Use of polyether ionophore antibiotics to control bacterial growth in sugar production
BACKGROUND OF THE INVENTION 
     This invention relates to the use of polyether ionophore antibiotics to control bacterial growth during sugar (sucrose) production. It can be used with a wide variety of feedstocks such as sugar beet juice, sugar cane juice, hydrolyzed grain (e.g., corn or wheat) or any other starch or sugar-containing material that can be used to produce simple sugars. 
     One of the key steps in sugar production is an extraction process where feedstock such as sugar beets or sugar cane is treated to extract sugar (as an aqueous solution referred to herein as &#34;sweet juice`˜) from the plant material. For instance, in the case of sugar beets, a diffusion process is commonly employed where the beets are soaked in warm water. This is typically-performed at about 70° C. under acid conditions (pH around 6) for a period of 1 to 2 hours. During that time, heat-tolerant bacteria can proliferate, feeding on sugar and thus reducing the amount that can ultimately be recovered and marketed. This negatively impacts plant productivity and is a significant problem for the industry. Sugar cane is commonly subjected to an extraction process involving milling in which similar problems are encountered. 
     The microcrganisms causing the problem are mostly gram positive bacteria that belong to the lactobacillus genus. Streptococcus, bacillus., clostridium, leuconostoc and pediococcus may also be present. In the past, formaldehyde has been used in an attempt to control bacterial growth, but this raises serious safety concerns. 
     This invention concerns a method for the production of sugar wherein a polyether ionophore antibiotic such as monensin, narasin, salinomycin, lasalocid, maduramycin or semduramycin is used to control or supress bacterial growth during the process. These compounds have good activity against gram positive bacteria and do not easily degrade over time or under high temperatures. This makes them attractive to the sugar industry because: 
     1. they remain active for many days under typical sugar plant operating conditions; and 
     2. they remain active at the high temperatures and acid pH used in the extraction step. 
     The bacterial population in the extraction bath is greatly reduced by the addition of a bacteriostatic or bactericidal concentration, for example 0.5 to 3.0 ppm, preferably 0.5 to 1.5 ppm, of a polyether ionophore such as monensin. This control greatly reduces the bacterial consumption of sugar leading to a significant improvement in plant productivity. Surprisingly, there are no detectable polyether residues in the final white sugar crystals. This result is particularly important because it makes the invention suitable for manufacture of food grade, white sugar crystals. 
     1. Field of the Invention 
     THE KEY STEPS IN SUGAR PRODUCTION 
     The 4 main steps performed in a typical sugar plant are described hereafter. 
     EXTRACTION 
     The purpose of this step is to extract the sugar from the feedstock. It yields a sweet juice with a pH of about 6 that is very susceptible to bacterial contamination. It also extracts water-soluble substances such as proteins which must be removed from the medium since they can hinder sugar crystallization. 
     PURIFICATION 
     Its purpose is to eliminate organic substances extracted with the sugar. It consists in adding a mixture of lime and water to the sweet juice and then sending through a flow of carbon dioxide to precipitate calcium as calcium carbonate. After filtration, one gets a clear juice, with little organic content other than sucrose. 
     CONCENTRATION 
     This clear juice, which is about 14% sugar, is heated and concentrated into a syrup with a sugar content comprised between 60% and 70% by weight. 
     CRYSTALLIZATION 
     This last step yields white sugar and a byproduct, molasses. It consists in concentrating further the syrup at 85° C. under vacuum to bring it beyond the saturation point of sucrose (in a state called &#34;supersaturation&#34;). Then, one introduces a small amount of sugar crystals (about 0.5 g) to trigger crystallization which spreads rapidly through the liquid, turning it into a mass of white sugar crystals bathing in a syrup colored by impurities. The white sugar crystals are separated by centrifugation, rinsed and dried. 
     This crystallization step is repeated twice on the non-crystallized syrup coming out of the centrifuge. The second and third time, it yields brown sugar that is not marketed. Instead, it is reinjected at the beginning of the crystallization phase with the syrup coming from the evaporation step to yield more valuable white sugar. Only white sugar is marketed. 
     After the third iteration, the dark, noncrystallized juice has become molasses. It contains about 50% sugar and 30% foreign matter that prevents further crystallization. 
     2. Description of a Preferred Embodiment 
    
    
     BRIEF DESCRIPTION OF THE DRAWING 
     The drawing depicts an example of a plant that processes 500 tons of sugar beets per hour. 
    
    
     DETAILED DISCLOSURE OF THE METHOD FOR THE PRODUCTION OF SUGAR 
     The process is described for a plant treating 500 tons of sugar beets per hour. 
     1. EXTRACTION 
     Operating conditions: 
     Temperature: 70° C.; pH=6; duration: 1-2 hours; process: continuous. 
     The extraction process uses a conveyor immersed in water. It is fed at one end with chopped beets and at the other with warm water to which various sugar-rich residues have been added for recycling. The beets move against the flow of water, so their sugar concentration declines as that of the water increases. 
     Sweet juice containing about 14% sugar (plus water-soluble proteins and other impurities) runs off from the end where fresh beets are added to the conveyor while spent beets (pulp) are evacuated from the other end. The 500 tons of beets processed per hour yield about 500 m 3  of sweet juice and 500 tons of pulp. 
     2. PURIFICATION 
     Operating conditions: 
     Temperature: 75° C.; pH=8.5; duration: 1 hour; process: continuous. 
     The sweet juice from the extraction step is passed into a vat where it is mixed with an aqueous suspension of lime (200 g of CaO per liter). A stream of carbon dioxide is blown into the vat causing calcium carbonate to precipitate taking along large molecules such as proteins that can interfere with crystallization. 
     The 500 m 3  of sweet juice processed per hour use about 30 m 3  of aqueous-lime suspension and yield about 500 m 3  of purified sweet juice. 
     3. CONCENTRATION 
     Operating conditions: 
     Temperature: declining from 130° C. to 85° C.; pH=8.5; process: continuous. 
     The purified sweet juice is boiled down. The 500 m 3  of sweet juice (14-16% sugar) processed per hour yield 110 m 3  of concentrated syrup (60-70% sugar). 
     4. CRYSTALLIZATION 
     The 100 m 3  of concentrated syrup are run through the various phases of the crystallization step during which another 106 m 3  of water are evaporated off. Finally, one ends up with 60 tons per hour of white sugar and 20 tons of molasses with a 50% sugar concentration. 
     KEY PROPERTIES OF POLYETHER IONOPHORE ANTIBIOTICS 
     Experiments were conducted with several polyether ionophore antibiotics such as monensin, lasalocid and salinomycin using sweet juice extracted from sugar beets. These experiments confirmed the existence of bacteriostatic and bactericidal concentrations which, for these molecules, can be as low as 0.5 ppm to 3.0 ppm. At bacteriostatic concentrations, the growth of the bacterial population is inhibited. At bactericidal concentrations, the bacterial population drops. 
     We also did sensitivity testing showing that polyether ionophore antibiotics are active against most bacteria commonly encountered in sugar plants. For instance. Table 1 shows the reduction in bacterial count observed 6 hours after treatment with 3.0 ppm of monensin. 
     
                       TABLE 1                                                     
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The impact of monensin on the bacterial                                   
count of various microorganisms                                           
              BACTERIAL                                                   
              COUNT                                                       
              at t = O                                                    
                     at t = 6h                                            
                              Pct reduct.                                 
______________________________________                                    
Lactobacillus plantarium                                                  
                1.2 × 10.sup.8                                      
                         4.1 × 10.sup.5                             
                                  -99.70                                  
Lactobacillus fermentum                                                   
                6.2 × 10.sup.8                                      
                         4.4 × 10.sup.4                             
                                  -99.99                                  
Lactobacillus vaccimostercus                                              
                2.8 × 10.sup.8                                      
                         2.1 × 10.sup.5                             
                                  -99.90                                  
Lactobacillus buchneri                                                    
                5.5 × 10.sup.8                                      
                         3.0 × 10.sup.3                             
                                  -99.99                                  
Lactobacillus yamanashiensis                                              
                1.8 × 10.sup.5                                      
                         4.6 × 10.sup.4                             
                                  -74.40                                  
Lactobacillus coryniformis                                                
                3.7 × 10.sup.8                                      
                         3.3 × 10.sup.6                             
                                  -99.10                                  
Leuconostoc mesenteroides                                                 
                8.0 × 10.sup.5                                      
                         5.4 × 10.sup.3                             
                                  -99.30                                  
Leuconostoc acidilactici                                                  
                8.2 × 10.sup.8                                      
                         3.7 × 10.sup.8                             
                                  -54.90                                  
Bacillus subtills                                                         
                3.1 × 10.sup.5                                      
                         5.5 × 10.sup.4                             
                                  -82.30                                  
Bacillus brevis 3.3 × 10.sup.8                                      
                         6.0 × 10.sup.3                             
                                  -99.99                                  
Bacillus megaterium                                                       
                1.3 × 10.sup.8                                      
                         5.8 × 10.sup.7                             
                                  -55.40                                  
Bacillus coagulans                                                        
                1.1 × 10.sup.5                                      
                         6.1 × 10.sup.4                             
                                  -44.60                                  
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     In addition, we observed that polyether ionophore antibiotics are stable at temperatures of about 70° C. and a pH of about 6, i.e. conditions similar to those encountered in extraction baths. They are thus active under normal plant operating conditions. They degrade partly however, at the higher temperatures encountered downstream from extraction, which helps to produce white sugar crystals free of monensin residues. 
     RESIDUE ANALYSIS 
     To ascertain that white sugar crystals were free of monensin residues, Trials were conducted with the help of the French Sugar Research Institute, an industry-funded research organization. All monensin assays were done by the European Institute for the Environment located in Bordeaux, France, a well-known independent lab using the officially approved assay method (H.P.L.C.). 
     MONENSIN IN THE PURIFICATION PHASE 
     A master solution of monensin was first prepared by dissolving monensin crystals in 96% alcohol to reach a concentration of 20 g of monensin per liter of solution. Part of this solution was further diluted with water down to a concentration of 150 mg of monensin per liter. This was then used to supplement the sweet juice from extraction. Three different trials were made using varying concentrations of monensin in the sweet juice, i.e., 0.5 ppm, 1.0 ppm and 1.5 ppm. 
     The monensin-supplemented juice was then subjected to a typical purification step. Samples of 500 ml of filtered, purified juice were taken from the output stream immediately after filtration. They were assayed using the officially approved H.P.L.C. method. Results are summarized in the table below. They show that nearly 90% of monensin is eliminated by the purification step. This is understandable given monensints affinity for positive ions: it combines with calcium ion and is eliminated with it. 
     
         ______________________________________                                    
                       Percent of                                         
                       monensin                                           
MONENSIN CONTENT       eliminated by                                      
before purification                                                       
              after purification                                          
                           purification                                   
______________________________________                                    
0.5           &lt;0.1         &gt;80                                            
1.0           0.13         87                                             
1.5           0.17         89                                             
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     MONENSIN IN THE CONCENTRATION PHASE 
     Purified juice from the purification step was first standardized to 14.7% dry matter by addition of distilled water. This standardized juice was then treated with 1.5 ppm of monensin using the 150 mg/l dilute alcohol solution prepared in the extraction step. The monensin containing juice was first heated to 120° C. for 10 minutes. The temperature was then lowered to 100° C. until the dry matter concentration reached about 61%. The syrup was assayed by H.P.L.C. and a monensin content of 2.2 ppm was measured. 
     This was less than could have been expected from the mere concentration of the juice. Indeed that concentration should have raised the monensin content from 1.5 ppm to 6.2 ppm. Since only 2.2 ppm were found, it means that the difference, i.e., 4 ppm or 64% of the original quantity introduced at the start of the experiment, was destroyed by heat. 
     MONENSIN IN THE CRYSTALLIZATION PHASE 
     Syrup from the concentration step was supplemented with 1.5 ppm of monensin using the dilute monensin-alcohol solution (150 mg/l ) prepared in the extraction step. After the white sugar was crystallized, rinsed and dried, both the sugar and the remaining non-crystallized molasses were assayed. Results show: 
     No detectable amount of monensin in the white sugar (assay sensitivity: 0.5 ppm) 
     1.5 ppm in the remaining non-crystallized molasses 
     This shows that monensin stays in the liquid and that the rinsed white sugar crystals are free of monensin. Monensin ends up in the molasses instead. 
     ECONOMIC BENEFITS 
     The normal bacterial count in the extraction bath of a sugar plant is about 10 5  to 10 6  organisms/ml. Concern starts building above that and the contamination becomes significant when it reaches 10 9  /ml. These bacteria feed on sugar and lower the amount that is eventually recovered. 
     The chart here-below illustrates what happens when 1.5 ppm of monensin is introduced into the extraction juice. Most of it is destroyed along the way. The rest ends up in the molasses at a concentration of 2.6 ppm. 
     These calculations, however, assume a continuous usage of monensin in sugar production. In practice, the management of bacterial contamination require that the juice from extraction need only be treated one day per week to bring the bacterial count down to the no-problem zone until the next treatment. Under these conditions, the average monensin concentration in molasses would be 0.4 ppm. 
     This compares favorably with the 30 ppm of monensin that are commonly used to supplement beef feed rations and should not interfere with the use of molasses as animal feed.