The LP system can be used to prevent bacterial deterioration of milk when refrigeration is not available. It can also be used to prolong the safe storage life of refrigerated milk. Arguably the LP-system, immunoglobulins and lactoferrin have potential to be of value in neonate nutrition. The remaining section largely concerns the exploitation of the LP-system in the protection of neonates.
Manufacture of milk powders containing a functional LP system
That milk provides neonates with nutrients and various protective antimicrobial factors has been discussed previously. Because many of these factors are denatured by the heat treatments used in milk replacer manufacture commercial products, unless specially produced, generally do not contain antimicrobial proteins in active form.
The author and colleagues at the Scottish Agricultural College (SAC) at Auchincruive developed milk replacers containing a functional LP system, in powder form, stable at ambient temperature for at least five months and suitable for use on the farm. These were the result of a collaborative project between SAC, the former Scottish Milk Marketing Board (SMMB) and the former Swedish animal nutrition company Astra-Ewos.
Milk replacers containing active antimicrobial proteins can be produced by two main methods. Method 1 involves careful control of time-temperature treatments employed during replacer manufacture to avoid denaturation of the antimicrobial agent. In the second, the agent is extracted from milk or whey e.g. lactoferrin, and is dry-mixed with other ingredients to give a final milk replacer.
Calf milk replacers
Conventional calf milk replacers
Production of skim milk powder containing high levels of LP activity
Raw milk contains high but variable levels of LP activity (e.g. Korhonen et al., 1977). LP activity equivalent to enzyme protein concentrations between 20-40 mg/L is found frequently. These are markedly in excess of the levels (0.5 μg/ml in absence of catalase) required for efficient LP system activation (Björck, 1978). Powders of high LP activity can be produced by carefully controlling the temperatures at key stages during the drying process. The temperatures used for pasteurisation, and preheating prior to evaporation are the major critical control points as far as conservation of LP activity is concerned. The author has found that if temperatures at these points should exceed 75°C that this will result in significant loss of LP activity. Another activity critical control point is the temperature of the concentrate in the 'temperature controlled" jacketed pipe going to the atomiser.
Low heat skim milk powder can easily be produced commercially, using modern plant, containing approximately 50% of the LP activity of raw milk. Powder of high LP content has a non-casein nitrogen/total nitrogen ratio of >0.2 and a heat or casein number of <80.
Selection of hydrogen peroxide donor or hydrogen peroxide generating system
The LP system in milk replacers has been activated by addition of hydrogen peroxide in liquid form, generation of hydrogen peroxide in situ by use of glucose oxidase and by the hydrolysis of magnesium peroxide. The author and colleagues have also investigated other methods for example the use of hydrogen peroxide producing lactic acid bacteria, for LP system activation.
The characteristics of an ideal hydrogen peroxide donor are shown in table 1. Magnesium peroxide meets most of these criteria. This material is available commercially as a mixture containing Mg (OH) 2 (75% w/w) and Mg02 (25% w/w). Hydrolysis of this compound in milk is pH dependant; hydrogen peroxide production requires the pH to be lowered to 6 or less. Consequently replacers containing magnesium peroxide produce hydrogen peroxide in the abomasium of the calf. The use of magnesium peroxide for LP system activation is described in British Patent No. 1546747. Optimal LP system activation requires magnesium peroxide to be added to milk replacers to as to give a concentration of 0.03% (w/v) in the reconstituted replacer.
Table 1: Characteristics of a hydrogen peroxide donor for use in milk replacer formulation
1. It should be available in powder form. |
2. It should be un-reactive with replacer components during storage. |
3. It should release hydrogen peroxide at a controlled rate in the reconstituted replacer. |
4. It should be relatively inexpensive. |
5. It should be non-toxic and have no detrimental effect on the palatability of the replacer |
Sodium thiocyanate addition
Extenders
The composition of a typical LP milk replacer is shown in table 2.
Table2. Typical composition (1) of a calf milk replacer containing an |
|
Ingredient |
% w/w |
Low heat skim milk powder |
31 |
Fat filled milk powder (36-38% fat) |
53 |
Low heat whey powder |
13 |
Mineral and vitamin premix (2) |
0.65 |
Starch (3) |
2 |
1. Figures have been 'rounded' and do not add up to 100%. |
Anti-microbial activity of LP-containing milk replacers
Reconstituted milk replacer, adjusted to pH 5.8 (to hydrolyse the magnesium peroxide and release hydrogen peroxide) has been shown to be bactericidal to a range of pathogens (Mullan et al., 1982b). Typical data showing the effectiveness of the system against presumptive pathogens isolated from calves at post mortem by staff at the veterinary laboratory at the SAC at Auchincruive are shown in table 3.
Table 3. Effect of the LP system in calf milk replacers on the viability of strains of E. coli isolated from post-mortem examination of calves |
||||
Strain |
T0 Inoculum (CFU/ml) |
CFU/ml after 2 hours at 37 °C |
||
No cysteine added |
+ 1 mM cysteine |
% Kill |
||
1 |
1.3 x 107 |
3.3 x 106 |
6.84 x 107 |
74.6 |
8 |
3.9 x 106 |
2.39 x 105 |
5.84 x 107 |
93.9 |
9 |
1.49 x 107 |
4.2 x 106 |
5.58 x 107 |
71.8 |
1.72 x 107 |
1.1 x 106 |
6.72 x 107 |
93.6 |
|
NCDO 2328 |
9.5 x 106 |
3.2 x 102 |
2.68 x 107 |
99.99 |
NCDO 904 |
3.1 x 107 |
7.1 x 106 |
5.84 x 107 |
76.87 |
2 |
7.0 x 106 |
3.5 x 105 |
7.2 x 106 |
50 |
3 |
1.0 x 107 |
2.6 x 106 |
1.03 x 109 |
74 |
4 |
6.0 x 107 |
3.0 x 107 |
1.45 x 109 |
50 |
5 |
7.6 x 106 |
1.3 x 106 |
5.1 x 106 |
83 |
6 |
6.0 x 107 |
7.0 x 105 |
7.5 x 108 |
98.8 |
7 |
3.0 x 107 |
2.0 x 105 |
5.6 x 108 |
99.3 |
Notes: Milk replacer was reconstituted to give 10% TS and was adjusted to pH 5.8. Reconstituted replacer contained 0.03% (w/w) Mg02, LP activity equivalent to 7 μg LP/ml and 0.35 mM SCN. E. coli strains were isolated from lesions in calves at post-mortem and were presumed to be calf pathogens. Mullan and Waterhouse, unpublished data. |
Shelf life of lactoperoxidase-containing calf-milk replacers
There can be marked differences in the decline of enzyme activity in milk powders stored at ambient temperatures. LP concentrations in powders after 5 months storage generally ranged from 0.05 — 0.5 ug/ml of the reconstituted replacer. While 0.05 ug/ml of LP is too low to give optimal bactericidal activity it is high enough to reduce a population of 1 x 107 CFU/ml of a sensitive E. coli by up to 3 log cycles (Mullan and Waterhouse, unpublished data).
Quality assurance of LP containing milk replacers
Some research on the use of immunoglobulin concentrates in calf feeding has been published. The limited data suggest that inclusion of non-specific antibodies in milk replacers does not improve calf health or performance (De Gregorio and Anexstad, 1991) but suggest that concentrates containing antibodies specific for pathogens may have positive effects (Annexstad et al., 1991).
Improved human milk replacers
Bovine immunoglobulins possess both antibacterial and antiviral activity, e.g. antirotavirus activity (Goldman et al., 1985). There is some research interest in the isolation of immunoglobulins from milk e.g. cheese whey and the addition of immunoglobulin concentrates to human milk replacers. If these antimicrobial proteins are to be effective then a local immune response within the gastrointestinal tract of the infant must be considered as the mechanism responsible; this remains to be demonstrated.
Acknowledgements
The LP project at the WSAC at Auchincruive (now SRUC) was a collaborative project involving the Dairy Technology, Animal Husbandry and Veterinary departments, the Scottish Milk Marketing Board and Astra-Ewos. Please see the acknowledgement's section.
How to cite this article
Mullan, W.M.A. (2003) .
[On-line]. Available from: https://www.dairyscience.info/index.php/exploitation-of-anti-microbial-proteins/168-lactoperoxidase-system.html?tmpl=component&print=1&layout=default&page= . Accessed: 19 April, 2024.
Revised August 2009.