Partial purification and characterization of protease extracted from kinema.

Proteases are large group of highly demanded enzymes having huge application in food and pharmaceutical industries. Numerous sources, including plants, microorganisms, and animals, can be used to obtain protease. Due to its affordability and safety consideration, fermented foods have recently attracted more attention as a source of microbial protease. The present study aimed to extract protease from kinema, partially purify the extracted protease following dialysis after precipitation with ammonium sulfate, and determine general characteristics of protease. The kinema having highest proteolysis activity after three days of control fermentation (Temperature 30±2 °C, RH 66 ± 2%) was taken for the study. About 2.45 fold of purification with overall recovery of 63.21% was achieved after precipitation with ammonium sulfate at 30-70% saturation level followed by dialysis of crude extracted protease. The dialysed kinema protease had specific activity of 7.90 U/mg. The enzyme remained actively functional across a wider pH (5-9) and temperature (40-60 °C) range. SDS-PAGE and Zymogram confirmed the presence of three major active bands respectively of 29.04 kDa, 36.09 kDa and 46.35 kDa in the kinema protease extract. The enzyme kinetics data on casein, fitted to Mechaelis Mentens' plots showed the protease had Vmax of 1.001 U/ml with corresponding Km value of 0.825 mg/ml. Metal ions such as iron, mercury and aluminium showed the inhibition effect whereas presence of sodium, zinc, and calcium shows the activation effect on protease performance. The enzyme was active over various natural substrates; showing maximal activity on casein, and subsequent to bovine serum albumin, gelatin, hemoglobin and whey protein respectively. Furthermore, molecular weight distribution of the protease extract and activity inhibition with ethylenediaminetetraacetic acid and phenylmethylsulfonyl fluoride, suggesting the protease from kinema could be a metal dependent serine protease or mixture of them.

Efficient three phase partitioning of actinidin from kiwifruit (Actinidia deliciosa) and its characterization.

Three phase partitioning (TPP) method was effectively utilized for the extraction and purification of milk clotting protease (actinidin) from the kiwifruit pulp. The different purification parameters of TPP such as ammonium sulfate saturation, ratio of the crude kiwifruit extract to tert-butanol, and the pH value of extract were optimized. The 40% (w/v) salt saturation having 1.0:0.75 (v/v) ratio of crude kiwifruit extract to tert-butanol at 6.0 pH value exhibited 3.14 purification fold along with 142.27% recovery, and the protease was concentrated exclusively at intermediate phase (IP). This fraction showed milk-clotting activity (MCA), but there was no such activity in lower aqueous phase (AP). The enzyme molecular weight was found to be 24 kDa from Tricine SDS-PAGE analysis. Recovered protease demonstrated greater stability at pH 7.0 and temperature 50 °C. The Vmax and Km values were 121.9 U/ml and 3.2 mg/ml respectively. Its cysteine nature was demonstrated by inhibition studies. This study highlighted that the TPP is an economic and effective method for extraction and purification of actinidin from kiwifruit, and it could be used as a vegetable coagulant for cheesemaking.

de Quervain's Disease among Patients Visiting the Orthopaedic Outpatient Department of Tertiary Care Center: A Descriptive Cross- sectional Study.

Introduction: de Quervain's disease is one of the common causes of wrist pain. It can cause serious disability and absence from work due to impaired functioning of the wrist and hand. The aim of this study is to find out the prevalence of de Quervain's disease among patients visiting the orthopaedic outpatient department of a tertiary care centre. Methods: This was a descriptive cross-sectional study conducted among patients visiting the orthopaedic outpatient department of a tertiary care centre after receiving ethical approval (IRC KAHS Reference: 078/079/56). This study was conducted from 1 January 2021 to 30 December 2021 from hospital medical records. A convenience sampling method was used. Patients with de Quervain's disease from 16 to 60 years were included in this study. Clinically diagnosis of de Quervain's disease was based on the tenderness of the radial styloid process, tenderness over the first extensor compartment on resisted thumb abduction or extension and positive Finkelstein test. Point estimate and 95% Confidence Interval were calculated. Results: Out of 9600 orthopaedic outpatients, de Quervain's disease was seen in 128 (1.33%) (2.68-4.52, 95% Confidence Interval). Conclusions: The prevalence of de Quervain's disease was similar when compared to other studies conducted in similar settings. Keywords: de Quervain's disease; surgery; tenosynovitis.

Arthroscopy-Assisted All-Suture Coracoclavicular and Acromioclavicular Joint Stabilization in Acute Acromioclavicular Joint Injuries.

The existing literature agrees on surgical management for Rockwood grade IV and V injuries, but there is no consensus which type of surgery is the most appropriate one. More than 150 surgeries have been described for this condition in the literature. In an injury of less than 3 weeks, most surgeons prefer suture-button devices for coracoclavicular stabilization. Recent biomechanical studies have demonstrated that coracoclavicular stabilization provides good vertical stability but poor horizontal stability of acromioclavicular joint. Hence, they recommend acromioclavicular stabilization along with coracoclavicular stabilization. The use of a suture-button device for coracoclavicular stabilization requires special implants and instruments along with high surgical skills to drill precisely placed holes in the clavicle and the coracoid in order to avoid fractures from the drill hole. Due to relatively smaller clavicle and coracoid in Asian population, making holes in the clavicle and the coracoid has increased risk of fracture. We describe a technique in which no drill holes are made in the clavicle or the coracoid. In our technique, suture tape is used, which is looped around the coracoid and the clavicle, and the limbs are tied over the clavicle to maintain the coracoclavicular distance. The remaining limbs of suture tape is further looped through the tunnels made in acromion and tied over the acromion to augment the acromioclavicular ligament. The potential advantages of this technique are no costly implant and instruments are required, avoidance of complications associated with drill holes in coracoid and clavicle, both coracoclavicular and acromioclavicular joints are stabilized, direct repair of the acromioclavicular ligament can be performed, and no need of second surgery for implant removal.

Outcome of Neglected Elbow Dislocation Treated with Open Reduction at Rural Tertiary Care Hospital in Nepal.

BACKGROUND: Chronic unreduced dislocation of elbow is a rare injury. Treatment options include open reduction internal fixation with K wire, replacement arthroplasty, excisional arthroplasty, arthrodesis, and hinged external fixator. The aim of this study is to determine the outcome of open reduction internal fixation with trans-olecranon K wire for neglected elbow dislocation. METHODS: This is a retrospective study done in three rural hospital of Karnali. Hospital records were reviewed from July 2015 to May 2018 to identify 11 cases who underwent open reduction internal fixation for neglected elbow dislocation. Pre and Postoperative outcome was assessed using range of motion and Mayo Elbow Performance Index. Data analysis was done using SPSS version 17. RESULTS: The average age of patient was 22.7 years (range 9-50 years). Non dominant hand was involved in 54.55%. The average preoperative elbow extension was 5.9 degree whereas postoperative extension was 15.9 degree. The average preoperative and postoperative elbow flexion was 24.5? and 113.6? respectively. Preoperative and postoperative Mayo elbow performance index was 18.6 and 86.3 respectively. Outcome was excellent in four patients, good in five patients and fair in two patients with one case having superficial infection. CONCLUSIONS: Open reduction and internal fixation with trans-olecranon k wire is an effective treatment method for neglected elbow dislocation. Postoperatively, elbow function is better with minimal complications.

Epidemiology of Femur Fractures in Children: A Descriptive Cross Sectional Study Based on a Rural Population of Nepal.

INTRODUCTION: Femur fracture in children is one of the most common lower limb fractures which require inpatient care. The aim of this study is to determine the epidemiology of femoral shaft fractures in children from a rural population of Karnali Nepal. METHODS: Hospital records were retrospectively reviewed from May 2017 to April 2020 to identify all the children with femur fracture. Sociodemographic profile, mode of injury, fracture pattern and location, time of presentation, initial treatment by traditional bone setters, treatment method and duration of hospital stay were noted. Data analysis was done using Statistical Package for Social Sciences version 20. RESULTS: Altogether 104 children were identified. The mean age was 5.55 years and boys predominated 65 (62.5%). Falls were the major mode of injury in 65 (62.4%) patients. Fractures were frequently noted between April 15 to August 15. There were four (3.8%) open fractures and concurrent fracture observed in eight (7.6%) patients. Ten (9.6%) children received prior treatment from traditional bonesetters. Treatment methods included hip spica 62 (59.6%), elastic intramedullary nailing 30 (28.8%) and plate fixation 12 (11.5%). The duration of hospital stay in the nailing and plate fixation group was 11.43 days and 18 days respectively. CONCLUSIONS: Fracture was common in 2-6 years of age group in boys during summer. Fall from cliff, rooftop and ladder were the major preventable cause of fracture. Delayed presentation and prior treatment with traditional bone setters add special challenges to orthopedic surgeons working in rural teaching hospital.

Pattern of Pediatric Supracondylar Fracture Operated at A Rural Teaching Hospital of Nepal: A Descriptive Cross-sectional Study.

INTRODUCTION: Supracondylar fracture of humerus is one of the common pediatric fractures encountered in our daily clinical practice. The purpose of this study is to determine the pattern of supracondylar fracture operated at rural teaching hospital of Jumla, Karnali Nepal. METHODS: A descriptive cross sectional study was conducted at Jumla, Karnali after Institutional Review Committee approval. Operating room notes from 15 May 2017 to 16 November 2019 were retrieved to gather the following information: patients address, age, sex, side, injury mechanism, displacement, neurovascular injury, concurrent injuries, initial management by traditional bone setters, time between injury and surgery, operative technique. Data analysis was done using Statistical Package for Social Sciences version 20. RESULTS: Left side predominated with 88 (63.7%) and extension type was common in 135 (97.8%). Thirteen (9.4%) patients were initially managed by traditional bonesetters. A total of 138 children underwent operative fixation with mean age of 7.47 years and gender ratio of 2:1 boy to girl. Fall from cliff, ladders and rooftops were the prevailing cause of injury 73 (52.8%). Average time between injury and surgery was 5.2 days. Closed reduction was done in 100 (72.4%) patients whereas open reduction was necessary in 38 (27.5%) patients. CONCLUSIONS: Closed extension type pediatric supracondylar fracture was common in this study. Fall from cliff, rooftop and ladder are the major cause of fracture. Delayed presentation and initial management of the fracture by the traditional bonesetters makes supracondylar fracture more challenging in resource limited setting like ours.

Medial versus Posterior Triceps Splitting Approach in Open Reduction Internal Fixation of Displaced Supracondylar Fracture of Humerus.

BACKGROUND: The purpose of this study is to compare the clinical and radiological outcome of medial versus posterior triceps splitting approach in open reduction internal fixation of displaced supracondylar fracture of humerus in children. METHODS: A retrospective review of total 70 children with medial approach (n=30) and posterior triceps splitting approach (n=40) children was made. Time to radiological union, postoperative Bauman's angle and Lateral humerocapitellar angle was compared for radiological outcome. Cosmetic and functional outcome was assessed and compared using Flynn's criteria. RESULTS: Radiological parameters like time to union, postoperative Bauman's angle and Lateral humerocapitellar angle were similar and non-significant between medial and posterior triceps splitting approach. Flynn's cosmetic outcome was similar between these approach (p=0.198). Loss of ROM was significant in posterior triceps splitting approach (p=0.00). Flynn's functional outcome was better with medial approach as compared to posterior triceps splitting approach with statistical significance (p=0.00). CONCLUSIONS: Medial approach has better functional outcome compared to posterior triceps splitting approach in open reduction internal fixation of displaced supracondylar fracture of humerus in children.

Orthopaedics Practice in a Rural Hospital During the COVID-19 Pandemic: A Descriptive Cross-Sectional Study.

INTRODUCTION: The coronavirus disease 2019  pandemic is one of the biggest public health challenges that we have faced, and has had a significant impact on the delivery of essential healthcare worldwide. Trauma and orthopaedic surgery was one of the most severely affected specialties as all non-emergency surgical cases were cancelled. With the possibility of future peak of corona virus, knowing the impact of Covid on orthopaedics would enable us to manage trauma patient in an effective manner and will help us to resume normal schedule of the trauma care. METHODS: This is a descriptive cross-sectional study. We studied the details of total number of patient at Orthopaedic and trauma outpatient, inpatient and operation theatre of Karnali Academy of Health Sciences from 14th March 2020 to 16 September 2020 as compared with the patient data from the same time period in 2019. RESULTS: There were 2288 patients during the non-Covid period who visited to the orthopaedic outpatient in comparison to 1618 during Covid period. Only 5 implant removals were performed in comparison to 50 removals (90% reduction) in elective cases. But the number of surgical cases was almost similar (204 vs 207 respectively). Total number of hospital stay had significantly reduced in the non-COVID period (average 4 days vs 6 days).There were 181 in-patient admissions during the COVID period while 241 admission during the non-COVID period. CONCLUSIONS: There is reduction in number of orthopaedic and trauma cases during COVID 19 with marked reduction in number of elective cases. Alternative methods of treatment like telemedicine and small scale health camps at peripheral districts should be conducted to treat non-urgent orthopaedic conditions during lockdown.

Early Versus Late Removal of Internally Fixated Kirschner's Wires for Displaced Lateral Condyle Fracture of Humerus in Children.

INTRODUCTION: Lateral condyle fracture of the distal humerus is the second most common paediatric elbow fracture. Unstable, rotated and displaced (>2 mm) fractures are managed with open reduction and internal fixation with Kirschner's wires or screws. Debate persists as for how long the Kirschner's wires should be placed in situ after internal fixation. We aimed to compare the functional and radiological outcome after early versus late removal of internally fixated Kirschner's wires for displaced lateral condyle fracture of distal humerus. METHODS: Children that underwent early (3-4 weeks) or late (5-7 weeks) removal of Kirschner's wire after open reduction and internal fixation for displaced lateral condyle fracture of humerus were observed for a period of minimum 6 months. Time to radiological union, carrying angle, range of motion was assessed and compared between early and late group. Functional outcome was compared using the Dhillon scoring system. RESULTS: We report the outcome of 40 cases (20 cases in each early and late group). Radiological union was achieved in all the cases of both group at 12 weeks follow up. The mean loss of carrying angle was statistically insignificant (p = 0.394) between the early and late group. There was no significant difference between the early and late group in relation to arc of motion at 12 weeks (p=0.724) and 6 months (p=0.638) follow up. Using the Dhillon scoring system, there was 100% excellent Dhillon score in early group, 80% excellent and 20% good Dhillon score in late group. Functional outcome was statistically insignificant between the two groups (p = 0.106). CONCLUSION: Early removal of internally fixated K-wires for displaced lateral condyle fracture of humerus in children showed similar radiological and functional results to late removal.