium. According to the observation (1929) of zeit (Chater), this stability increases towards the neutral region of pH and decreases towards the extreme. Therefore, the contraction temperature increases with the increase in chemical stability of collagen. The contraction temperature is defined by Tess (1946) as "a measure of the structural stability of collagen expressed in units of temperature". Thus, a treatment that increases the chemical stability of collagen by increasing the activation free energy of collagen denaturation also increases the thermal stability of collagen, since the contraction temperature increases with the treatment.

Thermal stability can be measured in various ways; for example, the shrinkage temperature (Ts) can be determined by studying the onset of shrinkage of a strip of leather immersed in water at a predetermined rate (Δ T/T) during a gradual heating process; however, the apparent shrink temperature is dependent on the heating rate, so that as the heating rate increases, the temperature measured as the shrink becomes visible is also higher. This makes this technique somewhat imprecise, particularly when the nature of the material can alter the kinetics of the reaction.

The thermal stability of collagen materials can also be measured using Differential Scanning Calorimetry (DSC) techniques, i.e., thermodynamic studies are conducted by measuring the change in heat absorbed by a sample relative to a reference sample heated to a certain rate (Δ T/T). Thus, in this test, the temperature increases linearly as a function of time. The phase change of the material may require more or less heat to flow to the sample, depending on whether the transition is endothermic or exothermic. For example, denaturation of collagen is an endothermic process, i.e., it requires heat. Thus DSC was used to determine collagen thermal stability. Furthermore, this technique allows the study of the thermal transition under known humidity conditions. In this technique, the thermal transition can be characterized by a starting temperature, assumed to be close to the conventionally measured shrinkage temperature, and an enthalpy associated with said thermal transition, equal to the area bounded by the heat flow curve between the starting and ending temperatures of the transition versus the temperature, normalized to the energy per unit weight of dry collagen material. Through this test, various additional and valuable information can be obtained, such as: a) the contraction temperature (Ts), which reflects the chemical state of collagen; b) enthalpy of contraction (Δ H) associated with the degree of degradation or retention of native collagen structure; for any degree of denaturation of collagen, this enthalpy will be less than the enthalpy of the native state of collagen. The increase in the shrinkage temperature can be in response to an increase in chemical stability through tanning, or in response to measurements made on the dried material, but the decrease in shrinkage temperature will be due to chemical destabilization (bond cleavage) or denaturation. Entropy can also be calculated, a thermodynamic parameter that is more related to the degree of disorder of the collagen structure, but this is a rarely used parameter. However, this type of analysis requires expensive and complex equipment, which is not available in any tannery facility or leather-related company. Furthermore, although the sensitivity of this method allows the use of very few material samples, this in turn represents a limitation facing biomaterials (e.g. collagen tissue) which may be very inhomogeneous on a macroscopic level, and produces a high variation between different microscopic samples depending on, for example, the fibrillar and fibrous structure of the selected sample, or depending on the pH or the different absorption of solutes of the selected microsample.

Another alternative test related to the estimation of collagen status is the technique of measuring isometric tension under certain hydrothermal conditions, the initials "HIT" of which are known. In this technique, the leather strip is held to restrain shrinkage while the tension caused by the shrinkage force is measured. Additional information about the molecular state of the material may be obtained, but a significant difference is found between the temperature values at which the transition starts.

In the obtained tension versus temperature curve, three steps are observed: a first step of increasing the tension to a maximum value, followed by a step of relatively constant tension; the third is a step that may be a constant tension or relaxation due to gradual destruction of the collagen structure or breaking of some of the crosslinks. The slope of the curve in the tension increase phase gives the concept of fibre stiffness due to the connection; the higher the slope, the greater the number of crosslinks present in the fiber. The relaxed regions represent the stability of these bonds. The steeper the speed of the relaxation curve, the less stable the bond will be. Finally, the shape of the HIT curves can provide useful information to compare the relative density and stability of the bonds of various samples.

This interesting hydrothermal behaviour of collagen also has its practical application in the collagen casing manufacturing industry, similar to leather and leather manufacturing industries, as both are based on aspects such as chemical modification of collagen, e.g. cross-linking or cross-linking polymerization, to look for certain physical properties and microbial stability over time; but also maximally retains the natural structure of collagen, and avoids the denaturation of the collagen in the leather pretreatment process and the subsequent process.

Some examples of the use of curves of, for example, the HIT test can be cited as reference in patents for collagen membranes such as US 3894158 or in application WO2004073407a1 (Morgan); Devro (2004), in which porcine collagen membranes are described, wherein this technique has been used to detect specific differences in collagen related to the source of this material, in particular in older females than the rest of the herd intended for meat. Based on the sudden contraction (about 25% to 33% of its initial length) that mammalian collagen undergoes when contacted with water at temperatures of 60-70 ℃, if the contraction is inhibited, considerable strain will be generated by rigidly mounting the specimen. The test investigated thermal contraction of collagen by measuring the tension produced by heating at a constant rate. Since the more collagen cross-linking, the older the animal, the more energy is required to cause denaturation of the molecules than collagen in young animals, which will also be reflected in an increase in temperature contraction. Also, the slope of the curve during increasing tension reports the stiffness of the fiber caused by the bonds: the steeper the slope, the greater the number of bonds present (collagen cross-linking) and therefore the greater the age of the animal. However, it is not possible to correlate the obtained map with the progress of the thermal transition process or collagen denaturation.

Disclosure of Invention

It is an object of the present invention to provide a simple, economical and reliable device capable of providing data comparable to that obtained by calorimetric techniques such as DSC and facilitating the testing of multiple samples in the same process.

To this end, the invention proposes a device for measuring the heat shrinkage of a collagen film, comprising:

a hydrothermal vessel tub and an adjustable heater for heating a liquid bath, such as water;

a temperature probe within the vessel;

an upper and a lower gripping jaw for at least one sample of the membrane;

a first bar configured to rest on a rim of the bucket and to receive the upper grip clamp such that the one or more test samples hang from the bar and extend into the bucket;

a set of pulleys fixed to the bottom of the tub and the second rod, and at least one wire attached to each lower gripping clip to transmit the tension of the specimen to the outside of the tub through the pulley block;

a unit for measuring the displacement of the specimens, the unit being provided with a displacement sensor for each specimen, wherein each sensor is attached to each wire;

a processor provided with program means for acquiring, processing and displaying displacement and temperature data.

The device may also be provided with means for securing the upper retaining clip to the rod and the lower retaining clip to the wire, which may be a ring, for example. The rod may be circular in shape to place the sample along a circle concentric with the surface of the barrel (the barrel is circular or polygonal in cross-section) and the plane of the rod lies horizontally and parallel to the bottom of the barrel. The temperature probe may be PT100 type. The heating system may be provided with means for thermal agitation by means of magnets. The measuring unit is preferably provided with analogue measuring elements for the displacement of the sample, such as flanges and metric scales, which allow a visual reading of said displacement.

The invention also relates to a method performed with the aforementioned device, comprising the following steps:

a sample of at least one collagen membrane in the form of a rectangular strip was prepared,

immersing the sample in the bath solution for 15 minutes until the water content of the osmotic absorption is balanced;

calibrating the starting position of each sample to zero;

once the bath heating ramp was selected and the approximate initial shrinkage temperature of the collagen was known, a pre-heated bath was introduced until nearly all of the sample was covered and heating by the heater was initiated;

the digital signals caused in the sensor by the contraction of the sample and the digital signals from the temperature probe are collected by a processor and a programming device.

Drawings

To facilitate a better understanding of the characteristics of the invention and to supplement the present description, the following figures are attached as an integral part thereof, these figures being illustrative and not restrictive in nature:

fig. 1 is a detail of the preparation of the test specimens.

Figure 2 shows the apparatus of the present invention.

Fig. 3 is a schematic view of the inside of the measuring unit.

Detailed Description

The test method of the invention is based on measuring the degree of linear shrinkage induced in a series of samples of predetermined length "L" hydrated by immersion in a water bath, wherein the composition of the linear bath may also be varied (e.g. pH, addition of solutes, other compatible solvents) during the increase of its temperature to obtain a shrinkage curve versus multiplet. The sample in the form of a specimen (1) is cut from the collagen material as an elongated strip (fig. 1) so that its longitudinal axis can be parallel or transverse to the direction of manufacture of the film from which it originates.

One advantage of designing an apparatus for performing the present testing method is that it allows for the simultaneous testing of "n" samples of material or other foreign material belonging to the same batch or different production batches. Another advantage of the method is that it can be applied to any material, whether collagen from casings or leather as its raw material.

The main advantage of the present test model is that when studying the degree of denaturation of a collagen-like sample, for example, we allow us to know the mathematical derivation of the individual curves of the obtained multiple graphs of the change in shrinkage with respect to temperature (or in other words, the shrinkage at each temperature), resulting in a graph type comparable to the thermograms obtained in DSC. This allows the use of macroscopic tests, where the test conditions (e.g. macroscopic composition of the collagen fibers, pH of the medium, concentration of solutes or other solvents) can be strongly varied simultaneously in several samples.

The present invention therefore becomes a very effective tool for evaluating collagen characteristics, which can provide predictive information about the behaviour of casings manufactured as collagen films, which must overcome the mechanical requirements during their application as packaging material and the suitable behaviour during consumption (e.g. when heated). By measuring the shrinkage, it is possible to assess in particular the degree of denaturation and/or crosslinking of the fibrin collagen used as the main raw material in the packaging material.

The apparatus shown in fig. 2 comprises a hydrothermal vessel tub 2 (for heating a water bath), preferably made of glass with high heat resistance, with the ability to accommodate a set of samples placed in a vertical position. The bucket may be polygonal or cylindrical depending primarily on whether the array of samples is to be a parallel row or a circular conveyor belt. The figures show a cylindrical container and a row of five samples as an example, although the number of samples and rows may be different and the cross-section of the container may also be different.

The barrel is also equipped with a temperature probe 8, preferably but not necessarily of the PT100 type, to control the temperature ramp from the processor. The probe also allows direct reading of the temperature through a digital display in order to manually record the temperature change.

The specimen is held in a good fastening with a clamp or spring finger 4, although unlike the model based on isometric contraction tension, it does not need to penetrate the specimen because the tension it must withstand is relatively much lower. Thus, the retention of the specimen is faster and more comfortable than the measurement of an isometric tension test, which represents an advantage over other previous models.

In a preferred embodiment, the specimen is suspended at one end from a clamp so that it always remains stretched due to its own weight (preferably less than 100g) attached to the weight of the clamp at the other end (fig. 1 and 2). In this vertical position, a group of test specimens consists of one or more horizontal bars 6 from which all test specimens to be tested are suspended. The retaining clip at the upper end of the sample is screwed onto the horizontal rod by means of a ring 7 (fig. 2).

In the preceding variant, the samples are arranged on a conveyor belt (not shown in the figures); the clamping bar is thus of circumferential form, in which all the clamps forming part of the upper clamping end of the test specimen are threaded. The plane of the circumference is in a horizontal position so that all the specimens are suspended at the same height.

In this position, the upper end of the sample, once clamped by the jig, becomes a fixed position in which displacement does not occur during hydrothermal shrinkage of the sample. It is the opposite lower end that enjoys freedom of movement and will rise as the contraction of each specimen occurs.

Each suspended lower gripping clip provided with its corresponding loop 7 is fastened independently to the end of a thin flexible and light wire 9 made of a high tenacity woven fibrous material with virtually zero elasticity (less than 10% longitudinal expansion between 0 and 100 °) to avoid deformation of the displacement measurement caused by thermal contraction of the test specimen. Each braided wire attached to the suspension claw of the test specimen is led from inside to outside the bath through a small pulley 10 made of, for example, plastic (preferably nylon or teflon) and finally attached at its distal end to an analog displacement measuring element located in a cell 11 in which all terminals of the test specimen are connected in the same way. The unit contains a displacement sensor for each sample connected by wires. It also allows visual reading by the position of the flange relative to the adjacent metric scale (in millimeters).

Below the hydrothermal vessel is placed an adjustable heating system 3, which is also equipped for magnetic stirring by means of magnets. Through which heating of the liquid in the bath will be performed.

The analog measuring and converting element (fig. 3) comprises a linear variable displacement converting unit (LVDT)17, which is a tubular unit, identical to each other, provided with three electromagnetic coils, each guided by an internal rod 14 and placed vertically in a group, duplicating the sequence of the group of samples from which it comes. The vertical displacement of the vertical tubular unit relative to its shaft is converted into a signal which is recorded in the processor 13. The displacement versus temperature graph may be implemented by a sequencer.

In order to make the testing process visually visible, each tubular unit has a flange 15 which protrudes through a vertical groove 16 so that all flanges are positioned vertically at the same height, thereby creating a horizontal alignment. The length of the vertical groove through which the flange projects is always considered to be greater than the maximum contraction length of the test specimen to be tested, the length thereof between the jaws also being predetermined for the described testing device. For example, the cut specimen is 15cm long and once clamped, the free distance between the jaws is 10 cm. Under these premises, the length of the groove of the measuring device is also set to 10cm, which can be measured by a silk-screen metric scale adjacent to the groove.

The detailed process is as follows:

1) cutting "n" samples into rectangular strips having a length of 15cm and a width of 2cm in the machine direction;

2) immersing the sample in the bath solution for 15 minutes until the water content of the osmotic absorption is balanced;

3) the strips bundled by the jaws are placed according to the schematic of fig. 1 and the cells are supported by horizontal rods in the mouth of the hydrothermal vessel;

4) calibrating the starting position of each sample to zero in the simulation apparatus by the programmer and the processor;

5) once the heating ramp of the bath has been selected, for example 2 ℃/min, and since the approximate initial shrinkage temperature of the collagen is known, water preheated from the bath, for example to 55 ℃, is introduced to cover almost all of the sample, and it begins to be heated by the heater;

6) the contraction of the specimen causes a displacement of the sensor element and the flange of the transducer unit and the digital signal of the device together with the digital signal from the temperature probe is received and integrated by the processor where a graph with recorded data will be generated, e.g. a graph of the ordinate L against the temperature, or other graphs, e.g. the derivative dL/dT of the ordinate against the temperature.

In view of this description and the accompanying drawings, it will be understood by those skilled in the art that the present invention has been described in terms of some preferred embodiments thereof, but that various changes may be introduced therein without departing from the subject matter of the invention as claimed.