Comparison of the Efficacies of Gaseous Ozone and Sodium Hypochlorite in Cleaning Stainless Steel Particles Fouled with Proteins

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Comparison of the Efficacies of Gaseous Ozone and Sodium Hypochlorite in Cleaning Stainless Steel Particles Fouled with Proteins Biocontrol Science, 2003, Vol. 8, No. 2, 87-91 Note Comparison of the Efficacies of Gaseous Ozone and Sodium Hypochlorite in Cleaning Stainless Steel Particles Fouled with Proteins KAZUHIRO TAKAHASHI', KUNIHIKO KOIKE2, AND SATOSHI FUKUZAKI1* Industrial Technology Center of Okayama Prefecture,1 5301 Haga, Okayama 701-1296, and 2Shiga Technology Center, lwatani International Corp., 4-5-1 Katsube, Moriyama, Shiga 524-0041, Japan Received 16 July 2002/Accepted 21 September 2002 The efficacies of gaseous ozone and sodium hypochlorite as an oxidizing agent for cleaning were compared in a laboratory model system using stainless steel particles fouled with pro- teins. Exposure of the protein-fouled stainless steel particles to 0.5% (v/v) gaseous ozone for 30 min enhanced the removal of the various proteins during subsequent alkali cleaning with NaOH solution to the same degree as the NaOH solution containing sodium hypochlorite at 0.2 to 0.4 g/l. The effect of ozone pretreatment on the removal of the proteins depended on the concentration of gaseous ozone. Pretreatment with a highly concentrated gaseous ozone of 20% for 30 min resulted in the almost complete removal of the proteins from stainless steel particles. Key words : Cleaning/Gaseous ozone/Hypochlorite/Protein/Stainless steel. In the food industry, the cleanliness and sterility of 1972). There are many advantages of using ozone as food-processing equipment is one of the most impor- an oxidizing agent in the food industry. Ozone is a tant factors in manufacturing wholesome and safe more powerful oxidant than chlorine and can kill a products. The combined use of detergent and chlo- large number of microorganisms (Graham, 1997; Kim, rine such as sodium hypochlorite has been practiced et al. 1999; Moore, et al. 2000). In addition, ozone can extensively in the dairy industry (Clegg, 1962). The be applied as a gas or in water, and it leaves no resid- main action of hypochlorite is due to the oxidation of ual toxicity after decomposition to oxygen. However, organic substances of soils and microorganisms. few published reports are available on the possible However, the repeated use of chlorinated alkaline de- use of gaseous ozone in the cleaning process. tergent results in the abrasion and corrosion of metal- The aim of this work is to compare the efficacy of lic materials. In addition, chlorination brings health gaseous ozone pretreatment with that of sodium hazards to human beings and wildlife through the pro- hypochlorite in the cleaning of stainless steel particles duction of harmful compounds in the environment fouled with proteins that have different properties, (Richardson et al., 1998). Therefore, alternative e.g., thermal stability, solubility, and structure. cleaning agents to chlorine are being explored for Nonporous stainless steel particles (SUS 304: 2-10 food-processing equipment. One such compound is m) were obtained from The Nilaco Corp. (Tokyo);u ozone (O3) which has been utilized as a sanitizing their specific surface area, as determined by BET agent in European water treatment plants (Gomella, method, was 0.43 m2/g. Crystalline BSA and ƒÀ -lacto- globulin ( ƒÀ -Lg) were purchased from Nacalai *Corresponding author . Tel : +81-82-286-9600, Fax:+ 81-82- Tesque Inc. (Kyoto). Casein and gelatin were pur- 286-9630 chased from Kanto Chemical Co., Inc. (Tokyo) and 88 K. TAKAHASHI ET AL. Wako Pure Chemical Ind. Ltd. (Osaka), respectively. The model protein-fouled stainless steel particles were prepared at 40•Ž by introducing a 50-ml aliquot of a protein solution (3g/l, 10-3 M KNO3) and 10 g of stainless steel particles into a 125-ml glass vial, which was then laid on its side in a water bath at 40 and reciprocally shaken (140 oscillations per min)•Ž for 2 h. The pH values of the protein solutions were adjusted to 5.6 for BSA and gelatin (at the point of zero charge) , and 6.8 for ƒÀ -Lg and casein (pH of milk). After being shaken, the protein-fouled stainless steel particles were collected and washed with 40 ml of 10-3M KNO3 solution by centrifugation (2300 •~g for 1 min). The washed protein-fouled particles were dried at 40•Ž for 16h and stored in a desiccator be- fore use for the cleaning experiments. The amounts of BSA, gelatin, ƒÀ -Lg, and casein adsorbed were 2.7, 1.8, 1.6, and 2.7 mg/m2, respectively. Gaseous ozone of 0.5 to 6.0% (v/v) was gener- ated from pure oxygen (99.999%, v/v) by a silent discharge ozonizer (M001; OHNIT Co. Ltd., Okayama) equipped with an ozone monitor (PG-620; Ebara Jitsugyo Co. Ltd., Tokyo). The above gaseous FIG. 1 . Effects of ozone pretreatment (0.5% for 30 min) ozone was concentrated to 15 and 20% using the and sodium hypochlorite (0.2 g AC//) on the removal of adsorption-desorption technique on silica gel (Koike proteins from stainless steel particles during cleaning with NaOH solutions of different pHs at 40•Ž. (A) BSA, (B) et al., 2000). Prior to cleaning, a 5-g portion of -Lg , (C) casein, (D) gelatin. Batchwise cleaning experi-ƒÀ protein-fouled stainless steel particles was treated ment was conducted at 40•Ž for 2 h with shaking (140 oscil- with gaseous ozone of 0.5 to 20% at room tempera- lations per min). Symbols: A, NaOH alone; O, ozone ture for 30 min. pretreatment; ^, sodium hypochlorite. Batchwise cleaning was conducted by introducing a 1.16-g portion of protein-fouled stainless steel parti- stainless steel particles, and that of the protein in so- cles (ca. 0.5m2) and 5 ml of cleaning solution into a lution were measured with a total organic carbon ana- 25-ml glass vial, which was then placed horizontally in lyzer (TOC-5000; Shimadzu Co., Kyoto) equipped a water bath (40 to 80•Ž) and reciprocally shaken with a solid sample module (SSM-5000; Shimadzu (140 oscillations per min) for 2 h. The pH values of Co., Kyoto). cleaning solutions were adjusted to pH 9.0 to 13.5 Figure 1 shows the effects of gaseous ozone with NaOH (1 M solution). After each cleaning experi- pretreatment (0.5% for 30 min) and sodium ment, the particles bearing the remaining proteins hypochlorite (0.2 g AC/l) on the removal of the pro- were collected by centrifugation (2300 xg for 1 min). teins when used in conjunction with the NaOH solu- To determine the effects of oxidizing agents, protein- tions of different pHs at 40•Ž. In the case of cleaning fouled particles were cleaned at 40•Ž for 2 h with with NaOH solution alone, the removal efficiency was NaOH solutions of pH 9.0 to 13.5 with and without so- markedly higher at pHs above 11 and increased with dium hypochlorite at a final concentration of 0.2 g increasing pH. This indicates that protein removal de- available chlorine (AC) per liter, respectively, as de- pends on the hydroxide ion concentration (Koopal, scribed above. Continuous cleaning was conducted at 1985; Takehara et al., 2001). It was noted that the re- 40•Ž in a stainless steel column (4 mm ƒ³ •~ 50 mm) moval of the proteins occurred markedly at the alka- by feeding 10-3 M KNO3 solution (pH 5.6) for 120 line pH region above the intrinsic dissociation min followed by feeding NaOH solution of pH 12 with constant (pKint) of a side-chain amino group of, for in- or without sodium hypochlorite (0.4g AC/l) for 180 stance, lysine, i.e., 10.8. The use of sodium min at a flow rate of 0.25 ml / min (Urano and hypochlorite and ozone significantly enhanced the re- Fukuzaki, 2001). The alkali eluate was fractionally moval of the proteins. The cleaning effects of ozone collected, and its protein concentration was meas- pretreatment against all the proteins were comparable ured. to those of alkaline hypochlorite solution at 0.2 g AC The amount of protein adsorbed or remaining on /l although small differences in the effects on various CLEANING EFFICACY OF GASEOUS OZONE 89 proteins were observed. The performance of hypochlorite and ozone in removing proteins is asso- ciated with the decomposition of proteins by their strong oxidative power (Merrill et al., 1962; Urano and Fukuzaki, 2001). As a result of decomposition, large fractions of the proteins could be removed even when using a low-pH NaOH solution. It is known that the effects of the combination of NaOH solution and temperature are also to solubilize adsorbed proteins through the chemical reaction be- tween NaOH and proteins, i.e., hydrolysis of proteins (Twomey, 1968). Figure 2 shows the effect of tem- perature on the protein cleaning efficiency by the NaOH solutions of different pHs. All the proteins were removed more efficiently by increasing the tempera- ture from 20 to 80•Ž at pHs especially above 12. Although BSA and p -Lg, which contain sulfhydryl ( SH) groups and are respectively denatured at tem- peratures above 60•Ž and 70•Ž at a neutral pH (De Wit and Swinkels, 1980; Steim, 1965), high tempera- tures under strongly alkaline conditions (pH>12) can enhance the removal of proteins irrespective of the Cleaning time (min) thermal stability of the proteins. As shown in Figs. 1 and 2, in the case of cleaning with a low-pH NaOH so- FIG.3. Time courses for the removal of proteins from stain- lution below 11, the effects of ozone and sodium less steel particles during continuous cleaning with the hypochlorite on the removal of the proteins were NaOH solution of pH 12 alone, NaOH solution (pH 12) com- bined with ozone pretreatment (0.5% for 30 min), or NaOH solution (pH 12) with hypochlorite of 0.4g AC/i.
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